# CB2 receptor in the CNS: From immune and neuronal modulation to behavior

Wanda Grabon, Sylvain Rheims, Jonathon Smith, Jacques Bodennec, Amor

Belmeguenai, Laurent Bezin

► To cite this version:

Wanda Grabon, Sylvain Rheims, Jonathon Smith, Jacques Bodennec, Amor Belmeguenai, et al. CB2 receptor in the CNS: From immune and neuronal modulation to behavior. Neuroscience and Biobehavioral Reviews, 2023, 150, pp.105226. 10.1016/j.neubiorev.2023.105226. hal-04129849

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Distributed under a Creative Commons Attribution 4.0 International License
CB2 receptor in the CNS:
From immune and neuronal modulation to behavior

Wanda Grabon¹,²*, Sylvain Rheims¹,²,³, Jonathon Smith¹,², Jacques Bodennec¹,², Amor Belmeguenai¹,², Laurent Bezin¹,²*

¹ Université Claude Bernard Lyon 1, CNRS, Inserm, Centre de Recherche en Neurosciences de Lyon, U10208 UMR5292, TIGER Team – F-69500 Bron, France

² Epilepsy Institute IDEE, 59 boulevard Pinel - F-69500 Bron, France

³ Department of Functional Neurology and Epileptology, Hospices Civils de Lyon - France

AB, LB: contributed equally.

PMID: 37164044, DOI: 10.1016/j.neubiorev.2023.105226

Abstract

Despite low levels of cannabinoid receptor type 2 (CB2R) expression in the central nervous system in human and rodents, a growing body of evidence shows CB2R involvement in many processes at the behavioral level, through both immune and neuronal modulations. Recent *in vitro* and *in vivo* evidence have highlighted the complex role of CB2R under physiological and inflammatory conditions. Under neuroinflammatory states, its activation seems to protect the brain and its functions, making it a promising target in a wide range of neurological disorders. Here, we provide a complete and updated overview of CB2R function in the central nervous system of rodents, spanning from modulation of immune function in microglia but also in other cell types, to behavior and neuronal activity, in both physiological and neuroinflammatory contexts.

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**Keywords** : Endocannabinoid system, cannabinoid receptor 2, neuroinflammation, neuronal activity, behavior.

**1. Introduction**

Although many cannabinoid receptors have been and continue to be described (Cristino et al., 2020), the roles of the cannabinoid receptors type 1 (CB1R) and type 2 (CB2R) within the central nervous system (CNS) are most documented (Galiègue et al., 1995; Iannotti et al., 2016). CB1R is the most prevalent G protein-coupled receptor (GPCR) in the CNS and is expressed extensively by most neuron types. Itis often referred to as the central cannabinoid receptor and is the major mediator of the psychoactive effects of *Cannabis sativa* and its derivatives (Mackie, 2005). CB2R was first described as a peripheral receptor involved in

immunomodulatory functions in leukocytes (Buckley et al., 2000). CB2R is a GPCR mainly coupled to G₁/o proteins. Its stimulation triggers inhibition of adenylate cyclase activity and activation of the mitogen-activated protein kinase (MAPK), and is followed by β-arrestin recruitment resulting in receptor desensitization (Bouaboula et al., 1999; Kibret et al., 2022). CB2R intracellular pathways have been recently reviewed (Iannotti et al., 2016) and may differ depending on the tissues, cell types and subcellular localization (Howlett and Abood, 2017). The main intracellular pathways regulated by CB2R are summarized in figure 1.

* Corresponding authors at: 59 boulevard Pinel, F-69500 Bron, France.
E-mail addresses: laurent.bezin@univ-lyon1.fr (L. Bezin), wanda.grabon@univ-lyon1.fr (W. Grabon)

ORCID :

Wanda Grabon - https://orcid.org/0000-0003-4443-476X

Sylvain Rheims - https://orcid.org/0000-0002-4663-8515

Amor Belmeguenai - https://orcid.org/0000-0003-3295-3095

Laurent Bezin - https://orcid.org/0000-0001-6411-4267
Nowadays, due to the emergence of higher sensitivity and specificity techniques, CB2R has been detected in the CNS and with its activity modulated using specific agonists and inverse agonists, we have gained a greater understanding of its involvement in cell functions and behavior (Whiting et al., 2022). Since then, CB2R has received increasing interest in neuroscience research, with targeting of CB2Rs demonstrating great therapeutic promise in many rodent models of neurological diseases. Over the past decades, its involvement in the control of microglial activity and behavior has been widely investigated, as recently reviewed (Jordan and Xi, 2019; Komorowska-Müller and Schmöle, 2020). However, this modulation of behavior is perhaps even broader than previously described, with direct or indirect intervention of other cell types involved, including neurons and infiltrating immune cells in pathological contexts. This review aims to provide a detailed and updated overview of CB2R function in the CNS, spanning from the modulation of immune function in microglia and other cell types, to neuronal activity and the resulting behavior, in both physiological and neuroinflammatory contexts. The specific details of experimental designs, such as; the doses of CB2R activators or inhibitors used, the frequency and the time of treatments (i.e., before, and/or during and/or after the brain injury), are provided in tables. This is to aide in comparisons between studies, to find the elements that may explain results sometimes presented as contradictory, and to guide future experimental design.

## 2. Tools to tackle CB2R functions

CB2R can be activated by endogenous cannabinoids (endocannabinoids, among which the most extensively studied are anandamide and 2-arachidonylglycerol) or exogenous cannabinoids (phytocannabinoids derived from the *Cannabis sativa* plant, such as Δ⁹-tetrahydrocannabinol or THC and cannabidiol or CBD, figure 1) (Peng et al., 2022; Pertwee, 2008; Schurman et al., 2019). These natural molecules are known to mediate a plethora of physiological effects by activating many distinct receptors. The main cannabinoid receptors are CB1R and CB2R, yet cannabinoids have been reported to interact with several other receptors, ranging from other GPCRs to ion channels and nuclear receptors (Howlett, 2002; Starowicz et al., 2007; Zou and Kumar, 2018). As these molecules identified so far are not selective for CB2R (Cristino et al., 2020), studies investigating their effects on the central nervous system have not been reported in this review. The availability of specific synthetic ligands to CB2R (agonists and inverse agonists) and CB2R Knock-Out (KO) mice (Buckley et al., 2000; López et al., 2018; Zhang et al., 2019) has allowed the study of the biological function of CB2R both *in vitro* and *in vivo*. The current state of CB2R ligand development and progress has been recently reviewed in depth (Whiting et al., 2022). In the present review are included studies that have demonstrated functional implication of CB2R using CB2R KO mice or specific ligands. Also are included studies that used non-selective

agonists, the specific effect of which on CB2R was demonstrated when counteracted by specific inverse agonists/antagonists. Table 1 lists the CB2R ligands used in studies included in this review. Among them, JWH-133 is the most commonly used agonist, yet GP1a currently appears to present the lowest $K_i$ and therefore the highest affinity to CB2R (Murineddu et al., 2006). AM630 is by far the most represented ligand among all the inverse agonists listed in this review, even with JTE-907 having the highest affinity to CB2R (Iwamura et al., 2001).

## 3. Neuro-immunomodulatory properties of CB2R

### 3.1. *In vitro* evidence of CB2R immunomodulatory functions

CB2R is known for having strong immunomodulatory properties widely demonstrated in peripheral leukocytes. In the CNS, CB2R expression can be induced in microglia under inflammatory conditions (Benito et al., 2003; Grabon et al., 2023; Komorowska-Müller and Schmöle, 2020; Maresz et al., 2005). CB2R activation by specific agonists has been widely studied *in vitro* under inflammatory conditions in brain cells, especially in primary cultured microglia activated by bacterial lipopolysaccharide (LPS).

#### 3.1.1. Effect on microglial reactivity and polarization

Activation of CB2R with different specific agonists was tested under inflammatory conditions on human, mouse and rat microglial cell lines. The results obtained are reported in Supplementary Table 1, having paid particular attention for each study to the time of CB2R ligand application, i.e., before, during, or following the inflammatory challenge. The results of all the reported studies converge in favor of a reduction in both the expression of pro-inflammatory cytokines and of inducible nitric oxide synthase (iNOS), as well as reduced levels of oxidative molecules related to the M1 polarization state, regardless of when the CB2R agonist was applied (Askari and Shafiee-Nick, 2019a; Ehrhart et al., 2005; Facchinetti et al., 2003; Ma et al., 2015; Malek et al., 2015; Ortega-Gutiérrez et al., 2005; Pan et al., 2020; Ramírez et al., 2005; Ramirez et al., 2012; Ribeiro et al., 2013; Romero-Sandoval et al., 2009; Young and Denovan-Wright, 2022). Activation of downstream pro-inflammatory pathways were also restrained by CB2R stimulation (Sheng et al., 2019; Tang et al., 2015). The expression of M2-related proteins, especially interleukin (IL)-10, arginase (Arg)-1 and cluster of differentiation (CD)206, as well as anti-inflammatory cytokines were found to be increased following CB2R specific activation (Askari and Shafiee-Nick, 2019a; Correa et al., 2010; Ma et al., 2015; Malek et al., 2015). Furthermore, microglia from CB2R deficient mice are not able to polarize toward the anti-inflammatory and cytoprotective M2 phenotype, as they showed a reduced expression of Arg-1 after stimulation with IL-4/IL-13 (Mecha et al., 2015).
<figure><img src="image_3.png" /><figcaption>**Figure 1. CB2R cellular signaling.** The different types of CB2R ligands are represented: phytocannabinoids, endocannabinoids, and synthetic ligands. The major intracellular pathways modulated by the G-protein coupled receptor CB2R are shown in the lower part of the figure. It should be noted that the pathways activated may depend on cell types, ligands and physiological conditions. Figure created with BioRender.com.
**Abbreviations:** Δ<sup>9</sup>-THC, Δ<sup>9</sup>-tetrahydrocannabinol; 2-AG, 2-arachidonyl-glycerol; ATP, adenosine triphosphate; cAMP, cyclic adenosine monophosphate; CBD, cannabidiol; ERK, Extracellular signal-regulated kinases; JNK, c-Jun N-terminal kinase; MAPK, mitogen-activated protein kinases.</figcaption></figure>

Taken together, these results suggest that CB2R activation plays a major role in resolving microglia-driven inflammation, promoting a switch from M1 to M2 polarization state, even if the distinction between these two states is now considered as caricatural and should be used cautiously (Paolicelli et al., 2022; Ransohoff, 2016).

Some studies have shown intriguing results comparing the effects of CB2R agonists and inverse agonists. In particular, it was shown that CD206 expression was decreased after CB2R activation, whereas it was increased by the specific inverse agonists SMM-189 and SR14428 (Presley et al., 2015). In addition, conflicting conclusions can be drawn when comparing studies that use different CB2R inverse agonists on different cell types and in response to distinct inflammatory challenges. For instance, inverse agonist applied before LPS induced either an increased expression of IL-1β, IL-18 and Tumor Necrosis Factor (TNF)-α with AM630 (Malek et al., 2015), or conversely a reduced release of IL-1β and prostaglandin E2 when using SMM-189 (Yu et al., 2020).

All the data collected so far highlights the full potential of CB2R modulation to resolve inflammation and alter microglial cell function, although our understanding of the mechanisms underlying this modulation remains largely limited due to their complexity.

### 3.1.2. Effect on other cell types

Production and secretion of cytokines, by monocytes, astrocytes, oligodendrocytes and endothelial cells, following inflammatory challenge were counteracted when stimulated by CB2R agonists (Askari and Shafiee-Nick, 2019b; Correa et al., 2005; Jia et al., 2020; Klegeris et al., 2003; Montecucco et al., 2008; Persidsky et al., 2015; Sheng et al., 2005). In addition, their release of chemokines, involved in the chemoattraction of immune cells toward the inflammatory site, was decreased following CB2R activation (Jia et al., 2020; Persidsky et al., 2015; Sheng et al., 2005) (Supplementary Table 1). Transwell migration assays demonstrated that migration of neutrophils (Murikinati et al., 2010; Porter et al., 2019) and macrophages (Montecucco et al., 2008; Persidsky et al., 2015; Rom et al., 2013) was inhibited when using specific CB2R agonists. Furthermore, a reduction in monocyte/endothelium interaction was demonstrated after CB2R activation in human primary brain microvascular endothelial cells challenged with TNF-α (Rom et al., 2013), concurrent with a decrease in adhesion molecules such as intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 expression (Persidsky et al., 2015; Ramirez et al., 2012). Hence, CB2R can exert its immunomodulatory role by decreasing the recruitment of peripheral leukocytes into inflamed neuronal tissue through limiting both leukocyte chemotaxis and extravasation.
<table><caption>**Table 1. CB2R Ligands.** Every CB2R ligands used in studies included in this review are listed in this table. Note that AM630, JTE-907, SMM-189, SR144528 have been first described as inverse agonists, as mentioned here, but have also been mentioned as antagonists in many studies. In the "frequency of use" column is presented the number of times each ligand was used among all included studies (studies having used one or more agonists to activate CB2R: n=155; studies having used one or more inverse agonists to study their own effect or to block the effect of an agonist: n=100)</caption><thead><tr><th>LIGAND</th><th>ACTION</th><th>IUPAC Nomenclature</th><th>Ki (nM)</th><th>CB2R/CB1R affinity</th><th>Frequency of use</th><th>REF</th></tr></thead><tbody><tr><td>AM1241</td><td>CB2R Agonist</td><td>(2-Iodo-5-nitrophenyl)-[1-[(1-methylpiperidin-2-yl)methyl]indol-3-yl]methanone</td><td>3.4</td><td>82:1</td><td>12/155</td><td>(Ibrahim et al., 2003)</td></tr><tr><td>β-caryophyllene</td><td>CB2R Agonist</td><td>(1R,4E,9S)-4,11,11-trimethyl-8-methylidenebicyclo[7.2.0]undec-4-ene</td><td>155</td><td>No affinity for CB1R</td><td>14/155</td><td>(Gertsch et al., 2008)</td></tr><tr><td>CB65</td><td>CB2R Agonist</td><td>N-Cyclohexyl-7-chloro-1-[2-(4-morpholinyl)ethyl]quinolin-4(1H)-one-3-carboxamide</td><td>3.3</td><td>303:1</td><td>1/155</td><td>(Manera et al., 2006)</td></tr><tr><td>GP1a</td><td>CB2R Agonist</td><td>N-(Piperidin-1-yl)-1-[2-(4-dichlorophenyl)-1,4-dihydro-6-methylindeno[1,2-c]pyrazole-3-carboxamide</td><td>0.037</td><td>>9000:1</td><td>10/155</td><td>(Murineddu et al., 2006)</td></tr><tr><td>GW833972A</td><td>CB2R Agonist</td><td>2-[(3-chlorophenyl)amino]-N-(4-pyridinylmethyl)-4-(trifluoromethyl)-5-Pyrimidinecarboxamide hydrochloride</td><td>7.8</td><td>1000:1</td><td>2/155</td><td>(Belvisi et al., 2008)</td></tr><tr><td>GW405833</td><td>CB2R Agonist</td><td>1-[2,3-Dichlorobenzoyl]-5-methoxy-2-methyl-3-[2-(4-morpholinyl)ethyl]-1H-indole</td><td>3.92</td><td>1217:1</td><td>4/155</td><td>(Gallant et al., 1996)</td></tr><tr><td>HU210</td><td>CB2R/CB1R Agonist</td><td>(6aR,10aR)-9-(hydroxymethyl)-6,6-dimethyl-3-[2-methyloctan-2-yl]-6H,6aH,7H,10H,10aH-benzo[c]isochromen-1-ol</td><td>0.52</td><td>1:10</td><td>3/155</td><td>(Felder et al., 1995)</td></tr><tr><td>HU308</td><td>CB2R Agonist</td><td>[(1R,2R,5R)-2-[2,6-Dimethoxy-4-(2-methyloctan-2-yl)phenyl]-7,7-dimethyl-4-bicyclo[3.1.1]hept-3-enyl]methanol</td><td>22.7</td><td>No affinity for CB1R</td><td>21/155</td><td>(Hanus et al., 1999)</td></tr><tr><td>HU910</td><td>CB2R Agonist</td><td>((1S,4R)-2-[2,6-dimethoxy-4-(2-methyloctan-2-yl)phenyl]-7,7-dimethylbicyclo[2.2.1]hept-2-en-1-yl)methanol</td><td>6</td><td>228:1</td><td>4/155</td><td>(Horváth et al., 2012)</td></tr><tr><td>HU914</td><td>CB2R Agonist</td><td>(1S,4R)-2-[2,6-dimethoxy-4-(2-methyloctan-2-yl)phenyl]-7,7-dimethylbicyclo[2.2.1] hept-2-ene-1-carboxylic acid</td><td>1500</td><td>No affinity for CB1R</td><td>1/155</td><td>(Magid et al., 2019)</td></tr><tr><td>JWH-015</td><td>CB2R Agonist</td><td>(2-Methyl-1-propyl-1H-indol-3-yl)-1-naphthalenylmethanone</td><td>13.8</td><td>28:1</td><td>18/155</td><td>(Showalter et al., 1996)</td></tr><tr><td>JWH-133</td><td>CB2R Agonist</td><td>(6aR,10aR)-3-[1,1-dimethylbutyl)-6a,7,10,10a-tetrahydro-6,6,9-trimethyl-6Hdibenzo[b,d]pyran</td><td>3.4</td><td>200:1</td><td>63/155</td><td>(Huffman et al., 1999)</td></tr><tr><td>MDA7</td><td>CB2R Agonist</td><td>1-[[(3-benzyl-3-methyl-2,3-dihydro-1-benzofuran-6-yl]carbonyl]piperidine</td><td>238</td><td>10:1</td><td>4/155</td><td>(Naguib et al., 2008)</td></tr><tr><td>O-1966</td><td>CB2R/CB1R Agonist</td><td>1-[4-(1,1-dimethyl-heptyl)-2,6-dimethoxy-phenyl]-3-methyl-cyclohexanol</td><td>23.0</td><td>220:1</td><td>9/155</td><td>(Wiley et al., 2002)</td></tr><tr><td>O-3853</td><td>CB2R Agonist</td><td>-</td><td>17.3</td><td>47:1</td><td>1/155</td><td>(Zhang et al., 2007)</td></tr><tr><td>Paeoniflorin</td><td>CB2R Agonist</td><td>[(1R,2S,3R,5R,6R,8S)-6-hydroxy-8-methyl-3-[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy-9,10-dioxatetracyclo[4.3.1.02,5.03,8]decan-2-yl]methyl benzoate</td><td>-</td><td>-</td><td>1/155</td><td>(Luo et al., 2018a)</td></tr><tr><td>RO6871304</td><td>CB2R Agonist</td><td>(3S)-1-[5-tert-butyl-3-[(1-cyclopropyl1H-tetrazol-5-yl)]methyl]-3H-[1,2,3]triazolo[4,5-d]pyrimidin-3-ol</td><td>34</td><td>588:1</td><td>1/155</td><td>(Porter et al., 2019)</td></tr><tr><td>RO6871085</td><td>CB2R Agonist</td><td>5-Cyclopropyl-N-[(2S)-1-cyclopropyl-2-(5-methyl-1,2,4-oxadiazol-3-yl)propan-2-yl]-4-(2,2,2-trifluoroethoxy)pyridine-2-carboxamide</td><td>6</td><td>44:1</td><td>1/155</td><td>(Porter et al., 2019)</td></tr><tr><td>UCM707</td><td>Inhibitor of anandamide uptake</td><td>(5Z,8Z,11Z,14Z)-N-(3-furylmethyl)eicosa-5,8,11,14-tetraenamide</td><td>-</td><td>-</td><td>1/155</td><td>(de Lago et al., 2002)</td></tr><tr><td>WIN55,212-2</td><td>CB2R/CB1R Agonist</td><td>(11R)-2-Methyl-11-[(morpholin-4-yl)methyl]-3-(naphthalene-1-carbonyl)-9-oxa-1-azatricyclo[6.3.1.0<sup>4,12</sup>]dodeca-2,4(12),5,7-tetraene</td><td>3.3</td><td>1:2</td><td>11/155</td><td>(Felder et al., 1995)</td></tr><tr><td>1-phenylisatin</td><td>CB2R Agonist</td><td>1-phenylindole-2,3-dione</td><td>-</td><td>-</td><td>2/155</td><td>(Jayant et al., 2016)</td></tr><tr><td>AM630</td><td>CB2R Inverse agonist</td><td>1-[2-(Morpholin-4-yl)ethyl]-2-methyl-3-(4-methoxybenzoyl)-6-iodoindole</td><td>31.2</td><td>165:1</td><td>67/100</td><td>(Ross et al., 1999)</td></tr><tr><td>JTE-907</td><td>CB2R Inverse agonist</td><td>N-(1,3-benzodioxol-5-ylmethyl)-1,2-dihydro-7-methoxy-2-oxo-8-(pentyloxy)-3-Quinolinecarboxamide</td><td>0.38</td><td>2760:1</td><td>3/100</td><td>(Iwamura et al., 2001)</td></tr><tr><td>SMM-189</td><td>CB2R Inverse agonist</td><td>30,50-dichloro-2,6-dihydroxy-biphenyl-4-yl)-phenyl-methanone</td><td>1.66</td><td>602:1</td><td>3/100</td><td>(Bhattacharjee et al., 2009)</td></tr><tr><td>SR144528</td><td>CB2R Inverse agonist</td><td>5-(4-Chloro-3-methylphenyl)-1-[4-(4-methylphenyl)methyl]-N-[(1S,2S,4R)-1,3,3-trimethylbicyclo[2.2.1]heptan-2-yl]-1H-pyrazole-3-carboxamide</td><td>0.6</td><td>700:1</td><td>32/100</td><td>(Rinaldi-Carmona et al., 1998)</td></tr></tbody></table>

**Abbreviations:** CB1R and CB2R, cannabinoid receptor type 1 and 2; IUPAC, International Union of Pure and Applied Chemistry; Ki, dissociation constant; Ref, reference first describing the compound.
## 3.2. Immunomodulatory properties of CB2R in rodent models of neuroinflammation

CB2R is upregulated in brain inflammatory tissue under different pathological contexts (Benito et al., 2008; Cabral et al., 2008; Grabon et al., 2023; Jordan and Xi, 2019). Numerous studies have provided evidence that CB2R is involved in the modulation of neuroinflammation, when triggered in various

animal models of diverse neurological disorders. Here multiple inflammatory hallmarks have been studied, namely the release of inflammatory cytokines, gliosis, cell polarization and the recruitment of peripheral leukocytes into the brain (Figure 2, Supplementary Table 2).

### 3.2.1. Inflammatory cytokine expression

The expression of pro-inflammatory cytokines in the neural tissue is one of the main features of neuroinflammation. Pro-inflammatory cytokines such as TNF-α, IL-1β, IL-6, and Interferon (IFN)-γ are strongly induced in many inflammatory-associated diseases or conditions affecting the nervous system, including Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), multiple sclerosis (MS), stroke, neuropathic pain, traumatic brain

injury (TBI), spinal cord injury, encephalitis, epilepsy, etc. In line with results obtained from *in vitro* investigations, the specific activation of CB2R was shown to significantly decrease the induction of pro-inflammatory cytokines in inflamed tissue in rodent models (Supplementary Table 2). In some studies using agonists with high CB2R affinity and selectivity, CB2R involvement has been ensured by the loss of effect with the use of inverse agonists (Chung et al., 2016; Javed et al., 2016; Li et al., 2018; Tang et al., 2015; Tao et al., 2016; Toguri et al., 2014; Torres et al., 2010; Viveros-Paredes et al., 2017a; Youssef et al., 2019) or by the lack of the expected effect in transgenic mice that do not express CB2R (CB2R KO) (Gómez-Gálvez et al., 2016; Porter et al., 2019).

This modulatory effect was observed in preventive strategies, when agonists were administered before induction of the disease or the injury, namely in rodent models of HD (Sagredo et al., 2009), LPS-induced inflammation (Sahu et al., 2019a), 3,4-methylenedioxy-N-methylamphetamine (MDMA)-induced inflammation (Torres et al., 2010), neonatal stroke (Tang et al., 2016), seizures (Karan et al., 2021) and PD (Chung et al., 2016; Javed et al., 2016).

<figure><img src="image_4.png" /><figcaption>**Figure 2. Summary of immunomodulatory effects of CB2R in the central nervous system.** This figure depicts the main outcomes on neuroinflammation following CB2R modulation described in section 3. The main effects of pharmacological activation of CB2R by treatment with specific agonists are summarized on the left side of the figure. The majority of the studies included in this review converge in favor of an anti-inflammatory effect of CB2R activation, with a reduction in the: 1) activation of both microglia and astrocytes, 2) expression of cytokines and other inflammatory molecules, 3) permeability of the blood-brain barrier (BBB), and 4) infiltration of leukocytes (neutrophils, T lymphocytes, monocytes depending on the context). The effects of blocking CB2R activity (genetically using KO mice or pharmacologically with specific antagonists) are shown on the right side of the figure. Blockade of CB2R mostly leads to pro-inflammatory effects, with exacerbation of microglia and astrocyte activation, inflammatory molecules expression, BBB permeability and peripheral immune cell infiltration. Figure created with BioRender.com</figcaption></figure>
When administered after triggering of the pathological condition in animal models, CB2R agonists attenuated overexpression of pro-inflammatory molecules, as shown in models of AD (Çakır et al., 2019; Fakhfouri et al., 2012; Wu et al., 2013), MS (Croxford and Miller, 2003; Kong et al., 2014a), uveitis (Toguri et al., 2014), vascular dementia (Luo et al., 2018a), stroke (Tang et al., 2015; Tao et al., 2016; Zarruk et al., 2012), sepsis (Çakır et al., 2020; Yang et al., 2022), intracerebral hemorrhage (Li et al., 2018; Lin et al., 2017), human immunodeficiency virus (HIV)-1 encephalitis (Gorantla et al., 2010), PD (Rentsch et al., 2020), TBI (Amenta et al., 2014; Braun et al., 2018; Magid et al., 2019), spinal cord injury (Adhikary et al., 2011), surgery-induced neuroinflammation (Sun et al., 2017), sub-chronic stress-induced inflammation (Zoppi et al., 2014), high fat diet (Youssef et al., 2019) and hypertension-induced neuroinflammation (Shi et al., 2020). Interestingly, peripheral treatment with inverse agonist SMM-189 was also shown to decrease the expression of pro-inflammatory cytokines induced by status epilepticus in mice (Yu et al., 2020), suggesting that both agonists and inverse-agonist may exert anti-inflammatory properties under certain conditions. Chronic administration of CB2R agonists in AD models after the onset on the neuroinflammatory state also resulted in a decrease in the expression of pro-inflammatory mediators (Aso et al., 2013; Martín-Moreno et al., 2012). In certain cases, the decrease in pro-inflammatory cytokines following CB2R activation was accompanied by an increase in the expression of anti-inflammatory cytokines, as observed in models of stroke (Tao et al., 2016), spinal cord injury (Adhikary et al., 2011) or TBI (Braun et al., 2018).

CB2R activation by agonists in the vast majority of cases, and sometimes by inverse agonists, can therefore exert powerful preventive and curative effects on the induction of pro-inflammatory cytokines.

However, as for the *in vitro* studies, the effects of CB2R inverse agonists used alone on inflammatory cytokine induction are more contrasted. Cytokine induction remained unchanged in a model of stroke (Tang et al., 2016), was upregulated in surgery-induced neuroinflammation (Sun et al., 2017), downregulated in a model of uveitis (Toguri et al., 2014) and inconsistent in TBI, being either unchanged (Braun et al., 2018) or upregulated (Amenta et al., 2014). These intriguing effects may be explained by the variety of the doses, time and modes of administration of the drugs, and by the time measurements were made.

### 3.2.2. Gliosis

#### 3.2.2.1. Microglia

Pre-treatment with CB2R agonists showed a significant reduction of microglial activation in inflamed tissue of animal models of chronic post-ischemia pain (Xu et al., 2016), MDMA-induced neuroinflammation (Torres et al., 2010), neonatal stroke (Tang et al., 2016), paclitaxel-induced neuro-

inflammation (Wu et al., 2019), Theiler's virus induced demyelination disease (Mecha et al., 2018), CO-induced cognitive deficit (Du et al., 2020), hemicerebellectomy (Oddi et al., 2012) and PD (Chung et al., 2016; Javed et al., 2016; Viveros-Paredes et al., 2017a). Activation of CB2R during or after induction also lead to a decrease in microgliosis in rodent models of high-fat diet induced-neuroinflammation (Rorato et al., 2022), morphine-hyperalgesia (Tumati et al., 2012), AD (Wu et al., 2013), HIV1-encephalitis (Gorantla et al., 2010), intracerebral hemorrhage (Li et al., 2018), MS (Alberti et al., 2017; Kong et al., 2014b, 2014a; Ortega-Gutiérrez et al., 2005), PD (Gómez-Gálvez et al., 2016; Rentsch et al., 2020), spinal cord injury (Adhikary et al., 2011), neuropathic pain (Klauke et al., 2014), stroke (Tang et al., 2015; Tao et al., 2016; Yu et al., 2015; Zarruk et al., 2012), surgery-induced neuroinflammation (Sun et al., 2017), TBI (Elliott et al., 2011), vascular dementia (Luo et al., 2018a) and retinal inflammation (Spyridakos et al., 2021). Finally, chronic activation of CB2R in AD (Aso et al., 2013; Li et al., 2019a; Martín-Moreno et al., 2012; Wu et al., 2017) and in amyotrophic lateral sclerosis (ALS) (Espejo-Porras et al., 2019) mouse also resulted in a decrease in microglial activation. Microglial cells sensitivity to CB2R activation under inflammatory state is consistent with *in vitro* observations and with the fact that CB2R expression was shown to be induced in this cell population in such conditions (Grabon et al., 2023; Komorowska-Müller and Schmöle, 2020).

Given that microglia are considered as one of the major contributors of inflammatory cytokine secretion under pathological conditions, the limitation of microglial activation may be at least one explanatory factor of the decrease in pro-inflammatory cytokine induction observed following CB2R activation.

#### 3.2.2.2. Astrocytes

In most studies reporting the effect of CB2R activation on astrogliosis, glial fibrillary acid protein (GFAP) expression was found to be either unchanged, as in a HIV-encephalitis (Gorantla et al., 2010) and AD (Martín-Moreno et al., 2012) models (Supplementary Table 2) or downregulated in ALS (Espejo-Porras et al., 2019), PD (Chung et al., 2016; Javed et al., 2016; Rentsch et al., 2020; Viveros-Paredes et al., 2017a), spinal cord ischemia (Jing et al., 2020), neuropathic pain (Klauke et al., 2014), morphine-mediated inflammation (Tumati et al., 2012), and in a different AD model (Wu et al., 2013). Unexpectedly, astrogliosis was found to be exacerbated following CB2R agonist administration and reduced with an inverse agonist in another AD mouse model (Esposito et al., 2007). CB2R activation appears to have potent opposite effects on astrogliosis depending on the animal model used, on the time-window of treatment and on the time of data collection, due to astrogliosis being a dynamic process and to CB2R having complex immunomodulatory properties. CB2R expression has been poorly investigated in astrocytes. It was detected in GFAP-positive cells in the
substansia nigra of PD patients (Navarrete et al., 2018) and in the striatum in a HD rat model (Sagredo et al., 2009), but the specificity of the antibodies used in these studies has not been validated. In vitro, CB2R mRNA was detected at low level in enriched primary cultured astrocytes (Carlisle et al., 2002). Nevertheless, CB2R activation may also have indirect effects on astrocytes through microglia for example.

### 3.2.3. Leukocyte brain invasion

#### 3.2.3.1. Peripheral leukocyte detection in the CNS

The number of infiltrating immune cells in inflamed neural tissue was found to be downregulated by both pre- and post-treatment with CB2R agonist. Namely, T-cells, macrophages and/or neutrophils were found to be fewer in rodent models of PD within the substansia nigra (Chung et al., 2016) and the striatum (Gómez-Gálvez et al., 2016), neuropathic pain within the spinal cord (Sheng et al., 2019), MS within the spinal cord (Kong et al., 2014b, 2014a), stroke within the ischemic cortex (Yu et al., 2015), TBI within the cortex (Braun et al., 2018), in injured spinal cord (although not significant) (Adhikary et al., 2011), and uveitis in eyes (Toguri et al., 2018).

#### 3.2.3.2. Chemotaxis (chemokine expression)

The induction of chemokines was shown to be decreased by CB2R activation in rodent models of PD, MS, spinal cord injury, stroke, surgery-induced neuroinflammation, TBI and uveitis (Supplementary Table 2). However, treatment with AM630, a CB2R inverse agonist, resulted in more contrasted outcomes as chemokine expression was either increased following a surgery-induced cognitive impairment model (Sun et al., 2017), unchanged in a TBI model (Braun et al., 2018) or decreased in an uveitis model (Toguri et al., 2014).

#### 3.2.3.3. Blood brain barrier (BBB) permeability

The migration of peripheral leukocytes into the CNS is made possible under neuroinflammatory conditions by the BBB leakage due to the induction of metalloproteases such as MM9 and the decrease in tight junction protein expression, such as zonula-occludens-1 or claudin-5. This permeability can be measured with different methodological approaches, namely via tracing labeled-proteins normally unable to cross the BBB under physiological conditions. The BBB permeability induced by neuroinflammatory conditions was found to be decreased by CB2R activation in rodent models of PD (Chung et al., 2016), spinal cord ischemia (Jing et al., 2020), LPS-induced inflammation (Ramirez et al., 2012), intracerebral hemorrhage (Li et al., 2018), and TBI (Supplementary Table 2).

#### 3.2.3.4. Leukocyte vessel adhesion

Following CB2R activation in cultured endothelial cells from human brain, there is a reduced expression of specific adhesion molecules (VCAM, ICAM), that play an essential role in the interaction between leukocytes and endothelial cells,

together with a reduced BBB permeability (Ramirez et al., 2012). Vessel adhesion of leukocytes has also been assessed *in vivo* following CB2R activation by intravital fluomicroscopy and revealed reduced leukocyte/endothelium interactions in a focal ischemia/reperfusion model (Supplementary Table 2).

Overall, these results argue that CB2R can modulate neuroinflammation by controlling peripheral leukocytes invasion in the nervous tissue.

### 3.2.4. Cell polarization

In agreement with the *in vitro* data, *in vivo* results obtained in rodent models of AD (Li et al., 2019a), intracerebral hemorrhage (Lin et al., 2017), TBI (Li et al., 2022), vascular dementia (Luo et al., 2018a) and stroke (Tao et al., 2016) show that CB2R activation promoted the change of microglial cell polarization from the proinflammatory M1 to the anti-inflammatory M2 phenotype. In a study that distinguished macrophages from microglial cells, CB2R activation caused cortex-infiltrating macrophages to display M2 phenotypic features in a TBI mouse model (Braun et al., 2018).

Hence, CB2R may act as a regulator of neuroinflammation not only by modulating activation of resident glial cells but also by controlling the number of peripheral leukocytes migrating to the CNS and their inflammatory state (Figure 2).

### 3.3. Insight from CB2R KO mice

CB2R's immunomodulatory role was demonstrated in models of MS, TBI and subchronic stress-induced neuroinflammation, that its deletion resulted in an increase in pro-inflammatory cytokine expression compared with wild-type (WT) littermates (Supplementary Table 3). Production of iNOS was also 10-fold higher in CB2R KO mice subjected to TBI (Amenta et al., 2014). Gliosis was exacerbated in hypoxia-ischemia (Kossatz et al., 2016), ALS (Rodríguez-Cueto et al., 2021), age-related cognitive-decline (Komorowska-Müller et al., 2021a), isoflurane-induced inflammation (Li et al., 2021), neuropathic pain (Racz et al., 2008), α-synuclein fibril-induced inflammation (Feng et al., 2022), MS (Shao et al., 2014) and PD (Gómez-Gálvez et al., 2016) models established in CB2R KO mice. Conversely, gliosis was reduced in a model of LPS and IFN-γ-induced inflammation (Reusch et al., 2022). BBB permeability was found to be enhanced in a TBI model (Amenta et al., 2014), alongside elevated endothelium/leukocyte interactions (Porter et al., 2019; Toguri et al., 2018). Adhesion of leukocytes to endothelium was not increased in LPS-induced neuroinflammation in CB2R KO mouse, though the complete resolution of inflammation took longer (Ramirez et al., 2012). Inconsistent results were obtained from genetic models of AD established in different CB2R KO mouse strains. Here, an exacerbation of the inflammatory state could have been expected as in other inflammatory models. In fact, CB2R deletion either resulted in unchanged IL-1β expression and microglial activation (Koppel et al., 2014; López et al., 2018) or it resulted in a
decrease in gliosis (Ruiz de Martín Esteban et al., 2022; Schmöle et al., 2018), pro-inflammatory cytokine and chemokine expression and macrophage infiltration (Schmöle et al., 2015) depending on AD mouse model used. These conflicting data may be due to the genetic etiology of the various AD models and the use of different CB2R KO mouse strains.

## 4. Neuronal activity modulation by CB2R

CB1R is highly expressed in the CNS at the neuronal level and functionally modulates presynaptic neurotransmitter release (Araque et al., 2017; Howlett, 1998). In stark contrast to the vast amount of literature on CB1R-mediated neuromodulation, very little is known about the relevance of CB2R in neuronal signaling. While the neuronal expression of CB2R has been a matter of debate over the past decade, accumulating evidence supports that CB2R plays a role in neuronal activity modulation. Here, we report evidence emerging from both *in vitro* (Supplementary Table 4) and *in vivo* (Supplementary Table 5) supporting CB2R involvement in the modulation of neuronal activity in various brain regions and neuronal subtypes (Figure 3).

### 4.1. Neurotransmitter release

The impact of CB2R on both excitatory and inhibitory neurotransmitter release has been studied *in vitro*. Bath application of specific agonists resulted in a reduction of evoked gamma-aminobutyric acid (GABA) release (Andó et al., 2012), capsaicin induced calcitonin gene-related peptide (CGRP) release (Beltramo et al., 2006) and K+-stimulated Glutamate (Sánchez-Zavaleta et al., 2018) and dopamine release (López-Ramírez et al., 2020) (Supplementary Table 4). Microdialysis studies have revealed additional insights into the effects of CB2R signaling on neurotransmitter release *in vivo*. The effect of CB2R specific activation has been particularly investigated on dopamine release in the nucleus accumbens of rodents. Under physiological conditions, CB2R local activation resulted in a significative drop in extracellular dopamine in a dose-dependent manner. This effect was blocked by an inverse agonist administration and was absent in CB2R KO mice (Xi et al., 2011). Likewise, CB2R pharmacological activation dose-dependently reduced the cocaine- and morphine-mediated rises of extracellular levels of dopamine (Grenald et al., 2017; Xi et al., 2011). However, the extracellular dopamine level in the nucleus accumbens of healthy rats remained unchanged when challenged with the same agonist in a different study, likely because of the systemic route of administration (Zhang et al., 2017). In a rodent model of PD, CB2R activation led to an increase in dopamine and serotonin levels in the striatum (Shi et al., 2017). In the hippocampus, local infusion of JWH-133 reduced ischemia-enhanced extracellular glutamate (Zheng et al., 2015). Taken together, these studies provide further evidence that CB2R activation acts in multiple brain regions to modulate neurotransmitter release.

### 4.2. Ionic currents

Several lines of electrophysiological evidence indicate that activation of CB2R can modify neuronal activity through ionic current modulation. *In vitro*, JWH-133 bath application on medial prefrontal cortex (mPFC) slices dose-dependently evoked Cl⁻ currents as measured by whole-cell current and voltage clamp recordings (den Boon et al., 2014) and increased intracellular calcium levels (den Boon et al., 2012). T-type Ca²⁺ current amplitude was shown to be increased by CB2R activation in isolated retinal ganglion cells (Qian et al., 2017), as well as M-type K⁺ current amplitude in dopaminergic neurons of VTA slices (Ma et al., 2019). These effects were both prevented by AM630 application (Supplementary Table 4). *In vivo*, calcium imaging revealed that local JWH-133 administration in the spinal cord attenuated capsaicin evoked intracellular calcium increase in a spinal nerve ligation rat model (Sagar et al., 2005).

### 4.3. Membrane potential

Neuronal membrane potential was shown to be differentially shifted by CB2R activation in a cell type-specific and a region-specific manner. CB2R agonist bath application resulted in a transient delayed depolarization in layer II/III pyramidal neurons of rat mPFC slices and had no effect in CB2R KO mice (den Boon et al., 2012), whereas CB2R specific activation hyperpolarized dopaminergic neurons isolated from mouse VTA (Zhang et al., 2017, 2014). HU-308 was also shown to induce a hyperpolarization in CA3 pyramidal cells of mouse hippocampus *in vivo*. This effect of CB2R activation was directly mediated by neurons as the membrane potential remained unchanged in mice with conditional KO of CB2R in synapsin-positive neurons (Syn-CB2R cKO mice) (Stempel et al., 2016), providing evidence that functional CB2Rs are indeed expressed by neurons in the hippocampus.

### 4.4. Neuronal excitability

Further *in vitro* and *in vivo* electrophysiological studies brought insights on CB2R involvement in neuronal excitability, namely by assessing neuronal spontaneous or evoked action potentials (AP) firing rate and duration, or by measuring the afterhyperpolarization (AHP). Bath application of CB2R specific agonists on mPFC, VTA, midbrain or substantia nigra pars compacta (SNC) slices resulted in a decreased excitability (den Boon et al., 2012; Ma et al., 2019; Yu et al., 2021; Zhang et al., 2021, 2017, 2014) of pyramidal and dopaminergic neurons, respectively. Conversely, inverse agonist application raised pyramidal neurons excitability in mPFC slices. Neither the agonist nor the inverse agonist had any effect on the excitability of neurons from CB2R KO mice (den Boon et al., 2012). *In vivo*, neuronal excitability was also shown to be decreased by CB2R activation in thalamic ventral posterior nucleus neurons (Jhaveri et al., 2008), in spinal wide dynamic range neurons (Elmes et al., 2004; Nackley et al., 2004), in dorsal horn neurons (Sokal et al., 2003), in CA3
<figure><img src="image_5.png" /><figcaption>**Figure 3. Neuronal activity outcomes following CB2R activation or blockade.** This figure summarizes the main mechanisms involved in the modulation of neuronal activity that can be modulated following CB2R activation or blockade, as described in section 4. It should be noted that these modulations were each demonstrated in specific neuronal subpopulations (supplementary table 4 and 5). Figure created with *BioRender.com*</figcaption></figure>

pyramidal cells (Stempel et al., 2016) and in VTA dopaminergic neurons (Zhang et al., 2014).

### 4.5. Synaptic transmission

The effect of CB2R activation has also been studied at the synaptic level in vitro by monitoring spontaneous, miniature, or evoked post-synaptic currents (PSC). In the hippocampus, activation of CB2R by specific agonists resulted in a higher miniature excitatory PSC (mEPSC) frequency without impacting their amplitude nor inhibitory currents. CB2R-mediated hyperpolarization observed in CA3 did not result from an alteration of evoked IPSC amplitude or field excitatory post-synaptic potential, suggesting that it would be a solely self-regulatory cell-intrinsic mechanism (Stempel et al., 2016). CB2R activation in hippocampus slices also resulted in higher field EPSC (Hwang et al., 2020). In entorhinal cortex and cerebellum slices, amplitude of spontaneous inhibitory PSC recorded in Layer II-V neurons (Morgan et al., 2009) and Purkinje cells (Sadanandan et al., 2020) was reduced by CB2R agonist application, further suggesting that neuronal activity can be modulated by CB2R at the synaptic level. In addition, when CB2R was transfected into CB1R null neurons, depolarization suppression of excitation (DSE) was restored (Atwood et al., 2012). DSE, known to be mediated by CB1R, is the result of a post-synaptic endocannabinoid production that leads to presynaptic inhibition at excitatory synapses and is a form of short-term synaptic plasticity that shortly suppresses neurotransmission (Kano et al., 2009). Restoration of DSE following CB2R transfection into CB1R null neurons further highlights CB2R potency for synaptic transmission modulation. Synaptic transmission can also be indirectly modulated by regulation

of neurotransmitter receptor expression, as it was shown for 5-HT2A receptor in a neuronal cell culture following CB2R specific activation (Franklin and Carrasco, 2012).

### 4.6. Synaptic plasticity

CB2R can also modulate neuronal function through synaptic plasticity mechanisms. Namely, in mouse hippocampal slices, HU-308 bath application was shown to mimic AP-Driven long-lasting hyperpolarization i.e. slow self-inhibition in CA3 pyramidal neurons. This effect was absent in CB2R KO mice and in Syn-CB2R cKO mice, demonstrating a cell-intrinsic mechanism specifically induced by neuronal CB2R (Stempel et al., 2016). The same phenomenon was observed in Layer II/III regular spiking non-pyramidal cells (RNSCP) interneurons recorded in somatosensory cortical slices and was prevented by inverse agonist application. As for the previous study, the agonist did not have any effect on cortical slices from KO animals (Stumpf et al., 2018). *In vivo*, chronic treatment with specific CB2R agonist MDA-7 in AD mice protected long term potentiation (LTP) in CA1 neurons (Wu et al., 2017, 2013).

CB2R's role in sustaining LTP has been further confirmed in a study which showed that LTP was reduced in CB2R KO mice (Li and Kim, 2016a).

### 4.7. Dendritic spines

Spine density modulation is a potent cellular mechanism underlying synaptic function. Spine density of CA1 pyramidal cells was enhanced after chronic CB2R activation using JWH-133 or GP1α in organotypic hippocampal slices (Kim and Li,
2015). In addition, CB2R has been shown to be involved in the protection of dendritic spine dynamics, which is impaired after isoflurane exposure, and exacerbated in the hippocampus of CB2R KO mice compared to WT mice (Li et al., 2021).

Overall, evidence shows that CB2R displays region- and cell type-specific modulation of neuronal activity by acting on both the intrinsic excitability and synaptic events at the network level. So far, direct evidence for the neuronal expression of CB2R is still sparse, due to the lack of specific antibodies and neuron-specific genetic manipulations. It is therefore challenging to hypothesize on the potential mode of action of CB2R on the modulation of neuronal activity at the synaptic level. The expression of CB2R in other cell types, including microglia, is more consensual. Since microglia is known to modulate neuronal transmission (Marinelli et al., 2019; Salter and Beggs, 2014), invalidating CB2R gene expression in specific neuronal populations may contribute to elucidate the role of neuronal CB2Rs on neuronal transmission, as already performed in CA3 hippocampal neurons (Stempel et al., 2016).

## 5. Behavior modulation by CB2R

Aside from its immunomodulatory role, CB2R is now widely known to influence behavior. The effects of CB2R modulation on behavior in neuroinflammatory contexts can be explained by the modulation that CBR2 exerts directly on inflammation (see section 3). However, the fact that CB2R also exerts effects on behavior in physiological contexts implies that CB2R dependent mechanisms other than neuroinflammation may govern behavior. Here, we report studies that have provided evidence for the involvement of CB2R in different functions, under physiological and pathophysiological conditions: feeding, motor function, learning and memory, mood regulation, reward, analgesia and seizure.

### 5.1. Feeding

CB2R pharmacological activation led to different outcomes on feeding behavior depending on studies (Supplementary Table 6). Food intake was either unchanged (Martín-Sánchez et al., 2019) or reduced (Verty et al., 2015) following one single administration of a CB2R specific agonist. In diet-induced obesity models, chronic CB2R activation reduced food intake during the first six days and the body weight along the entire treatment duration, while decreasing feed efficiency ratio (Verty et al., 2015), and diminishing the visceral fat index (Youssef et al., 2019). In a PD model, treatment with a CB2R agonist prevented weight loss (Shi et al., 2017). In mice overexpressing CB2R, basal food intake remained unchanged but fasting-induced intake was reduced (Romero-Zerbo et al., 2012). Inversely, CB2R KO mouse food intake and visceral fat mass are greater than in WT mice (Agudo et al., 2010; Schmitz et al., 2016). The use of inverse agonists brought more contrasted results. Here, AM630 administration led to either unchanged (Martín-Sánchez et al., 2019) or lower (Adamczyk

et al., 2012; Ishiguro et al., 2010) food intake in rodents, or also a higher food intake after 12h food deprivation (Ishiguro et al., 2010). Of the 9 studies that provided evidence of CB2R-mediated modulation of feeding behavior, one showed a concomitant anti-inflammatory effect measured in the cortex (Youssef et al., 2019) and one showed an increase in dopamine and serotonin levels when measured in the striatum (Shi et al., 2017).

### 5.2. Motor function

The effect of CB2R activation has been studied on motor function, either as a marker of functional recovery under pathological conditions, as a proper object of study or as a control when measuring other behavioral readouts (Supplementary Table 7). When peripherally administered, CB2R agonists tend to lower locomotor activity in healthy rodents in most of the reported studies (Cortez et al., 2022; Liu et al., 2017; Onaivi et al., 2008; Spiller et al., 2019; Valenzano et al., 2005; Xi et al., 2011), whereas administration of CB2R specific inverse agonists either had no effect on locomotion (Adamczyk et al., 2012; Li and Kim, 2016b; Martín-Sánchez et al., 2019; Viveros-Paredes et al., 2017a) or enhanced it (Onaivi et al., 2008). However, basal motor activity in mice overexpressing CB2R was similar to that of WT mice as measured in open field tasks (Aracil-Fernández et al., 2012; Li and Kim, 2017) and decreased in CB2R KO mice (Ortega-Alvaro et al., 2011). Intranigral microinjection of a CB2R agonist induced contralateral turning in healthy rats (Sánchez-Zavaleta et al., 2018). The fact that CB2R modulations mediate effects on motor activity in healthy animals, i.e. in non-inflammatory contexts, and that locomotor activity is increased when CB2R is specifically deleted in dopaminergic neurons (Canseco-Alba et al., 2019; Liu et al., 2017) demonstrate that CB2R can act directly on neuronal activity.

Under pathological conditions, treatment with CB2R agonists enhanced functional recovery in models of ALS (Espejo-Porras et al., 2019), high fat diet (Youssef et al., 2019), intracerebral hemorrhage (Li et al., 2018), spinal cord injury (Adhikary et al., 2011; Latini et al., 2014), TBI (Braun et al., 2018; Elliott et al., 2011; Magid et al., 2019), stroke (Yu et al., 2015), LPS-induced sickness behavior (Sahu et al., 2019a), chronic mild stress (Onaivi, 2006) and PD (He et al., 2020; Palomo-Garo et al., 2016; Shi et al., 2017; Viveros-Paredes et al., 2017a; Yu et al., 2021). In addition, motor impairments were worsened with CB2R inverse agonist in a model of stroke (Bravo-Ferrer et al., 2017) and when CB2R gene was deleted in models of HD (Palazuelos et al., 2009) and ALS (Rodríguez-Cueto et al., 2021). Among the 33 studies that demonstrated an effect of CB2R modulation on motor function, 12 showed a concurrent anti-inflammatory effect in the different regions of interest (Adhikary et al., 2011; Braun et al., 2018; Elliott et al., 2011; Espejo-Porras et al., 2019; Li et al., 2018; Magid et al., 2019; Reiner et al., 2014; Rodríguez-Cueto et al., 2021; Sahu et al., 2019b; Viveros-Paredes et al., 2017b; Youssef et al., 2019; Yu et al., 2015) and four showed
effects on neuronal functions (Sánchez-Zavaleta et al., 2018; Xi et al., 2011; Yu et al., 2021; Zhang et al., 2017).

## 5.3. Learning and memory

CB2R signaling seems to play an important regulatory role in learning and memory (Supplementary Table 8). Acute systemic administration of CB2R agonists to healthy WT rodent was shown to enhance recognition memory (Oliveira et al., 2016), spatial learning (Tchekalarova et al., 2018) and aversive memory consolidation (García-Gutiérrez et al., 2013; Kruk-Slomka et al., 2022, 2016). Beside CB2R pharmacological activation, genetic manipulation of CB2R further highlights its implication in learning and memory. Deletion of CB2R in mice disrupted consolidation of short and long term aversive memories (García-Gutiérrez et al., 2013; Ortega-Alvaro et al., 2011). CB2R is also involved in different hippocampus-dependent memory mechanisms, as its deletion impaired contextual fear memory (Li and Kim, 2016b; Wang et al., 2018) and either enhanced or decreased spatial memory depending on the task (Supplementary Table 8), without impacting cued fear memory (Li and Kim, 2016b). Furthermore, cell-specific genetic models made it possible to pinpoint specific cellular actors of CB2R-mediated memory enhancement. Microglia-specific overexpression of CB2R in the CA1 enhanced contextual fear memory while over-expression in pyramidal neurons or interneurons did not exert any effect on memory. Conversely, CA1 pyramidal cell-specific deletion improved performances in spatial memory task and microglia-specific deletion impaired contextual fear memory while enhancing novelty memory (Li and Kim, 2017).

Pharmacological manipulation of CB2R in pathological conditions can alleviate cognitive decline. Chronic systemic administration of CB2R specific agonists protected memory performances in different genetic and pharmacologic rodent models of AD (Aso et al., 2013; Fakhfouri et al., 2012; Jayant et al., 2016; Li et al., 2019a; Martín-Moreno et al., 2012; Wu et al., 2013, 2017), age-related cognitive decline (Lindsey et al., 2019), stroke (Ronca et al., 2015), HIV-induced neuronal injury (Wang et al., 2022). high fat diet-induced (Youssef et al., 2019) and surgery-induced (Sun et al., 2017) cognitive impairment, schizophrenia (Cortez et al., 2022), okadaic acid-induced neurodegeneration (Çakır et al., 2019), sepsis-induced encephalopathy (Yang et al., 2022), and vascular dementia (Lou et al., 2017; Luo et al., 2018a). Intra-CA1 GP1α microinjection resulted similarly in enhanced memory consolidation in a step-through task in a D-AP5 (selective NMDA receptor antagonist)-induced memory impairment rat model (Nasehi et al., 2017). Conversely, inverse agonists administration decreased cognitive abilities in healthy animals (García-Gutiérrez et al., 2013; Nasehi et al., 2017) and worsened cognitive deficits in rodent models of vascular dementia (Luo et al., 2018a) and D-AP5-induced (Nasehi et al., 2017) and surgery-induced cognitive impairments (Sun et al., 2017). Cognitive impairments were worsened when CB2R was deleted in models of aging (Komorowska-Müller et al.,

2021b) and isoflurane-induced cognitive decline (Li et al., 2021) but improved in models of AD (Galán-Ganga et al., 2021; Schmöle et al., 2018, 2015). It is to note that CB2R KO mice displayed AD-like tau hyperphosphorylation alongside their cognitive impairments (Wang et al., 2018).

Of the 34 studies that demonstrated an effect of CB2R modulation on memory and/or learning, 16 showed a concomitant immunomodulatory effect in the hippocampus and/or the cortex in pathological contexts (Aso et al., 2013; Çakır et al., 2019; Fakhfouri et al., 2012; Jayant et al., 2016; Li et al., 2021, 2019b; Luo et al., 2018b; Martín-Moreno et al., 2012; Schmöle et al., 2018, 2015; Sun et al., 2017; Wang et al., 2022; Wu et al., 2017, 2013; Yang et al., 2022; Youssef et al., 2019) and two showed protection of hippocampal long-term potentiation (Wu et al., 2017, 2013).

## 5.4. Mood

Chronic peripheral administration of CB2R agonists produced different effects on anxiety and depressive-like behavior in healthy rodents depending on the behavioral task (García-Gutiérrez et al., 2012; Onaivi et al., 2008; Valenzano et al., 2005; Verty et al., 2015) (Supplementary Table 9). CB2R pharmacological activation under pathological conditions counteracted anxiety- and/or depression-like behaviors induced by chronic stress (Hwang et al., 2020; Onaivi et al., 2008), neuropathic pain (Cabañero et al., 2020), LPS (Sahu et al., 2019a), high-fat diet (Youssef et al., 2019), sepsis-induced encephalopathy (Yang et al., 2022) and TBI (Braun et al., 2018). Interestingly, treatment with inverse agonists also reduced the basal level of anxiety in healthy laboratory-reared rodents (García-Gutiérrez et al., 2010; García-Gutiérrez and Manzanares, 2011) and anxiety and depression-like traits in models of anxiety (García-Gutiérrez and Manzanares, 2011; Ten-Blanco et al., 2022b), orexin-induced fear extinction deficit (Ten-Blanco et al., 2022a) and also TBI (Reiner et al., 2014).

Transgenic mice specifically overexpressing CB2R in the central nervous system exhibited an endophenotypic resistance to acute and chronic anxiogenic- and depression-like stimuli (García-Gutiérrez et al., 2010; García-Gutiérrez and Manzanares, 2011; Romero-Zerbo et al., 2012). In contrast, CB2R KO mice displayed high vulnerability to stressful stimuli (Liu et al., 2017; Ortega-Alvaro et al., 2011). It is to note that these phenotypes were not observed when CB2R overexpression or deletion was specifically induced in CA1 cells (Li and Kim, 2017). CB2R genetic deletion either induced anxiety-like behavior (Ortega-Alvaro et al., 2011) or had no effect (Li and Kim, 2016b) depending on CB2R KO mouse strain and test. Furthermore, depression-like behaviors are induced when CB2R is specifically deleted in dopaminergic neurons (Liu et al., 2017). However, in CB2R KO mice, mood dysregulation was lowered in aging (Komorowska-Müller et al., 2021b) and chronic mild stress (Liu et al., 2017) models.
Among the 19 studies that evidenced an effect of CB2R modulation on mood, 6 showed a simultaneous immunomodulatory effect (Braun et al., 2018; Hwang et al., 2020; Reiner et al., 2014; Sahu et al., 2019b; Yang et al., 2022; Youssef et al., 2019) and one showed an increase in field excitatory post-synaptic potential and a reduction in long term depression induced by LPS exposure in hippocampal slices (Hwang et al., 2020).

## 5.5. Reward and addiction

Accumulating evidence supports the potency of cannabinoids in treating drug addiction (Gonzalez-Cuevas et al., 2018) (Supplementary Table 10). Presently, the area of treating addiction is relatively unexplored with a current lack of viable therapies aside from lower doses or analogues of the offending substance. Coinciding with CB2R ability to modulate behaviors and mood, accumulating evidence supports the potency of cannabinoids in treating drug addiction.

### 5.5.1. Alcohol

CB2R appears to be involved in alcohol abuse as CB2R KO mice developed more robust alcohol conditioned place preference (CPP) and have a higher alcohol consumption than their WT littermates (Ortega-Álvaro et al., 2015; Powers et al., 2015). Intriguingly, specific deletion of CB2R in dopaminergic neurons reduced alcohol CPP (Liu et al., 2020). Chronic activation of CB2R signaling using specific agonists reduced alcohol rewarding behavior (Al Mansouri et al., 2014; Martín-Sánchez et al., 2019). Nevertheless, one single administration of CB2R agonist failed to modify alcohol intake and alcohol-CPP expression (Powers et al., 2015). It should be noted that different results were obtained under various stress conditions. CB2R pharmacological stimulation, in mice subjected to chronic mild stress, enhanced alcohol intake (Ishiguro et al., 2007; Onaivi et al., 2008). Moreover, CB2R invalidation in dopaminergic neurons, in mice exposed to subacute stress, displayed lower alcohol consumption as opposed to non-stressed mice (Liu et al., 2017). Taken together, these studies highlight the fact that CB2R may exert different roles depending on the cell type and physiological conditions.

### 5.5.2. Psychostimulants (Cocaine, nicotine, amphetamine)

The role of CB1R on the effects of psychostimulants has already been widely addressed (Chaperon et al., 1998; Peters et al., 2021; Solinas et al., 2008) but the role of CB2R is now receiving greater attention. Both peripheral, intra-accumbens and intra-VTA administration of CB2R selective agonists blocked cocaine-induced CPP (Canseco-Alba et al., 2019; Delis et al., 2016; Ignatowska-Jankowska et al., 2013; Lopes et al., 2020; Xi et al., 2011) and reduced cocaine self-administration (Xi et al., 2011; Zhang et al., 2015, 2014). This inhibition was reversed by the inverse agonist AM630 (Delis et al., 2016; Xi

et al., 2011; Zhang et al., 2014) and absent in CB2R KO mice (Xi et al., 2011; Zhang et al., 2015, 2014). Genetic manipulation studies support the inhibitory role of CB2R in cocaine-induced behavioral alteration as CB2R overexpressing mice display a conditioned place aversion and reduced cocaine self-administration (Aracil-Fernández et al., 2012), while specific deletion of the CB2R gene in dopaminergic neurons leads to an increased cocaine- (Liu et al., 2017), amphetamine- and methamphetamine-induced CPP (Canseco-Alba et al., 2019). These results bring a new insight on the involvement of CB2Rs expressed by dopaminergic neurons in the modulation of the reinforcing effects of cocaine, amphetamine and methamphetamine. However, CB2R activation seems to mediate opposite effects on consumption of nicotine, another psychostimulants, as both genetic invalidation and pharmacological blockade of CB2R abolished nicotine rewarding properties (Gamaleddin et al., 2012; Ignatowska-Jankowska et al., 2013; Navarrete et al., 2013). In the case of nicotine, the CNS stimulant effect is associated with the cholinergic system. Therefore, CB2R seems to be differently involved in cholinergic and dopaminergic pathways. Taken together, these data support the notion that CB1R and CB2R have different and perhaps opposing roles in modulating cocaine's rewarding and psychomotor stimulant effects. A study using electrical intracranial brain-stimulation reward (BSR) paradigm provides support for this idea as animals treated with selective CB1R and CB2R receptor agonists produced significant BSR enhancement and inhibition, respectively (Spiller et al., 2019).

### 5.5.3. Opioids

Although few studies have examined the impact of CB2R ligands on opioid abuse so far, CB2R is known to interact with mu opioid receptors to attenuate chronic pain. Co-administration of a CB2R agonist alongside morphine produces synergistic increases in the anti-nociceptive effects of morphine (Grenald et al., 2017) (see section 5.6). In the same study, peripheral treatment with JWH-015, a CB2R agonist, decreased morphine-induced CPP. Hence, co-treatment with CB2R agonists allows significant synergistic inhibition of preclinical pain while reducing opioid-induced unwanted side effects. Further investigations are needed to understand the role of CB2R at the cellular and molecular level.

### 5.5.4. Other

Peripheral administration of CB2R agonist JWH-133 reduced food- (Martín-Sánchez et al., 2019) and sucrose- (Zhang et al., 2015) induced CPP in mice. The inhibitory effects of CB2R activation on reward circuits observed with alcohol and psychostimulants like cocaine appear therefore to be generalized to natural rewards such as food or sucrose. Nonetheless, the food-induced CPP was not diminished by a single dose of the CB2R inverse agonist AM630 unlike alcohol
<table><caption>**Table 2. Co-incidence of immunomodulatory and/or neuromodulatory effects with behavioral outcomes following modulation of CB2R activity.** Inflammatory, neuronal or behavioral outcome following CB2R modulation is either enhanced (✓) or decreased (✓). The symbol “-” was used when the type of outcome has not been investigated by the corresponding study.</caption><thead><tr><th>MODEL</th><th>CB2R MODULATION</th><th>BRAIN REGION</th><th>INFLAMMATORY OUTCOME</th><th>NEURONAL OUTCOME</th><th>BEHAVIORAL OUTCOMES</th><th>REF</th></tr></thead><tbody><tr><td rowspan="3">Healthy (Mouse)</td><td rowspan="3">Pharmacological activation</td><td>nucleus accumbens</td><td>-</td><td>✓ extracellular DA</td><td>✓ basal and cocaine-enhanced locomotion</td><td>(Xi et al., 2011)</td></tr><tr><td>VTA</td><td>-</td><td>✓ excitability</td><td>✓ cocaine self-administration</td><td>(Zhang et al., 2014)</td></tr><tr><td>Substansia nigra</td><td>-</td><td>✓ autonomous firing rate</td><td>✓ motor function</td><td>(Yu et al., 2021)</td></tr><tr><td>Healthy (Rat)</td><td>Pharmacological activation</td><td>Substansia nigra</td><td>-</td><td>✓K-stimulated [³H]-Glutamate release</td><td>Induction of contralateral turning (2-4 turns/min)</td><td>(Sánchez-Zavaleta et al., 2018)</td></tr><tr><td rowspan="3">Alzheimer's disease (APP/PS1 Tg Mouse)</td><td rowspan="3">Pharmacological activation</td><td>Cortex</td><td>✓ Inflammation</td><td>-</td><td>✓ recognition memory</td><td>(Aso et al., 2013)</td></tr><tr><td>Cortex</td><td>✓ Inflammation</td><td>-</td><td>✓ recognition memory</td><td>(Li et al., 2019a)</td></tr><tr><td>Hippocampus, entorhinal cortex</td><td>✓ Inflammation</td><td>✓ LTP</td><td>✓ memory</td><td>(Wu et al., 2017)</td></tr><tr><td>Alzheimer's disease (APP2576 Tg Mouse)</td><td>Pharmacological activation</td><td>Cortex</td><td>✓ Inflammation</td><td>-</td><td>✓ recognition memory</td><td>(Martín-Moreno et al., 2012)</td></tr><tr><td>Alzheimer's disease (icv STZ, Mouse)</td><td>Pharmacological activation</td><td>Brain</td><td>✓ Inflammation</td><td>-</td><td>✓ memory performance</td><td>(Jayant et al., 2016)</td></tr><tr><td>Alzheimer's disease (intrahippocampal Aβ<sub>1-40</sub>, Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>✓ LTP, ✓ eEPSC amplitude</td><td>✓ spatial learning</td><td>(Wu et al., 2013)</td></tr><tr><td>Alzheimer's disease (intrahippocampal Aβ<sub>1-42</sub>, Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>-</td><td>✓ spatial learning</td><td>(Fakhfouri et al., 2012)</td></tr><tr><td rowspan="2">Alzheimer's disease (APP/PS1 Tg Mouse)</td><td rowspan="2">Genetic blockade</td><td>Peri-plaques areas</td><td>✓ Inflammation</td><td>-</td><td>✓ spatial memory</td><td>(Schmölle et al., 2018)</td></tr><tr><td>Brain</td><td>✓ Inflammation</td><td>-</td><td>✓ spatial memory at 6 months</td><td>(Schmölle et al., 2015)</td></tr><tr><td rowspan="2">Amyotrophic lateral sclerosis (Mouse)</td><td rowspan="2">Pharmacological activation</td><td>Ventral and dorsal horn</td><td>✓ Inflammation</td><td>-</td><td>✓ locomotor activity</td><td>(Espejo-Porras et al., 2019)</td></tr><tr><td>Spinal cord</td><td>✓ Inflammation</td><td>-</td><td>✓ motor function</td><td>(Rodríguez-Cueto et al., 2021)</td></tr><tr><td>Chronic restraint stress (Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>✓ fEPSPs, ✓ LPS-induced intensification of LTD</td><td>✓ depression</td><td>(Hwang et al., 2020)</td></tr><tr><td>Cocaine (Mouse)</td><td>Pharmacological activation</td><td>Nucleus accumbens</td><td>-</td><td>✓ cocaine-enhanced extracellular DA</td><td>✓ cocaine self-administration (dose-dep), ✓ cocaine's rewarding efficacy.</td><td>(Xi et al., 2011)</td></tr><tr><td>Diet-induced inflammation (Rat)</td><td>Pharmacological activation</td><td>Prefrontal cortex</td><td>✓ Inflammation</td><td>-</td><td>✓ visceral fat index, ✓ spontaneous locomotor activity, ✓ memory, ✓ anxiety and depression</td><td>(Youssef et al., 2019)</td></tr><tr><td>HIV-1 glycoprotein 120 induced neural injury (Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>-</td><td>✓ learning and memory performance</td><td>(Wang et al., 2022)</td></tr><tr><td>Intracerebral Hemorrhage (Rat)</td><td>Pharmacological activation</td><td>Peri hematoma-brain</td><td>✓ Inflammation</td><td>-</td><td>✓ sensorymotor and balance function</td><td>(Li et al., 2018)</td></tr><tr><td>Isoflurane-induced inflammation (mouse)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>-</td><td>✓ spatial learning and memory</td><td>(Li et al., 2021)</td></tr><tr><td>LPS-induced inflammation (Mouse)</td><td>Pharmacological activation</td><td>Brain</td><td>✓ Inflammation</td><td>-</td><td>✓ locomotor activity, ✓ anxiety, depression</td><td>(Sahu et al., 2019a)</td></tr><tr><td>Morphine (Rat)</td><td>Pharmacological activation</td><td>Nucleus accumbens</td><td>-</td><td>✓ morphine-enhanced extracellular DA</td><td>✓ Morphine-induced CPP, Alone: analgesic effect (✓ Morphine-induced analgesia</td><td>(Grenald et al., 2017)</td></tr><tr><td>Morphine-mediated hyperalgesia (Rat)</td><td>Pharmacological activation</td><td>Spinal Cord</td><td>✓ Inflammation</td><td>-</td><td>✓ Morphine-mediated thermal hyperalgesia and tactile allodynia</td><td>(Tumati et al., 2012)</td></tr><tr><td rowspan="2">Multiple sclerosis (Mouse)</td><td rowspan="2">Pharmacological activation</td><td>Lumbar spinal cord</td><td>✓ Inflammation</td><td>-</td><td>✓ mechanical hyperalgesia</td><td>(Alberti et al., 2017)</td></tr><tr><td>Spinal cord</td><td>✓ Inflammation</td><td>-</td><td>✓ basal nociceptive response, ✓ thermal hyperalgesia, mechanical and thermal allodynia</td><td>(Racz et al., 2008)</td></tr><tr><td rowspan="2">Neuropathic pain (Mouse)</td><td rowspan="2">Genetic blockade</td><td>Spinal cord</td><td>✓ Inflammation</td><td>-</td><td>✓ Analgesic effect, ✓ mechanical and thermal hyperalgesia</td><td>(Klauke et al., 2014)</td></tr><tr><td>Pharmacological activation</td><td>✓ Inflammation</td><td>-</td><td>✓ spatial memory performance</td><td>(Çakır et al., 2019)</td></tr><tr><td>Okadaic acid-induced neurodegeneration (Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>✓ Inflammation</td><td>-</td><td>✓ MPTP-induced movement impairment, ✓ stride length impairment, ✓ balance impairment</td><td>(Viveros-Paredes et al., 2017a)</td></tr><tr><td rowspan="2">Parkinson's disease (Mouse)</td><td rowspan="2">Pharmacological activation</td><td>Substansia Nigra and striatum</td><td>✓ Inflammation</td><td>-</td><td>Prevention of MPTP-induced weight loss, ✓ motor function</td><td>(Shi et al., 2017)</td></tr><tr><td>Striatum</td><td>-</td><td>✓ DA and 5-HT levels</td><td></td><td></td></tr></tbody></table>
<table><thead><tr><th></th><th></th><th></th><th></th><th></th><th></th><th></th></tr></thead><tbody><tr><td>Retrovirus Infection-Induced Neuropathic Pain (Mouse)</td><td>Pharmacological activation</td><td>Dorsal root ganglia & lumbar spinal cord</td><td>Inflammation</td><td>-</td><td>allodynia (mechanical hypersensitivity)</td><td>(Sheng et al., 2019)</td></tr><tr><td>Sepsis-induced encephalopathy (Mouse)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>Inflammation</td><td>-</td><td>recognition and spatial memory, anxiety</td><td>(Yang et al., 2022)</td></tr><tr><td>Spinal cord injury (Mouse)</td><td>Pharmacological activation</td><td>Spinal cord</td><td>Inflammation</td><td>-</td><td>motor function</td><td>(Adhikary et al., 2011)</td></tr><tr><td>Stroke (Rat)</td><td>Pharmacological activation</td><td>Ischemic cortex</td><td>Inflammation</td><td>-</td><td>Functional recovery</td><td>(Yu et al., 2015)</td></tr><tr><td>Surgery-induced cognitive impairment (Mouse)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>Inflammation</td><td>-</td><td>contextual fear memory</td><td>(Sun et al., 2017)</td></tr><tr><td rowspan="4">Traumatic Brain Injury (Mouse)</td><td>Pharmacological blockade</td><td>Optic tract</td><td>Inflammation</td><td>-</td><td>locomotor activity, depression, fear extinction, fear</td><td>(Reiner et al., 2014)</td></tr><tr><td>Pharmacological activation</td><td>Cortex</td><td>Inflammation</td><td>-</td><td>locomotor performance, usage of forelimb</td><td>(Elliott et al., 2011)</td></tr><tr><td>Pharmacological activation</td><td>Cortex</td><td>Inflammation</td><td>-</td><td>motor coordination, anxiety</td><td>(Braun et al., 2018)</td></tr><tr><td>Pharmacological activation</td><td>Cortex and hippocampus</td><td>Inflammation</td><td>-</td><td>locomotor performance</td><td>(Magid et al., 2019)</td></tr><tr><td rowspan="2">Vascular dementia (Rat)</td><td>Pharmacological activation</td><td>Hippocampus</td><td>Inflammation</td><td>-</td><td>spatial memory</td><td>(Luo et al., 2018a)</td></tr><tr><td>Pharmacological blockade</td><td>Hippocampus</td><td>Inflammation</td><td>-</td><td>spatial memory</td><td>(Luo et al., 2018a)</td></tr></tbody></table>

**Abbreviations:** 5-HT, serotonin; CPP, conditioned place preference; DA, dopamine; EPSP, excitatory post-synaptic potential; LPS, lipopolysaccharide; LTD, long term depression; LTP, long term potentiation; MPTP, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Ref, reference; Tg, transgenic; VTA, ventral tegmental area.

-induced CPP. In this regard, CB2R inactivation appears to specifically interfere with drug-induced CPP induction, but not when a natural reinforcer is presented, such as food.

Overall, these findings support brain CB2R involvement in drug reward and addiction and suggest that brain CB2R may constitute a new target in therapeutics development for the treatment of substance abuse. Among the 20 studies that demonstrated an effect of CB2R modulation on addiction and reward behaviors, three showed an effect on neuronal activity, namely on extracellular dopamine levels in the nucleus accumbens (Grenald et al., 2017; Xi et al., 2011) and on the excitability of dopaminergic neurons in the VTA (Zhang et al., 2014). Knowing that CB2R expression has been robustly shown in these neurons, these results suggest a direct effect of CB2R activation on their activity which results in a behavioral outcome.

## 5.6. Analgesia

A growing body of evidence suggests that CB2R plays a role in pain modulation (Supplementary Table 11). Systemic treatment with CB2R specific agonists in rodents promoted an analgesic effect by itself, or potentiated opioid-induced analgesia (Altun et al., 2015; Grenald et al., 2017; Valenzano et al., 2005). Depending on the pain assay, GP1a alone may exert analgesic effects or enhance analgesia when combined with morphine (Chen et al., 2019). Hence, using cannabinoids may reduce the dose of opioids needed for analgesia in certain pain conditions. In addition, morphine-mediated thermal hyperalgesia and tactile allodynia were reduced following systemic CB2R agonist administration, suggesting that CB2R activation may be useful to prevent the neuroinflammatory consequences of sustained morphine treatment (Tumati et al., 2012). Other CB2R agonists were also shown to exert analgesic effects when administered to healthy rodents

(Altun et al., 2015; Cortez et al., 2022; Grenald et al., 2017; Klauke et al., 2014; Valenzano et al., 2005).

In different pathologic rodent models, CB2R agonists also presented antinociceptive properties, such as in a chronic inflammatory pain model (Negrete et al., 2011), multiple sclerosis (Alberti et al., 2017), high-fat diet (Schmitz et al., 2016), neuropathic pain (Cabañero et al., 2020; Grenald et al., 2017; Ibrahim et al., 2003; Klauke et al., 2014; Sheng et al., 2019; Shiue et al., 2017; Valenzano et al., 2005), incisional and inflammatory (Valenzano et al., 2005) and post-operative pain (Grenald et al., 2017).

Genetic models of invalidation further support the antinociceptive potency of CB2R. In fact, basal nociceptive responses were enhanced in several behavioral tests in CB2R KO mice (Racz et al., 2008). Dopaminergic neurons may be differentially involved in mechanisms underlying various pain conditions, as the pain threshold was raised in the tail flick test but tended to be lowered in the hot plate test in mice with CB2R deletion in dopaminergic neurons (DAT-CB2R cKO mice) (Liu et al., 2020, 2017). Irradiated WT mice reconstituted with bone marrow (BM) cells from CB2R KO mice developed the same enhanced manifestations of neuropathic pain as CB2R KO animals, demonstrating the protective role of CB2R in BM-derived cells that are recruited to the spinal cord in the development of neuropathic pain (Racz et al., 2008).

Of the 17 studies that showed an effect of CB2R modulation on pain perception, five showed a concomitant immunomodulatory effect (Alberti et al., 2017; Klauke et al., 2014; Racz et al., 2008; Sheng et al., 2019; Tumati et al., 2012) and one showed an effect on extracellular dopamine levels measured in the nucleus accumbens (Grenald et al., 2017).
## 5.7. Seizure activity

CB1R modulation can improve seizure outcomes, but is accompanied by several psychotropic effects, limiting its potential as a therapeutic target (Kow et al., 2014; Pertwee, 2012). CB2R has received less attention for its role in seizure activity so far. The few studies that have directly examined the effect of modulating CB2R activity on seizure susceptibility have yielded inconsistent results (Supplementary Table 12). The CB2R agonist β-caryophyllene (BCP) decreases seizure frequency and spread in mice undergoing maximal electroshock seizures (Tchekalarova et al., 2018). BCP and HU308 were found to increase the latency to myoclonic jerks induced by PTZ and to lower the seizure threshold (Ghanbari et al., 2020; Oliveira et al., 2016). However, it is also reported that direct brain injection of the CB2R agonist, AM1241, increased seizure severity in rats (de Carvalho et al., 2016). Additionally, the CB2R agonists HU-308 and BCP had no effect on PTZ-induced seizures in young rodents (Huizenga et al., 2017; Tchekalarova et al., 2018). JWH-133 administration, following pilocarpine-induced status epilepticus, reduced mortality and increased the latent period for the first seizure attack (Cao et al., 2021). Overall, while there is evidence that modulating CB2R activity can affect seizure susceptibility, this relationship remains unclear, highlighting the impact of differences in study design, including species, seizure model, CB2R agonist and dosing (Supplementary Table 12). The use of genetic manipulation brought new insight into the role of CB2R in seizure activity. Deletion of CB2R in mice resulted in a higher severity score of handling induced seizures after alcohol consumption (Ortega-Álvaro et al., 2015), and a higher average Racine score in a 6 Hz seizure induction paradigm (Shapiro et al., 2019). Furthermore, both heterozygous and homozygous KO mice exhibited increased susceptibility to PTZ-induced seizures. Among double KO mice with both CB1R and CB2R deletions, 27.6% of mice exhibited handling induced seizures and 80% mice had at least 1 spontaneous electrographic seizure, but no seizure were recorded from CB2R KO mice (Rowley et al., 2017), suggesting a synergic role of both CB1R and CB2R. Administration of the CB2R inverse agonist, SR144528, in WT mice increased seizure susceptibility, suggesting that increasing CB2R expression or activity may grant seizure protection (Shapiro et al., 2019).

Of the studies listed in this review that investigated the effect of CB2R modulation on epileptic activity, none looked for a parallel effect on inflammation or on neuronal activity. To better understand the role of CB2R in seizure susceptibility, it is crucial that future studies take into account these potential effects on neuroinflammation, which is known to be involved in the occurrence of seizures and comorbidities of epilepsy (Vezzani, 2020), and to investigate possible effects on neuronal activity whose imbalance leads to seizure occurrence.

Given the broad involvement of CB2R in behavior, understanding brain CB2R signaling may provide new insights into treatments for brain disorders ranging from dementia to epilepsy and drug addiction. Nonetheless, the underlying mechanisms at a cellular and molecular level remain unclear. To date, studies using cell-specific genome-editing techniques are still scarce. Studies in DAT-CB2R cKO mice have already demonstrated the involvement of CB2R in dopaminergic neurons in motor activities, anxiety and depression-like behaviors and in the differential rewarding properties of alcohol and various psychostimulants (Canseco-Alba et al., 2019; Liu et al., 2017). CB2R expression in microglia or in CA1 pyramidal neurons was shown to be involved in contextual fear memory or in spatial working memory, respectively (Li and Kim, 2016b). Once identity of cells expressing CB2R in brain have been accurately established (Atwood and Mackie, 2010; Grabon et al., 2023), future studies using novel manipulations of CB2R expression in specific types or subtypes of cells in different brain regions will allow robust isolation of the CB2R's role in behavior.

## 6. Targeting CB2R in clinical trials

To date, only three of the CB2R agonists listed in this review have been included in clinical studies. β-caryophyllene was tested in a phase 2 trial for the treatment of pain related to osteoarthritis of the knee as a topical cream in 56 participants (NCT03152578). The safety evaluation of the MDA7 agonist tested in phase 1 (NCT04375436) supports feasibility in clinical use and is likely to be tested in further clinical studies (Foss et al., 2020). Finally, paeoniflorin - in combination with other compounds - has been tested for the treatment of rheumatoid arthritis (NCT01600521). Although not as effective as current disease-modifying antirheumatic drugs (methotrexate and leflunomide), paoniflorin appears to be a safer option as a substitute for these drugs in long-term rheumatoid arthritis treatment when their toxicity is an issue (Chen et al., 2013).

## 7. Conclusion

Evidence has accumulated over the last decade showing the broad involvement of CB2R in the control of brain functions, particularly in behavior, through neuroimmune processes. Of the 100 studies included in this review that showed behavioral outcomes following CB2R modulation, 33 observed a concomitant modulation of the neuro-inflammatory state and nine showed changes in neuronal activity (Table 2). This suggests that CB2R dependent modulation of behavior is indeed mediated by immunomodulatory and/or neural mechanisms, either directly or indirectly. Highly expressed by resident microglial cells, and presumably leukocytes infiltrating the CNS, CB2R appears as a promising candidate in drug development for targeting immunomodulatory processes, in particular those mediating changes in the polarization of brain immune cells towards a pro-healing state. The findings of discrete CB2R expression by certain neuronal populations and direct
control of neuronal activity by CB2R may explain some
modulations of behavior observed following CB2R activation.
Further developments of new genetic models of inducible
*cn2r* gene invalidation, restricted to certain cell populations,
is crucial to decipher the multifactorial mechanisms of which
the endocannabinoid system, and CB2R in particular, can
influence brain function and behavior.

Competing Interests

The authors declare no competing interests.

References

Adamczyk, P., Miszkiel, J., McCreary, A.C., Filip, M., Papp, M., Przegaliński, E., 2012. The effects of cannabinoid CB1, CB2 and vanilloid TRPV1 receptor antagonists on cocaine addictive behavior in rats. Brain Research 1444, 45-54. https://doi.org/10.1016/j.brainres.2012.01.030

Adhikary, S., Li, H., Heller, J., Skarica, M., Zhang, M., Ganea, D., Tuma, R.F., 2011. Modulation of Inflammatory Responses by a Cannabinoid-2-Selective Agonist after Spinal Cord Injury. J Neurotrauma 28, 2417-2427. https://doi.org/10.1089/neu.2011.1853

Agudo, J., Martin, M., Roca, C., Molas, M., Bura, A.S., Zimmer, A., Bosch, F., Maldonado, R., 2010. Deficiency of CB2 cannabinoid receptor in mice improves insulin sensitivity but increases food intake and obesity with age. Diabetologia 53, 2629-2640. https://doi.org/10.1007/s00125-010-1894-6

Al Mansouri, S., Ojha, S., Al Maamari, E., Al Ameri, M., Nurulain, S.M., Bahi, A., 2014. The cannabinoid receptor 2 agonist, β-caryophyllene, reduced voluntary alcohol intake and attenuated ethanol-induced place preference and sensitivity in mice. Pharmacology Biochemistry and Behavior 124, 260-268. https://doi.org/10.1016/j.pbb.2014.06.025

Alberti, T.B., Barbosa, W.L.R., Vieira, J.L.F., Raposo, N.R.B., Dutra, R.C., 2017. (-)-β-Caryophyllene, a CB2 Receptor-Selective Phytocannabinoid, Suppresses Motor Paralysis and Neuroinflammation in a Murine Model of Multiple Sclerosis. *Int J Mol Sci* 18, 691. https://doi.org/10.3390/ijms18040691

Altun, A., Yildirim, K., Ozdemir, E., Bagcivan, I., Gursoy, S., Durmus, N., 2015. Attenuation of morphine antinociceptive tolerance by cannabinoid CB1 and CB2 receptor antagonists. *J Physiol Sci* 65, 407-415. https://doi.org/10.1007/s12576-015-0379-2

Amenta, P.S., Jallo, J.I., Tuma, R.F., Hooper, D.C., Elliott, M.B., 2014. Cannabinoid receptor type-2 stimulation, blockade, and deletion alter the vascular inflammatory responses to traumatic brain injury. *J Neuroinflammation* 11. https://doi.org/10.1186/s12974-014-0191-6

Andó, R.D., Bíró, J., Csölle, C., Ledent, C., Sperlágh, B., 2012. The inhibitory action of exo- and endocannabinoids on [3H]GABA release are mediated by both CB1 and CB2 receptors in the mouse hippocampus. *Neurochemistry International* 60, 145-152. https://doi.org/10.1016/j.neuint.2011.11.012

Aracil-Fernández, A., Trigo, J.M., García-Gutiérrez, M.S., Ortega-Alvaro, A., Ternianov, A., Navarro, D., Robledo, P., Berbel, P., Maldonado, R., Manzanares, J., 2012. Decreased Cocaine Motor Sensitization and Self-Administration in Mice Overexpressing Cannabinoid CB2 Receptors. *Neuropsychopharmacology* 37, 1749-1763. https://doi.org/10.1038/npp.2012.22

Araque, A., Castillo, P.E., Manzoni, O.J., Tonini, R., 2017. Synaptic functions of endocannabinoid signaling in health and disease. Neuropharmacology 124, 13-24. https://doi.org/10.1016/j.neuropharm.2017.06.017

Askari, V.R., Shafiee-Nick, R., 2019a. The protective effects of β-caryophyllene on LPS-induced primary microglia M1/M2 imbalance: A mechanistic evaluation. *Life Sciences* 219, 40-73. https://doi.org/10.1016/j.lfs.2018.12.059

Askari, V.R., Shafiee-Nick, R., 2019b. Promising neuroprotective effects of β-caryophyllene against LPS-induced oligodendrocyte toxicity: A mechanistic study. *Biochemical Pharmacology* 159, 154-171. https://doi.org/10.1016/j.bcp.2018.12.001

Aso, E., Juvés, S., Maldonado, R., Ferrer, I., 2013. CB 2 Cannabinoid Receptor Agonist Ameliorates Alzheimer-Like Phenotype in AβPP/PS1 Mice. *Journal of Alzheimer's Disease* 35, 847-858. https://doi.org/10.3233/JAD-130137

Atwood, B.K., Mackie, K., 2010. CB2: a cannabinoid receptor with an identity crisis. *Br J Pharmacol* 160, 467-479. https://doi.org/10.1111/j.1476-5381.2010.00729.x

Atwood, B.K., Straiker, A., Mackie, K., 2012. CB2 cannabinoid receptors inhibit synaptic transmission when expressed in cultured autaptic neurons. *Neuropharmacology* 63, 514-523. https://doi.org/10.1016/j.neuropharm.2012.04.024

Beltramo, M., Bernardini, N., Bertorelli, R., Campanella, M., Nicolussi, E., Fredduzzi, S., Reggiani, A., 2006. CB2 receptor-mediated antihyperalgesia: possible direct involvement of neural mechanisms. *European Journal of Neuroscience* 23, 1530-1538. https://doi.org/10.1111/j.1460-9568.2006.04684.x

Belvisi, M.G., Patel, H.J., Freund-Michel, V., Hele, D.J., Crispino, N., Birrell, M.A., 2008. Inhibitory activity of the novel CB2 receptor agonist, GW833972A, on guinea-pig and human sensory nerve function in the airways. *Br J Pharmacol* 155, 547-557. https://doi.org/10.1038/bjp.2008.298

Benito, C., Núñez, E., Tolón, R.M., Carrier, E.J., Rábano, A., Hillard, C.J., Romero, J., 2003. Cannabinoid CB2 Receptors and Fatty Acid Amide Hydrolase Are Selectively Overexpressed in Neuritic Plaque-Associated Glia in Alzheimer's Disease Brains. *J Neurosci* 23, 11136-11141. https://doi.org/10.1523/JNEUROSCI.23-35-11136.2003

Benito, C., Tolón, R.M., Pazos, M.R., Núñez, E., Castillo, A.I., Romero, J., 2008. Cannabinoid CB2 receptors in human brain inflammation. *Br J Pharmacol* 153, 277-285. https://doi.org/10.1038/sj.bjp.0707505

Bhattacharjee, H., Gurley, S.N., Moore, B.M., 2009. Design and synthesis of novel tri-aryl CB2 selective cannabinoid ligands. *Bioorganic & Medicinal Chemistry Letters* 19, 1691-1693. https://doi.org/10.1016/j.bmcl.2009.01.100

Bouaboula, M., Desnoyer, N., Carayon, P., Combes, T., Casellas, P., 1999. Gi protein modulation induced by a selective inverse agonist for the peripheral cannabinoid receptor CB2: implication for intracellular signalization cross-regulation. *Mol Pharmacol* 55, 473-480.

Braun, M., Khan, Z.T., Khan, M.B., Kumar, M., Ward, A., Achyut, B.R., Arbab, A.S., Hess, D.C., Hoda, Md.N., Baban, B., Dhandapani, K.M., Vaibhav, K., 2018. Selective activation of cannabinoid receptor-2 reduces neuroinflammation after traumatic brain injury via alternative macrophage polarization. *Brain Behav Immun* 68, 224-237. https://doi.org/10.1016/j.bbi.2017.10.021

Bravo-Ferrer, I., Cuartero, M.I., Zarruk, J.G., Pradillo, J.M., Hurtado, O., Romera, V.G., Díaz-Alonso, J., García-Segura, J.M., Guzmán, M., Lizasoain, I., Galve-Roperh, I., Moro, M.A., 2017. Cannabinoid Type-2 Receptor Drives Neurogenesis and Improves Functional Outcome After Stroke. *Stroke* 48, 204-212. https://doi.org/10.1161/STROKEAHA.116.014793

Buckley, N.E., McCoy, K.L., Mezey, É., Bonner, T., Zimmer, Anne, Felder, C.C., Glass, M., Zimmer, Andreas, 2000. Immunomodulation by cannabinoids is absent in mice deficient for the cannabinoid CB2 receptor. *European Journal of Pharmacology* 396, 141-149. https://doi.org/10.1016/S0014-2999(00)00211-9

Cabañero, D., Ramírez-López, A., Drews, E., Schmöle, A., Otte, D.M., Wawrzczak-Bargiela, A., Huerga Encabo, H., Kummer, S., Ferrer-Montiel, A., Przewlocki, R., Zimmer, A., Maldonado, R., 2020. Protective role of neuronal and lymphoid cannabinoid CB2 receptors in neuropathic pain. *eLife* 9, e55582. https://doi.org/10.7554/eLife.55582

Cabral, G.A., Raborn, E.S., Griffin, L., Dennis, J., Marciano-Cabral, F., 2008. CB2 receptors in the brain: role in central immune function. *Br J Pharmacol* 153, 240-251. https://doi.org/10.1038/sj.bjp.0707584

Çakır, M., Tekin, S., Doğanyiğit, Z., Erden, Y., Soytürk, M., Çiğremiş, Y., Sandal, S., 2019. Cannabinoid type 2 receptor agonist JWH-133, attenuates Okadaic acid induced spatial memory impairment and neurodegeneration in rats. *Life Sci* 217, 25-33. https://doi.org/10.1016/j.lfs.2018.11.058
Çakir, M., Tekin, S., Okan, A., Çakan, P., Doğanyiğit, Z., 2020. The ameliorating effect of cannabinoid type 2 receptor activation on brain, lung, liver and heart damage in cecal ligation and puncture-induced sepsis model in rats. International Immunopharmacology 78, 105978.
https://doi.org/10.1016/j.intimp.2019.105978

Canseco-Alba, A., Schanz, N., Sanabria, B., Zhao, J., Lin, Z., Liu, Q.-R., Onaivi, E.S., 2019. Behavioral effects of psychostimulants in mutant mice with cell-type specific deletion of CB2 cannabinoid receptors in dopamine neurons. Behav Brain Res 360, 286-297.
https://doi.org/10.1016/j.bbr.2018.11.043

Cao, Q., Yang, F., Wang, H., 2021. CB2R induces a protective response against epileptic seizures through ERK and p38 signaling pathways. International Journal of Neuroscience 131, 735-744.
https://doi.org/10.1080/00207454.2020.1796661

Carlisle, S.J., Marciano-Cabral, F., Staab, A., Ludwick, C., Cabral, G.A., 2002. Differential expression of the CB2 cannabinoid receptor by rodent macrophages and macrophage-like cells in relation to cell activation. International Immunopharmacology 2, 69-82.
https://doi.org/10.1016/S1567-5769(01)00147-3

Chaperon, F., Soubrié, P., Puech, A.J., Thiébot, M.-H., 1998. Involvement of central cannabinoid (CB 1) receptors in the establishment of place conditioning in rats. Psychopharmacology 135, 324-332.
https://doi.org/10.1007/s002130050518

Chen, L., Qi, H., Jiang, D., Wang, R., Chen, A., Yan, Z., Xiao, J., 2013. The New Use of an Ancient Remedy: A Double-Blinded Randomized Study on the Treatment of Rheumatoid Arthritis. Am. J. Chin. Med. 41, 263-280.
https://doi.org/10.1142/S0192415X13500195

Chen, X., Cowan, A., Inan, S., Geller, E.B., Meissler, J.J., Rawls, S.M., Tallarida, R.J., Tallarida, C.S., Watson, M.N., Adler, M.W., Eisenstein, T.K., 2019. Opioid-sparing effects of cannabinoids on morphine analgesia: participation of CB1 and CB2 receptors. British Journal of Pharmacology 176, 3378-3389.
https://doi.org/10.1111/bph.14769

Chung, Y.C., Shin, W.-H., Baek, J.Y., Cho, E.J., Baik, H.H., Kim, S.R., Won, S.-Y., Jin, B.K., 2016. CB2 receptor activation prevents glial-derived neurotoxic mediator production, BBB leakage and peripheral immune cell infiltration and rescues dopamine neurons in the MPTP model of Parkinson's disease. Exp Mol Med 48, e205.
https://doi.org/10.1038/emm.2015.100

Correa, F., Hernangómez, M., Mestre, L., Loría, F., Spagnolo, A., Docagne, F., Marzo, V.D., Guaza, C., 2010. Anandamide enhances IL-10 production in activated microglia by targeting CB2 receptors: Roles of ERK1/2, JNK, and NF-κB. Glia 58, 135-147.
https://doi.org/10.1002/glia.20907

Correa, F., Mestre, L., Docagne, F., Guaza, C., 2005. Activation of cannabinoid CB2 receptor negatively regulates IL-12p40 production in murine macrophages: role of IL-10 and ERK1/2 kinase signaling. Br J Pharmacol 145, 441-448.
https://doi.org/10.1038/sj.bjp.0706215

Cortez, I.L., Silva, N.R., Rodrigues, N.S., Pedrazzi, J.F.C., Del Bel, E.A., Mechoulam, R., Gomes, F.V., Guimarães, F.S., 2022. HU-910, a CB2 receptor agonist, reverses behavioral changes in pharmacological rodent models for schizophrenia. Progress in Neuro-Psychopharmacology and Biological Psychiatry 117, 110553.
https://doi.org/10.1016/j.pnpbp.2022.110553

Cristino, L., Bisogno, T., Di Marzo, V., 2020. Cannabinoids and the expanded endocannabinoid system in neurological disorders. Nat Rev Neurol 16, 9-29.
https://doi.org/10.1038/s41582-019-0284-z

Croxford, J.L., Miller, S.D., 2003. Immunoregulation of a viral model of multiple sclerosis using the synthetic cannabinoid R(+)WIN55,212. J Clin Invest 111, 1231-1240.
https://doi.org/10.1172/JCI200317652

Davis, E.J., Foster, T.D., Thomas, W.E., 1994. Cellular forms and functions of brain microglia. Brain Research Bulletin 34, 73-78.
https://doi.org/10.1016/0361-9230(94)90189-9

de Carvalho, C.R., Hoeller, A.A., Franco, P.L.C., Martini, A.P.S., Soares, F.M.S., Lin, K., Prediger, R.D., Whalley, B.J., Walz, R., 2016. The cannabinoid CB2 receptor-specific agonist AM1241 increases pentylenetetrazole-induced seizure severity in Wistar rats. Epilepsy Research 127, 160-167.
https://doi.org/10.1016/j.epilepsyres.2016.08.011

de Lago, E., Fernández-Ruiz, J., Ortega-Gutiérrez, S., Viso, A., López-Rodríguez, M.L., Ramos, J.A., 2002. UCM707, a potent and selective inhibitor of endocannabinoid uptake, potentiates hypokinetic and

antinociceptive effects of anandamide. Eur. J. Pharmacol. 449, 99-103.
https://doi.org/10.1016/s0014-2999(02)01996-9

Delis, F., Polissidis, A., Poulia, N., Justinova, Z., Nomikos, G.G., Goldberg, S.R., Antoniou, K., 2016. Attenuation of Cocaine-Induced Conditioned Place Preference and Motor Activity via Cannabinoid CB2 Receptor Agonism and CB1 Receptor Antagonism in Rats. Int J Neuropsychopharmacol 20, 269-278.
https://doi.org/10.1093/ijnp/pyw102

den Boon, F.S., Chameau, P., Houthuijs, K., Bolijn, S., Mastrangelo, N., Kruse, C.G., Maccarrone, M., Wadman, W.J., Werkman, T.R., 2014. Endocannabinoids produced upon action potential firing evoke a Cl-current via type-2 cannabinoid receptors in the medial prefrontal cortex. Pflugers Arch - Eur J Physiol 466, 2257-2268.
https://doi.org/10.1007/s00424-014-1502-6

den Boon, F.S., Chameau, P., Schaafsma-Zhao, Q., van Aken, W., Bari, M., Oddi, S., Kruse, C.G., Maccarrone, M., Wadman, W.J., Werkman, T.R., 2012. Excitability of prefrontal cortical pyramidal neurons is modulated by activation of intracellular type-2 cannabinoid receptors. Proc Natl Acad Sci U S A 109, 3534-3539.
https://doi.org/10.1073/pnas.1118167109

Du, J.-J., Liu, Z.-Q., Yan, Y., Xiong, J., Jia, X.-T., Di, Z.-L., Ren, J.-J., 2020. The Cannabinoid WIN 55,212-2 Reduces Delayed Neurologic Sequelae After Carbon Monoxide Poisoning by Promoting Microglial M2 Polarization Through ST2 Signaling. J Mol Neurosci 70, 422-432.
https://doi.org/10.1007/s12031-019-01429-2

Ehrhart, J., Obregon, D., Mori, T., Hou, H., Sun, N., Bai, Y., Klein, T., Fernandez, F., Tan, J., Shyle, R., 2005. Stimulation of cannabinoid receptor 2 (CB2) suppresses microglial activation. J Neuroinflammation 2, 29.
https://doi.org/10.1186/1742-2094-2-29

Elliott, M.B., Tuma, R.F., Amenta, P.S., Barbe, M.F., Jallo, J.I., 2011. Acute Effects of a Selective Cannabinoid-2 Receptor Agonist on Neuroinflammation in a Model of Traumatic Brain Injury. Journal of Neurotrauma 28, 973-981.
https://doi.org/10.1089/neu.2010.1672

Elmes, S.J.R., Jhaveri, M.D., Smart, D., Kendall, D.A., Chapman, V., 2004. Cannabinoid CB2 receptor activation inhibits mechanically evoked responses of wide dynamic range dorsal horn neurons in naïve rats and in rat models of inflammatory and neuropathic pain. European Journal of Neuroscience 20, 2311-2320.
https://doi.org/10.1111/j.1460-9568.2004.03690.x

Espejo-Porras, F., García-Toscano, L., Rodríguez-Cueto, C., Santos-García, I., de Lago, E., Fernandez-Ruiz, J., 2019. Targeting glial cannabinoid CB2 receptors to delay the progression of the pathological phenotype in TDP-43 (A315T) transgenic mice, a model of amyotrophic lateral sclerosis. Br J Pharmacol 176, 1585-1600.
https://doi.org/10.1111/bph.14216

Esposito, G., Iuvone, T., Savani, C., Scuderi, C., De Filippis, D., Papa, M., Di Marzo, V., Steardo, L., 2007. Opposing Control of Cannabinoid Receptor Stimulation on Amyloid-β-Induced Reactive Gliosis: In Vitro and in Vivo Evidence. I Pharmacol Exp Ther 322, 1144-1152.
https://doi.org/10.1124/jpet.107.121566

Facchinetti, F., Giudice, E.D., Furegato, S., Passarotto, M., Leon, A., 2003. Cannabinoids ablate release of TNFα in rat microglial cells stimulated with lyopolysaccharide. Glia 41, 161-168.
https://doi.org/10.1002/glia.10177

Fakhouri, G., Ahmadiani, A., Rahimian, R., Grolla, A.A., Moradi, F., Haeri, A., 2012. WIN55212-2 attenuates amyloid-beta-induced neuroinflammation in rats through activation of cannabinoid receptors and PPAR-γ pathway. Neuropharmacology 63, 653-666.
https://doi.org/10.1016/j.neuropharm.2012.05.013

Fani Maleki, A., Rivest, S., 2019. Innate Immune Cells: Monocytes, Monocyte-Derived Macrophages and Microglia as Therapeutic Targets for Alzheimer's Disease and Multiple Sclerosis. Frontiers in Cellular Neuroscience 13.

Felder, C.C., Joyce, K.E., Briley, E.M., Mansouri, J., Mackie, K., Blond, O., Lai, Y., Ma, A.L., Mitchell, R.L., 1995. Comparison of the pharmacology and signal transduction of the human cannabinoid CB1 and CB2 receptors. Mol. Pharmacol. 48, 443-450.

Feng, L., Lo, H., You, H., Wu, W., Cheng, X., Xin, J., Ye, Z., Chen, X., Pan, X., 2022. Loss of cannabinoid receptor 2 promotes α-Synuclein-induced
microglial synaptic pruning in nucleus accumbens by modulating the pCREB-c-Fos signaling pathway and complement system. Exp Neurol 359, 114230. https://doi.org/10.1016/j.expneurol.2022.114230

Foss, J., Naguib, M., Giordano, T., 2020. A phase I single ascending dose safety study of NTRX-07 in normal volunteers. Alzheimer's & Dementia 16, e039150. https://doi.org/10.1002/alz.039150

Franklin, J.M., Carrasco, G.A., 2012. Cannabinoid-Induced Enhanced Interaction and Protein Levels of Serotonin 5-HT2A and Dopamine D2 Receptors in Rat Prefrontal Cortex. J Psychopharmacol 26, 1333-1347. https://doi.org/10.1177/0269881112450786

Galán-Ganga, M., Rodríguez-Cueto, C., Merchán-Rubira, J., Hernández, F., Ávila, J., Posada-Ayala, M., Lanciego, J.L., Luengo, E., Lopez, M.G., Rábano, A., Fernández-Ruiz, J., Lastres-Becker, I., 2021. Cannabinoid receptor CB2 ablation protects against TAU induced neurodegeneration. acta neuropathol commun 9, 90. https://doi.org/10.1186/s40478-021-01196-5

Galiègue, S., Mary, S., Marchand, J., Dussossoy, D., Carrière, D., Carayon, P., Bouaboula, M., Shire, D., Fur, G.L., Casellas, P., 1995. Expression of Central and Peripheral Cannabinoid Receptors in Human Immune Tissues and Leukocyte Subpopulations. European Journal of Biochemistry 232, 54-61. https://doi.org/10.1111/j.1432-1033.1995.tb20780.x

Gallant, M., Dufresne, C., Gareau, Y., Guay, D., Leblanc, Y., Prasit, P., Rochette, C., Sawyer, N., Slipetz, D.M., Tremblay, N., Metters, K.M., Labelle, M., 1996. New class of potent ligands for the human peripheral cannabinoid receptor. Bioorganic & Medicinal Chemistry Letters 6, 2263-2268. https://doi.org/10.1016/0960-894X(96)00426-X

Gamaleddin, I., Zvonok, A., Makriyannis, A., Goldberg, S.R., Le Foll, B., 2012. Effects of a selective cannabinoid CB2 agonist and antagonist on intravenous nicotine self administration and reinstatement of nicotine seeking. PLoS One 7, e29900. https://doi.org/10.1371/journal.pone.0029900

García-Gutiérrez, M., Pérez-Ortiz, J., Gutiérrez-Adán, A., Manzanares, J., 2010. Depression-resistant endophenotype in mice overexpressing cannabinoid CB2 receptors. Br J Pharmacol 160, 1773-1784. https://doi.org/10.1111/j.1476-5381.2010.00819.x

García-Gutiérrez, M.S., García-Bueno, B., Zoppi, S., Leza, J.C., Manzanares, J., 2012. Chronic blockade of cannabinoid CB2 receptors induces anxiolytic-like actions associated with alterations in GABAA receptors. Br J Pharmacol 165, 951-964. https://doi.org/10.1111/j.1476-5381.2011.01625.x

García-Gutiérrez, M.S., Manzanares, J., 2011. Overexpression of CB2 cannabinoid receptors decreased vulnerability to anxiety and impaired anxiolytic action of alprazolam in mice. J Psychopharmacol 25, 111-120. https://doi.org/10.1177/0269881110379507

García-Gutiérrez, M.S., Ortega-Álvaro, A., Busquets-García, A., Pérez-Ortiz, J.M., Caltana, L., Ricatti, M.J., Brusco, A., Maldonado, R., Manzanares, J., 2013. Synaptic plasticity alterations associated with memory impairment induced by deletion of CB2 cannabinoid receptors. Neuropharmacology 73, 388-396. https://doi.org/10.1016/j.neuropharm.2013.05.034

Gertsch, J., Leonti, M., Raduner, S., Racz, I., Chen, J.-Z., Xie, X.-Q., Altmann, K.-H., Karsak, M., Zimmer, A., 2008. Beta-caryophyllene is a dietary cannabinoid. Proc Natl Acad Sci U S A 105, 9099-9104. https://doi.org/10.1073/pnas.0803601105

Ghanbari, M.-M., Joneidi, M., Kiani, B., Babaie, J., Sayyah, M., 2020. Cannabinoid receptors and the proconvulsant effect of toxoplasmosis in mice. Microbial Pathogenesis 144, 104204. https://doi.org/10.1016/j.micpath.2020.104204

Gómez-Gálvez, Y., Palomo-Garo, C., Fernández-Ruiz, J., García, C., 2016. Potential of the cannabinoid CB2 receptor as a pharmacological target against inflammation in Parkinson's disease. Progress in Neuro-Psychopharmacology and Biological Psychiatry 64, 200-208. https://doi.org/10.1016/j.pnpbp.2015.03.017

Gonzalez-Cuevas, G., Martin-Fardon, R., Kerr, T.M., Stouffer, D.G., Parsons, L.H., Hammell, D.C., Banks, S.L., Stinchcomb, A.L., Weiss, F., 2018. Unique treatment potential of cannabidiol for the prevention of relapse to drug use: preclinical proof of principle. Neuropsychopharmacology 43, 2036-2045. https://doi.org/10.1038/s41386-018-0050-8

Gorantla, S., Makarov, E., Roy, D., Finke-Dwyer, J., Murrin, L.C., Gendelman, H.E., Poluektova, L., 2010. Immunoregulation of a CB2 Receptor Agonist in a Murine Model of NeuroAIDS. J Neuroimmune Pharmacol 5, 456-468. https://doi.org/10.1007/s11481-010-9225-8

Grabon, W., Bodennec, J., Rheims, S., Belmeguenai, A., Bezin, L., 2023. Update on the controversial identity of cells expressing cnr2 gene in the nervous system. CNS Neurosci Ther 1-11. https://doi.org/10.1111/cns.13977

Grenald, S.A., Young, M.A., Wang, Y., Ossipov, M.H., Ibrahim, M.M., Largent-Milnes, T.M., Vanderah, T.W., 2017. Synergistic attenuation of chronic pain using mu opioid and cannabinoid receptor 2 agonists. Neuropharmacology 116, 59-70. https://doi.org/10.1016/j.neuropharm.2016.12.008

Hanus, L., Breuer, A., Tchilibon, S., Shiloah, S., Goldenberg, D., Horowitz, M., Pertwee, R.G., Ross, R.A., Mechoulam, R., Fride, E., 1999. HU-308: a specific agonist for CB(2), a peripheral cannabinoid receptor. Proc. Natl. Acad. Sci. U.S.A. 96, 14228-14233. https://doi.org/10.1073/pnas.96.25.14228

He, X., Yang, L., Huang, R., Lin, L., Shen, Y., Cheng, L., Jin, L., Wang, S., Zhu, R., 2020. Activation of CB2R with AM1241 ameliorates neurodegeneration via the Xist/miR-133b-3p/Pitx3 axis. Journal of Cellular Physiology 235, 6032-6042. https://doi.org/10.1002/jcp.29530

Horváth, B., Magid, L., Mukhopadhyay, P., Bátkai, S., Rajesh, M., Park, O., Tanchian, G., Gao, R.Y., Goodfellow, C.E., Glass, M., Mechoulam, R., Pacher, P., 2012. A new cannabinoid CB2 receptor agonist HU-910 attenuates oxidative stress, inflammation and cell death associated with hepatic ischaemia/reperfusion injury. Br J Pharmacol 165, 2462-2478. https://doi.org/10.1111/j.1476-5381.2011.01381.x

Howlett, A.C., 2002. International Union of Pharmacology. XXVII. Classification of Cannabinoid Receptors. Pharmacological Reviews 54, 161-202. https://doi.org/10.1124/pr.54.2.161

Howlett, A.C., 1998. The CB1Cannabinoid Receptor in the Brain. Neurobiology of Disease 5, 405-416. https://doi.org/10.1006/nbdi.1998.0215

Howlett, A.C., Abood, M.E., 2017. CB1 & CB2 Receptor Pharmacology. Adv Pharmacol 80, 169-206. https://doi.org/10.1016/bs.apha.2017.03.007

Huffman, J.W., Liddle, J., Yu, S., Aung, M.M., Abood, M.E., Wiley, J.L., Martin, B.R., 1999. 3-(1',1'-Dimethylbutyl)-1-deoxy-delta8-THC and related compounds: synthesis of selective ligands for the CB2 receptor. Bioorg. Med. Chem. 7, 2905-2914. https://doi.org/10.1016/s0968-0896(99)00219-9

Huizenga, M.N., Wicker, E., Beck, V.C., Forcelli, P.A., 2017. Anticonvulsant effect of cannabinoid receptor agonists in models of seizures in developing rats. Epilepsia 58, 1593-1602. https://doi.org/10.1111/epi.13842

Hwang, E.-S., Kim, H.-B., Lee, S., Kim, M.-J., Kim, K.-J., Han, G., Han, S.-Y., Lee, E.-A., Yoon, J.-H., Kim, D.-O., Maeng, S., Park, J.-H., 2020. Antidepressant-like effects of β-caryophyllene on restraint plus stress-induced depression. Behavioural Brain Research 380, 112439. https://doi.org/10.1016/j.bbr.2019.112439

Iannotti, F.A., Di Marzo, V., Petrosino, S., 2016. Endocannabinoids and endocannabinoid-related mediators: Targets, metabolism and role in neurological disorders. Progress in Lipid Research 62, 107-128. https://doi.org/10.1016/j.plipres.2016.02.002

Ibrahim, M.M., Deng, H., Zvonok, A., Cockayne, D.A., Kwan, J., Mata, H.P., Vanderah, T.W., Lai, J., Porreca, F., Makriyannis, A., Malan, T.P., 2003. Activation of CB2 cannabinoid receptors by AM1241 inhibits experimental neuropathic pain: pain inhibition by receptors not present in the CNS. Proc. Natl. Acad. Sci. U.S.A. 100, 10529-10533. https://doi.org/10.1073/pnas.1834309100

Ignatowska-Jankowska, B.M., Muldoon, P.P., Lichtman, A.H., Damaj, M.I., 2013. The cannabinoid CB2 receptor is necessary for nicotine-conditioned place preference, but not other behavioral effects of nicotine in mice. Psychopharmacology (Berl) 229, 591-601. https://doi.org/10.1007/s00213-013-3117-6

Ishiguro, H., Carpio, O., Horiuchi, Y., Shu, A., Higuchi, S., Schanz, N., Benno, R., Arinami, T., Onaivi, E.S., 2010. A nonsynonymous polymorphism in cannabinoid CB2 receptor gene is associated with eating disorders in
humans and food intake is modified in mice by its ligands. Synapse 64, 92-96. https://doi.org/10.1002/syn.20714

Ishiguro, H., Iwasaki, S., Teasenfitz, L., Higuchi, S., Horiuchi, Y., Saito, T., Arinami, T., Onaivi, E.S., 2007. Involvement of cannabinoid CB2 receptor in alcohol preference in mice and alcoholism in humans. Pharmacogenomics 7, 380-385. https://doi.org/10.1038/sj.tpj.6500431

Iwamura, H., Suzuki, H., Ueda, Y., Kaya, T., Inaba, T., 2001. In Vitro and in Vivo Pharmacological Characterization of JTE-907, a Novel Selective Ligand for Cannabinoid CB2 Receptor 6.

Javed, H., Azimullah, S., Haque, M.E., Ojha, S.K., 2016. Cannabinoid Type 2 (CB2) Receptors Activation Protects against Oxidative Stress and Neuroinflammation Associated Dopaminergic Neurodegeneration in Rotenone Model of Parkinson's Disease. Front. Neurosci. 10. https://doi.org/10.3389/fnins.2016.00321

Jayant, S., Sharma, B.M., Bansal, R., Sharma, B., 2016. Pharmacological benefits of selective modulation of cannabinoid receptor type 2 (CB2) in experimental Alzheimer's disease. Pharmacology Biochemistry and Behavior 140, 39-50. https://doi.org/10.1016/j.pbb.2015.11.006

Jhaveri, M.D., Elmes, S.J.R., Richardson, D., Barrett, D.A., Kendall, D.A., Mason, R., Chapman, V., 2008. Evidence for a novel functional role of cannabinoid CB2 receptors in the thalamus of neuropathic rats. Eur J Neurosci 27, 1722-1730. https://doi.org/10.1111/j.1460-9568.2008.06162.x

Jia, Y., Deng, H., Qin, Q., Ma, Z., 2020. JWH133 inhibits MPP+-induced inflammatory response and iron influx in astrocytes. Neurosci Lett 720, 134779. https://doi.org/10.1016/j.neulet.2020.134779

Jing, N., Fang, B., Li, Z., Tian, A., 2020. Exogenous activation of cannabinoid-2 receptor modulates TLR4/MMP9 expression in a spinal cord ischemia reperfusion rat model. J Neuroinflammation 17. https://doi.org/10.1186/s12974-020-01784-7

Jordan, C.J., Xi, Z.-X., 2019. Progress in Brain Cannabinoid CB2 Receptor Research: From Genes to Behavior. Neurosci Biobehav Rev 98, 208-220. https://doi.org/10.1016/j.neubiorev.2018.12.026

Kano, M., Ohno-Shosaku, T., Hashimotodani, Y., Uchigashima, M., Watanabe, M., 2009. Endocannabinoid-mediated control of synaptic transmission. Physiol Rev 89, 309-380. https://doi.org/10.1152/physrev.00019.2008

Karan, A.A., Spivak, Y.S., Gerasimov, K.A., Suleymanova, E.M., Volobueva, M.N., Kvichansky, A.A., Vinogradova, L.V., Bolshakov, A.P., 2021. CB2 receptors modulate seizure-induced expression of pro-inflammatory cytokines in the hippocampus but not neocortex. Mol Neurobiol 58, 4028-4037. https://doi.org/10.1007/s12035-021-02395-w

Kibret, B.G., Ishiguro, H., Horiuchi, Y., Onaivi, E.S., 2022. New Insights and Potential Therapeutic Targeting of CB2 Cannabinoid Receptors in CNS Disorders. Int J Mol Sci 23, 975. https://doi.org/10.3390/ijms23020975

Kim, J., Li, Y., 2015. Chronic activation of CB2 cannabinoid receptors in the hippocampus increases excitatory synaptic transmission: Synaptic regulation mediated by CB2 cannabinoid receptors. J Physiol 593, 871-886. https://doi.org/10.1113/jphysiol.2014.286633

Klauke, A.-L., Racz, I., Pradier, B., Markert, A., Zimmer, A.M., Gertsch, J., Zimmer, A., 2014. The cannabinoid CB2 receptor-selective phytocannabinoid beta-caryophyllene exerts analgesic effects in mouse models of inflammatory and neuropathic pain. European Neuropsychopharmacology 24, 608-620. https://doi.org/10.1016/j.euroneuro.2013.10.008

Klegeris, A., Bissonnette, C.J., McGeer, P.L., 2003. Reduction of human monocytic cell neurotoxicity and cytokine secretion by ligands of the cannabinoid-type CB2 receptor. Br J Pharmacol 139, 775-786. https://doi.org/10.1038/sj.bjp.0705304

Komorowska-Müller, J.A., Rana, T., Olabiyi, B.F., Zimmer, A., Schmöle, A.-C., 2021a. Cannabinoid Receptor 2 Alters Social Memory and Microglial Activity in an Age-Dependent Manner. Molecules 26, 5984. https://doi.org/10.3390/molecules26195984

Komorowska-Müller, J.A., Ravichandran, K.A., Zimmer, A., Schürmann, B., 2021b. Cannabinoid receptor 2 deletion influences social memory and

synaptic architecture in the hippocampus. Sci Rep 11, 16828. https://doi.org/10.1038/s41598-021-96285-9

Komorowska-Müller, J.A., Schmöle, A.-C., 2020. CB2 Receptor in Microglia: The Guardian of Self-Control. Int J Mol Sci 22. https://doi.org/10.3390/ijms22010019

Kong, W., Li, H., Tuma, R., Ganea, D., 2014a. Selective CB2 receptor agonist Gp1a attenuates EAE through modulating CD4 T cell differentiation and immune cell infiltration in the CNS (BA8P.131). The Journal of Immunology 192, 113.14-113.14.

Kong, W., Li, H., Tuma, R.F., Ganea, D., 2014b. Selective CB2 receptor activation ameliorates EAE by reducing Th17 differentiation and immune cell accumulation in the CNS. Cell Immunol 287, 1-17. https://doi.org/10.1016/j.cellimm.2013.11.002

Koppel, J., Vingtdeux, V., Marambaud, P., d'Abramo, C., Jimenez, H., Stauber, M., Friedman, R., Davies, P., 2014. CB2 Receptor Deficiency Increases Amyloid Pathology and Alters Tau Processing in a Transgenic Mouse Model of Alzheimer's Disease. Mol Med 20, 29-36. https://doi.org/10.2119/molmed.2013.00140.revised

Kossatz, E., Maldonado, R., Robledo, P., 2016. CB2 cannabinoid receptors modulate HIF-1α and TIM-3 expression in a hypoxia-ischemia mouse model. European Neuropsychopharmacology 26, 1972-1988. https://doi.org/10.1016/j.euroneuro.2016.10.003

Kow, R.L., Jiang, K., Naydenov, A.V., Le, J.H., Stella, N., Nathanson, N.M., 2014. Modulation of Pilocarpine-Induced Seizures by Cannabinoid Receptor 1. PLoS One 9. https://doi.org/10.1371/journal.pone.0095922

Kruk-Slomka, M., Boguszewska-Czubara, A., Slomka, T., Budzynska, B., Biala, G., 2016. Correlations between the Memory-Related Behavior and the Level of Oxidative Stress Biomarkers in the Mice Brain, Provoked by an Acute Administration of CB Receptor Ligands. Neural Plast 2016, 9815092. https://doi.org/10.1155/2016/9815092

Kruk-Slomka, M., Dzik, A., Biala, G., 2022. The Influence of CB2-Receptor Ligands on the Memory-Related Responses in Connection with Cholinergic Pathways in Mice in the Passive Avoidance Test. Molecules 27, 4252. https://doi.org/10.3390/molecules27134252

Latini, L., Bisicchia, E., Sasso, V., Chiurchiù, V., Cavallucci, V., Molinari, M., Maccarrone, M., Viscomi, M.T., 2014. Cannabinoid CB2 receptor (CB2R) stimulation delays rubrospinal mitochondrial-dependent degeneration and improves functional recovery after spinal cord hemisection by ERK1/2 inactivation. Cell Death & Disease 5, e1404. https://doi.org/10.1038/cddis.2014.364

Li, C., Shi, J., Sun, J., Shi, Y., Jia, H., 2021. Cannabinoid receptor 2 deficiency enhances isoflurane-induced spatial cognitive impairment in adult mice by affecting neuroinflammation, neurogenesis and neuroplasticity. Exp Ther Med 22, 908. https://doi.org/10.3892/etm.2021.10340

Li, C., Shi, J., Wang, B., Li, J., Jia, H., 2019a. CB2 cannabinoid receptor agonist ameliorates novel object recognition but not spatial memory in transgenic APP/PS1 mice. Neuroscience Letters 707, 134286. https://doi.org/10.1016/j.neulet.2019.134286

Li, C., Shi, J., Wang, B., Li, J., Jia, H., 2019b. CB2 cannabinoid receptor agonist ameliorates novel object recognition but not spatial memory in transgenic APP/PS1 mice. Neuroscience Letters 707, 134286. https://doi.org/10.1016/j.neulet.2019.134286

Li, L., Luo, Q., Shang, B., Yang, X., Zhang, Y., Pan, Q., Wu, N., Tang, W., Du, D., Sun, X., Jiang, L., 2022. Selective activation of cannabinoid receptor-2 reduces white matter injury via PERK signaling in a rat model of traumatic brain injury. Experimental Neurology 347, 113899. https://doi.org/10.1016/j.expneurol.2021.113899

Li, L., Yun, D., Zhang, Y., Tao, Y., Tan, Q., Qiao, F., Luo, B., Liu, Y., Fan, R., Xian, J., Yu, A., 2018. A cannabinoid receptor 2 agonist reduces blood-brain barrier damage via induction of MKP-1 after intracerebral hemorrhage in rats. Brain Research 1697, 113-123. https://doi.org/10.1016/j.brainres.2018.06.006

Li, Y., Kim, J., 2017. Distinct roles of neuronal and microglial CB2 cannabinoid receptors in the mouse hippocampus. Neuroscience 363, 11-25. https://doi.org/10.1016/j.neuroscience.2017.08.053

Li, Y., Kim, J., 2016a. Deletion of CB2 cannabinoid receptors reduces
synaptic transmission and long-term potentiation in the mouse
hippocampus. Hippocampus 26, 275-281.
https://doi.org/10.1002/hipo.22558

Li, Y., Kim, J., 2016b. CB2 Cannabinoid Receptor Knockout in Mice Impairs Contextual Long-Term Memory and Enhances Spatial Working Memory. Neural Plast 2016. https://doi.org/10.1155/2016/9817089

Lin, L., Yihao, T., Zhou, F., Yin, N., Qiang, T., Haowen, Z., Qianwei, C., Jun, T., Yuan, Z., Gang, Z., Hua, F., Yunfeng, Y., Zhi, C., 2017. Inflammatory Regulation by Driving Microglial M2 Polarization: Neuroprotective Effects of Cannabinoid Receptor-2 Activation in Intracerebral Hemorrhage. Front Immunol 8. https://doi.org/10.3389/fimmu.2017.00112

Lindsey, L.P., Daphney, C.M., Oppong-Damoah, A., Uchakin, P.N., Abney, S.E., Uchakina, O.N., Khusial, R.D., Akil, A., Murnane, K.S., 2019. The cannabinoid receptor 2 agonist, β-caryophyllene, improves working memory and reduces circulating levels of specific proinflammatory cytokines in aged male mice. Behav Brain Res 372, 112012. https://doi.org/10.1016/j.bbr.2019.112012

Liu, Q.-R., Canseco-Alba, A., Liang, Y., Ishiguro, H., Onaivi, E.S., 2020. Low Basal CB2R in Dopamine Neurons and Microglia Influences Cannabinoid Tetrad Effects. Int J Mol Sci 21, 9763. https://doi.org/10.3390/ijms21249763

Liu, Q.-R., Canseco-Alba, A., Zhang, H.-Y., Tagliaferro, P., Chung, M., Dennis, E., Sanabria, B., Schanz, N., Escosteguy-Neto, J.C., Ishiguro, H., Lin, Z., Sgro, S., Leonard, C.M., Santos-Junior, J.G., Gardner, E.L., Egan, J.M., Lee, J.W., Xi, Z.-X., Onaivi, E.S., 2017. Cannabinoid type 2 receptors in dopamine neurons inhibits psychomotor behaviors, alters anxiety, depression and alcohol preference. Sci Rep 7. https://doi.org/10.1038/s41598-017-17796-y

Lopes, J.B., Bastos, J.R., Costa, R.B., Aguiar, D.C., Moreira, F.A., 2020. The roles of cannabinoid CB1 and CB2 receptors in cocaine-induced behavioral sensitization and conditioned place preference in mice. Psychopharmacology (Berl.) 237, 385-394. https://doi.org/10.1007/s00213-019-05370-5

López, A., Aparicio, N., Pazos, M.R., Grande, M.T., Barreda-Manso, M.A., Benito-Cuesta, I., Vázquez, C., Amores, M., Ruiz-Pérez, G., García-García, E., Beatka, M., Tolón, R.M., Dittel, B.N., Hillard, C.J., Romero, J., 2018. Cannabinoid CB2 receptors in the mouse brain: relevance for Alzheimer's disease. J Neuroinflammation 15. https://doi.org/10.1186/s12974-018-1174-9

López-Ramírez, G., Sánchez-Zavaleta, R., Ávalos-Fuentes, A., José Sierra, J., Paz-Bermúdez, F., Leyva-Gómez, G., Segovia Vila, J., Cortés, H., Florán, B., 2020. D2 autoreceptor switches CB2 receptor effects on [3H]-dopamine release in the striatum. Synapse 74, e22139. https://doi.org/10.1002/syn.22139

Lou, J., Teng, Z., Zhang, L., Yang, J., Ma, L., Wang, F., Tian, X., An, R., Yang, M., Zhang, Q., Xu, L., Dong, Z., 2017. β-Caryophyllene/Hydroxypropyl-β-Cyclodextrin Inclusion Complex Improves Cognitive Deficits in Rats with Vascular Dementia through the Cannabinoid Receptor Type 2 -Mediated Pathway. Front Pharmacol 8, 2. https://doi.org/10.3389/fphar.2017.00002

Luo, X.-Q., Li, A., Yang, X., Xiao, X., Hu, R., Wang, T.-W., Dou, X.-Y., Yang, D.-J., Dong, Z., 2018a. Paeoniflorin exerts neuroprotective effects by modulating the M1/M2 subset polarization of microglia/macrophages in the hippocampal CA1 region of vascular dementia rats via cannabinoid receptor 2. Chin Med 13. https://doi.org/10.1186/s13020-018-0173-1

Luo, X.-Q., Li, A., Yang, X., Xiao, X., Hu, R., Wang, T.-W., Dou, X.-Y., Yang, D.-J., Dong, Z., 2018b. Paeoniflorin exerts neuroprotective effects by modulating the M1/M2 subset polarization of microglia/macrophages in the hippocampal CA1 region of vascular dementia rats via cannabinoid receptor 2. Chin Med 13. https://doi.org/10.1186/s13020-018-0173-1

Ma, L., Jia, J., Liu, X., Bai, F., Wang, Q., Xiong, L., 2015. Activation of murine microglial N9 cells is attenuated through cannabinoid receptor CB2 signaling. Biochemical and Biophysical Research Communications 458, 92-97. https://doi.org/10.1016/j.bbrc.2015.01.073

Ma, Z., Gao, F., Larsen, B., Gao, M., Luo, Z., Chen, D., Ma, X., Qiu, S., Zhou, Y., Xie, J., Xi, Z.-X., Wu, J., 2019. Mechanisms of cannabinoid CB2 receptor-mediated reduction of dopamine neuronal excitability in mouse ventral tegmental area. EBioMedicine 42, 225-237.

https://doi.org/10.1016/j.ebiom.2019.03.040

Mackie, K., 2005. Distribution of Cannabinoid Receptors in the Central and Peripheral Nervous System, in: Pertwee, R.G. (Ed.), Cannabinoids, Handbook of Experimental Pharmacology. Springer, Berlin, Heidelberg, pp. 299-325. https://doi.org/10.1007/3-540-26573-2_10

Magid, L., Heymann, S., Elgali, M., Avram, L., Cohen, Y., Liraz-Zaltsman, S., Mechoulam, R., Shohami, E., 2019. Role of CB2 Receptor in the Recovery of Mice after Traumatic Brain Injury. J Neurotrauma 36, 1836-1846. https://doi.org/10.1089/neu.2018.6063

Malek, N., Popiolek-Barczyk, K., Mika, J., Przewlocka, B., Starowicz, K., 2015. Anandamide, Acting via CB2 Receptors, Alleviates LPS-Induced Neuroinflammation in Rat Primary Microglial Cultures. Neural Plast 2015. https://doi.org/10.1155/2015/130639

Manera, C., Benetti, V., Castelli, M.P., Cavallini, T., Lazzarotti, S., Pibiri, F., Saccomanni, G., Tuccinardi, T., Vannacci, A., Martinelli, A., Ferrarini, P.L., 2006. Design, Synthesis, and Biological Evaluation of New 1,8-Naphthyridin-4(1 H)-on-3-carboxamide and Quinolin-4(1 H)-on-3-carboxamide Derivatives as CB₂ Selective Agonists. J. Med. Chem. 49, 5947-5957. https://doi.org/10.1021/jm0603466

Maresz, K., Carrier, E.J., Ponomarev, E.D., Hillard, C.J., Dittel, B.N., 2005. Modulation of the cannabinoid CB2 receptor in microglial cells in response to inflammatory stimuli. Journal of Neurochemistry 95, 437-445. https://doi.org/10.1111/j.1471-4159.2005.03380.x

Marinelli, S., Basilico, B., Marrone, M.C., Ragozzino, D., 2019. Microglia-neuron crosstalk: Signaling mechanism and control of synaptic transmission. Seminars in Cell & Developmental Biology, SI: Calcium signalling 94, 138-151. https://doi.org/10.1016/j semcdb.2019.05.017

Martín-Moreno, A.M., Brera, B., Spuch, C., Carro, E., García-García, L., Delgado, M., Pozo, M.A., Innamorato, N.G., Cuadrado, A., de Ceballos, M.L., 2012. Prolonged oral cannabinoid administration prevents neuroinflammation, lowers β-amyloid levels and improves cognitive performance in Tg APP 2576 mice. J Neuroinflammation 9, 8. https://doi.org/10.1186/1742-2094-9-8

Martín-Sánchez, A., Warnault, V., Montagud-Romero, S., Pastor, A., Mondragón, N., De La Torre, R., Valverde, O., 2019. Alcohol-induced conditioned place preference is modulated by CB2 cannabinoid receptors and modifies levels of endocannabinoids in the mesocorticolimbic system. Pharmacology Biochemistry and Behavior 183, 22-31. https://doi.org/10.1016/j.pbb.2019.06.007

Mecha, M., Feliú, A., Carrillo-Salinas, F.J., Rueda-Zubiaurre, A., Ortega-Gutiérrez, S., de Sola, R.G., Guaza, C., 2015. Endocannabinoids drive the acquisition of an alternative phenotype in microglia. Brain, Behavior, and Immunity 49, 233-245. https://doi.org/10.1016/j.bbi.2015.06.002

Mecha, M., Feliú, A., Machín, I., Cordero, C., Carrillo-Salinas, F., Mestre, L., Hernández-Torres, G., Ortega-Gutiérrez, S., López-Rodríguez, M.L., de Castro, F., Clemente, D., Guaza, C., 2018. 2-AG limits Theiler's virus induced acute neuroinflammation by modulating microglia and promoting MDSCs. Glia 66, 1447-1463. https://doi.org/10.1002/glia.23317

Montecucco, F., Burger, F., Mach, F., Steffens, S., 2008. CB2 cannabinoid receptor agonist JWH-015 modulates human monocyte migration through defined intracellular signaling pathways. Am J Physiol Heart Circ Physiol 294, H1145-1155. https://doi.org/10.1152/ajpheart.01328.2007

Morgan, N.H., Stanford, I.M., Woodhall, G.L., 2009. Functional CB2 type cannabinoid receptors at CNS synapses. Neuropharmacology 57, 356-368. https://doi.org/10.1016/j.neuropharm.2009.07.017

Murikinati, S., Jüttler, E., Keinert, T., Ridder, D.A., Muhammad, S., Waibler, Z., Ledent, C., Zimmer, A., Kalinke, U., Schwaninger, M., 2010. Activation of cannabinoid 2 receptors protects against cerebral ischemia by inhibiting neutrophil recruitment. The FASEB Journal 24, 788-798. https://doi.org/10.1096/fj.09-141275

Murineddu, G., Lazzari, P., Ruiu, S., Sanna, A., Loriga, G., Manca, I., Falzoi, M., Dessì, C., Curzu, M.M., Chelucci, G., Pani, L., Pinna, G.A., 2006. Tricyclic pyrazoles. 4. Synthesis and biological evaluation of analogues of the robust and selective CB2 cannabinoid ligand 1-(2',4'-dichlorophenyl)-6-methyl-N-piperidin-1-yl-1,4-dihydroindeno[1,2-c]pyrazole-3-carboxamide. J. Med. Chem. 49, 7502-7512. https://doi.org/10.1021/jm060920d
Nackley, A.G., Zvonok, A.M., Makriyannis, A., Hohmann, A.G., 2004. Activation of Cannabinoid CB₂ Receptors Suppresses C-Fiber Responses and Windup in Spinal Wide Dynamic Range Neurons in the Absence and Presence of Inflammation. Journal of Neurophysiology 92, 3562-3574. https://doi.org/10.1152/jn.00886.2003

Naguib, M., Diaz, P., Xu, J.J., Astruc-Diaz, F., Craig, S., Vivas-Mejia, P., Brown, D.L., 2008. MDA7: a novel selective agonist for CB2 receptors that prevents allodynia in rat neuropathic pain models. Br J Pharmacol 155, 1104-1116. https://doi.org/10.1038/bjp.2008.340

Nasehi, M., Hajikhani, M., Ebrahimi-Ghiri, M., Zarrindast, M.-R., 2017. Interaction between NMDA and CB2 function in the dorsal hippocampus on memory consolidation impairment: an isobologram analysis. Psychopharmacology 234, 507-514. https://doi.org/10.1007/s00213-016-4481-9

Navarrete, F., García-Gutiérrez, M.S., Aracil-Fernández, A., Lanciego, J.L., Manzanares, J., 2018. Cannabinoid CB1 and CB2 Receptors, and Monoacylglycerol Lipase Gene Expression Alterations in the Basal Ganglia of Patients with Parkinson's Disease. Neurotherapeutics 15, 459-469. https://doi.org/10.1007/s13311-018-0603-x

Navarrete, F., Rodríguez-Arias, M., Martín-García, E., Navarro, D., García-Gutiérrez, M.S., Aguilar, M.A., Aracil-Fernández, A., Berbel, P., Miñarro, J., Maldonado, R., Manzanares, J., 2013. Role of CB2 Cannabinoid Receptors in the Rewarding, Reinforcing, and Physical Effects of Nicotine. Neuropsychopharmacology 38, 2515-2524. https://doi.org/10.1038/npp.2013.157

Negrete, R., Hervera, A., Leánez, S., Martín-Campos, J.M., Pol, O., 2011. The Antinociceptive Effects of JWH-015 in Chronic Inflammatory Pain Are Produced by Nitric Oxide-cGMP-PKG-KATP Pathway Activation Mediated by Opioids. PLoS One 6. https://doi.org/10.1371/journal.pone.0026688

Oddi, S., Latini, L., Viscomi, M.T., Bisicchia, E., Molinari, M., Maccarrone, M., 2012. Distinct regulation of nNOS and iNOS by CB2 receptor in remote delayed neurodegeneration. J Mol Med 90, 371-387. https://doi.org/10.1007/s00109-011-0846-z

Oliveira, C.C. de, Oliveira, C.V. de, Grigoletto, J., Ribeiro, L.R., Funck, V.R., Grauncke, A.C.B., Souza, T.L. de, Souto, N.S., Furian, A.F., Menezes, I.R.A., Oliveira, M.S., 2016. Anticonvulsant activity of β-caryophyllene against pentylenetetrazol-induced seizures. Epilepsy & Behavior 56, 26-31. https://doi.org/10.1016/j.yebeh.2015.12.040

Onaivi, E.S., 2006. Neuropsychobiological Evidence for the Functional Presence and Expression of Cannabinoid CB2 Receptors in the Brain. Neuropsychobiology 54, 231-246. https://doi.org/10.1159/000100778

Onaivi, E.S., Ishiguro, H., Gong, J.-P., Patel, S., Meozzi, P.A., Myers, L., Perchuk, A., Mora, Z., Tagliaferro, P.A., Gardner, E., Brusco, A., Akinshola, B.E., Hope, B., Lujilde, J., Inada, T., Iwasaki, S., Macharia, D., Teasenfitz, L., Arinami, T., Uhl, G.R., 2008. Brain Neuronal CB2 Cannabinoid Receptors in Drug Abuse and Depression: From Mice to Human Subjects. PLoS One 3. https://doi.org/10.1371/journal.pone.0001640

Ortega-Alvaro, A., Aracil-Fernández, A., García-Gutiérrez, M.S., Navarrete, F., Manzanares, J., 2011. Deletion of CB2 Cannabinoid Receptor Induces Schizophrenia-Related Behaviors in Mice. Neuropsychopharmacology 36, 1489-1504. https://doi.org/10.1038/npp.2011.34

Ortega-Alvaro, A., Ternianov, A., Aracil-Fernández, A., Navarrete, F., García-Gutiérrez, M.S., Manzanares, J., 2015. Role of cannabinoid CB2 receptor in the reinforcing actions of ethanol. Addiction Biology 20, 43-55. https://doi.org/10.1111/adb.12076

Ortega-Gutiérrez, S., Molina-Holgado, E., Arévalo-Martín, Á., Correa, F., Viso, A., López-Rodríguez, M.L., Di Marzo, V., Guaza, C., 2005. Activation of the endocannabinoid system as a therapeutic approach in a murine model of multiple sclerosis. FASEB J. 19, 1338-1340. https://doi.org/10.1096/fj.04-2464fje

Palazuelos, J., Aguado, T., Pazos, M.R., Julien, B., Carrasco, C., Resel, E., Sagredo, O., Benito, C., Romero, J., Azcoitia, I., Fernández-Ruiz, J., Guzmán, M., Galve-Roperh, I., 2009. Microglial CB2 cannabinoid receptors are neuroprotective in Huntington's disease excitotoxicity. Brain 132, 3152-3164. https://doi.org/10.1093/brain/awp239

Palomo-Garo, C., Gómez-Gálvez, Y., García, C., Fernández-Ruiz, J., 2016. Targeting the cannabinoid CB2 receptor to attenuate the progression of

motor deficits in LRRK2-transgenic mice. Pharmacol Res 110, 181-192. https://doi.org/10.1016/j.phrs.2016.04.004

Pan, S.D., Grandgirard, D., Leib, S.L., 2020. Adjuvant Cannabinoid Receptor Type 2 Agonist Modulates the Polarization of Microglia Towards a Non-Inflammatory Phenotype in Experimental Pneumococcal Meningitis. Front Cell Infect Microbiol 10, 588195. https://doi.org/10.3389/fcimb.2020.588195

Peng, J., Fan, M., An, C., Ni, F., Huang, W., Luo, J., 2022. A narrative review of molecular mechanism and therapeutic effect of cannabidiol (CBD). Basic & Clinical Pharmacology & Toxicology 130, 439-456. https://doi.org/10.1111/bcpt.13710

Persidsky, Y., Fan, S., Dykstra, H., Reichenbach, N.L., Rom, S., Ramirez, S.H., 2015. Activation of Cannabinoid Type Two Receptors (CB2) Diminish Inflammatory Responses in Macrophages and Brain Endothelium. J Neuroimmune Pharmacol 10, 302-308. https://doi.org/10.1007/s11481-015-9591-3

Pertwee, R.G., 2012. Targeting the endocannabinoid system with cannabinoid receptor agonists: pharmacological strategies and therapeutic possibilities. Philos Trans R Soc Lond B Biol Sci 367, 3353-3363. https://doi.org/10.1098/rstb.2011.0381

Pertwee, R.G., 2008. The diverse CB1 and CB2 receptor pharmacology of three plant cannabinoids: Δ9-tetrahydrocannabinol, cannabidiol and Δ9-tetrahydrocannabivarin. British Journal of Pharmacology 153, 199-215. https://doi.org/10.1038/sj.bjp.0707442

Peters, K.Z., Oleson, E.B., Cheer, J.F., 2021. A Brain on Cannabinoids: The Role of Dopamine Release in Reward Seeking and Addiction. Cold Spring Harb Perspect Med 11, a039305. https://doi.org/10.1101/cshperspect.a039305

Porter, R.F., Szczesniak, A.-M., Toguri, J.T., Gebremeskel, S., Johnston, B., Lehmann, C., Fingerle, J., Rothenhäusler, B., Perret, C., Rogers-Evans, M., Kimbara, A., Nettekoven, M., Guba, W., Grether, U., Ullmer, C., Kelly, M.E.M., 2019. Selective Cannabinoid 2 Receptor Agonists as Potential Therapeutic Drugs for the Treatment of Endotoxin-Induced Uveitis. Molecules 24. https://doi.org/10.3390/molecules24183338

Powers, M.S., Breit, K.R., Chester, J.A., 2015. Genetic Versus Pharmacological Assessment of the Role of Cannabinoid Type 2 Receptors in Alcohol Reward-Related Behaviors. Alcohol Clin Exp Res 39, 2438-2446. https://doi.org/10.1111/acer.12894

Presley, C., Abidi, A., Suryawanshi, S., Mustafa, S., Meibohm, B., Moore, B.M., 2015. Preclinical evaluation of SMM-189, a cannabinoid receptor 2-specific inverse agonist. Pharmacol Res Perspect 3. https://doi.org/10.1002/prp2.159

Qian, W.-J., Yin, N., Gao, F., Miao, Y., Li, Q., Li, F., Sun, X.-H., Yang, X.-L., Wang, Z., 2017. Cannabinoid CB1 and CB2 receptors differentially modulate L- and T-type Ca 2+ channels in rat retinal ganglion cells. Neuropharmacology 124, 143-156. https://doi.org/10.1016/j.neuropharm.2017.04.027

Racz, I., Nadal, X., Alferink, J., Baños, J.E., Rehnelt, J., Martín, M., Pintado, B., Gutierrez-Adan, A., Sanguino, E., Manzanares, J., Zimmer, A., Maldonado, R., 2008. Crucial Role of CB2 Cannabinoid Receptor in the Regulation of Central Immune Responses during Neuropathic Pain. J Neurosci 28, 12125-12135. https://doi.org/10.1523/JNEUROSCI.3400-08.2008

Ramírez, B.G., Blázquez, C., del Pulgar, T.G., Guzmán, M., de Ceballos, M.L., 2005. Prevention of Alzheimer's Disease Pathology by Cannabinoids: Neuroprotection Mediated by Blockade of Microglial Activation. J Neurosci 25, 1904-1913. https://doi.org/10.1523/JNEUROSCI.4540-04.2005

Ramirez, S.H., Haskó, J., Skuba, A., Fan, S., Dykstra, H., McCormick, R., Reichenbach, N., Krizbai, I., Mahadevan, A., Zhang, M., Tuma, R., Son, Y.-J., Persidsky, Y., 2012. Activation of Cannabinoid Receptor 2 Attenuates Leukocyte-Endothelial Cell Interactions and Blood-Brain Barrier Dysfunction under Inflammatory Conditions. J Neurosci 32, 4004-4016. https://doi.org/10.1523/JNEUROSCI.4628-11.2012

Reiner, A., Heldt, S.A., Presley, C.S., Guley, N.H., Elberger, A.J., Deng, Y., D'Sumey, L., Rogers, J.T., Ferrell, J., Bu, W., Del Mar, N., Honig, M.G., Gurley, S.N., Moore, B.M., 2014. Motor, Visual and Emotional Deficits in Mice after Closed-Head Mild Traumatic Brain Injury Are Alleviated by the Novel CB2
Inverse Agonist SMM-189. Int J Mol Sci 16, 758-787.
https://doi.org/10.3390/ijms16010758

Rentsch, P., Stayte, S., Egan, T., Clark, I., Vissel, B., 2020. Targeting the cannabinoid receptor CB2 in a mouse model of l-dopa induced dyskinesia. Neurobiology of Disease 134, 104646.
https://doi.org/10.1016/j.nbd.2019.104646

Reusch, N., Ravichandran, K.A., Olabiyi, B.F., Komorowska-Müller, J.A., Hansen, J.N., Ulas, T., Beyer, M., Zimmer, A., Schmöle, A.-C., 2022. Cannabinoid receptor 2 is necessary to induce toll-like receptor-mediated microglial activation. Glia 70, 71-88. https://doi.org/10.1002/glia.24089

Ribeiro, R., Wen, J., Li, S., Zhang, Y., 2013. Involvement of ERK1/2, cPLA2 and NF-κB in microglia suppression by cannabinoid receptor agonists and antagonists. Prostaglandins & Other Lipid Mediators 100-101, 1-14.
https://doi.org/10.1016/j.prostaglandins.2012.11.003

Rinaldi-Carmona, M., Barth, F., Millan, J., Derocq, J.-M., Casellas, P., Congy, C., Oustric, D., Sarran, M., Bouaboula, M., Calandra, B., Portier, M., Shire, D., Brelière, J.-C., Fur, G.L., 1998. SR 144528, the First Potent and Selective Antagonist of the CB2 Cannabinoid Receptor. J Pharmacol Exp Ther 284, 644-650.

Rodríguez-Cueto, C., Gómez-Almería, M., García Toscano, L., Romero, J., Hillard, C.J., de Lago, E., Fernández-Ruiz, J., 2021. Inactivates the CB2 receptor accelerated the neuropathological deterioration in TDP-43 transgenic mice, a model of amyotrophic lateral sclerosis. Brain Pathol 31, e12972. https://doi.org/10.1111/bpa.12972

Rom, S., Zuluaga-Ramirez, V., Dykstra, H., Reichenbach, N.L., Pacher, P., Persidsky, Y., 2013. Selective activation of cannabinoid receptor 2 in leukocytes suppresses their engagement of the brain endothelium and protects the blood-brain barrier. Am J Pathol 183, 1548-1558.
https://doi.org/10.1016/j.ajpath.2013.07.033

Romero-Sandoval, E.A., Horvath, R., Landry, R.P., DeLeo, J.A., 2009. Cannabinoid receptor type 2 activation induces a microglial anti-inflammatory phenotype and reduces migration via MKP induction and ERK dephosphorylation. Mol Pain 5, 25. https://doi.org/10.1186/1744-8069-5-25

Romero-Zerbo, S.Y., Garcia-Gutierrez, M.S., Suárez, J., Rivera, P., Ruz-Maldonado, I., Vida, M., Fonseca, F.R. de, Manzanares, J., Bermúdez-Silva, F.J., 2012. Overexpression of Cannabinoid CB2 Receptor in the Brain Induces Hyperglycaemia and a Lean Phenotype in Adult Mice. Journal of Neuroendocrinology 24, 1106-1119. https://doi.org/10.1111/j.1365-2826.2012.02325.x

Ronca, R.D., Myers, A.M., Ganea, D., Tuma, R.F., Walker, E.A., Ward, S.J., 2015. A selective cannabinoid CB2 agonist attenuates damage and improves memory retention following stroke in mice. Life Sci 138, 72-77.
https://doi.org/10.1016/j.lfs.2015.05.005

Rorato, R., Ferreira, N.L., Oliveira, F.P., Fideles, H.J., Camilo, T.A., Antunes-Rodrigues, J., Mecawi, A.S., Elias, L.L.K., 2022. Prolonged Activation of Brain CB2 Signaling Modulates Hypothalamic Microgliosis and Astrogliosis in High Fat Diet-Fed Mice. Int J Mol Sci 23, 5527.
https://doi.org/10.3390/ijms23105527

Ross, R.A., Brockie, H.C., Stevenson, L.A., Murphy, V.L., Templeton, F., Makriyannis, A., Pertwee, R.G., 1999. Agonist-inverse agonist characterization at CB1 and CB2 cannabinoid receptors of L759633, L759656 and AM630. Br J Pharmacol 126, 665-672.
https://doi.org/10.1038/sj.bjp.0702351

Rowley, S., Sun, X., Lima, I.V., Tavenier, A., de Oliveira, A.C.P., Dey, S.K., Danzer, S., 2017. Cannabinoid receptor 1/2 double-knockout mice develop epilepsy. Epilepsia 58, e162-e166.
https://doi.org/10.1111/epi.13930

Ruiz de Martín Esteban, S., Benito-Cuesta, I., Terradillos, I., Martínez-Relimpio, A.M., Arnanz, M.A., Ruiz-Pérez, G., Korn, C., Raposo, C., Sarott, R.C., Westphal, M.V., Elezgarai, I., Carreira, E.M., Hillard, C.J., Grether, U., Grandes, P., Grande, M.T., Romero, J., 2022. Cannabinoid CB2 Receptors Modulate Microglia Function and Amyloid Dynamics in a Mouse Model of Alzheimer's Disease. Front Pharmacol 13, 841766.
https://doi.org/10.3389/fphar.2022.841766

Sadanandan, S.M., Kreko-Pierce, T., Khatri, S.N., Pugh, J.R., 2020. Cannabinoid type 2 receptors inhibit GABAA receptor-mediated currents

in cerebellar Purkinje cells of juvenile mice. PLoS One 15, e0233020.
https://doi.org/10.1371/journal.pone.0233020

Sagar, D.R., Kelly, S., Millns, P.J., O'Shaughnessey, C.T., Kendall, D.A., Chapman, V., 2005. Inhibitory effects of CB1 and CB2 receptor agonists on responses of DRG neurons and dorsal horn neurons in neuropathic rats. European Journal of Neuroscience 22, 371-379.
https://doi.org/10.1111/j.1460-9568.2005.04206.x

Sagredo, O., González, S., Aroyo, I., Pazos, M.R., Benito, C., Lastres-Becker, I., Romero, J.P., Tolón, R.M., Mechoulam, R., Brouillet, E., Romero, J., Fernández-Ruiz, J., 2009. Cannabinoid CB2 receptor agonists protect the striatum against malonate toxicity: relevance for Huntington's disease. Glia 57, 1154-1167. https://doi.org/10.1002/glia.20838

Sahu, P., Mudgal, J., Arora, D., Kinra, M., Mallik, S.B., Rao, C.M., Pai, K.S.R., Nampoothiri, M., 2019a. Cannabinoid receptor 2 activation mitigates lipopolysaccharide-induced neuroinflammation and sickness behavior in mice. Psychopharmacology 236, 1829-1838.
https://doi.org/10.1007/s00213-019-5166-y

Sahu, P., Mudgal, J., Arora, D., Kinra, M., Mallik, S.B., Rao, C.M., Pai, K.S.R., Nampoothiri, M., 2019b. Cannabinoid receptor 2 activation mitigates lipopolysaccharide-induced neuroinflammation and sickness behavior in mice. Psychopharmacology 236, 1829-1838.
https://doi.org/10.1007/s00213-019-5166-y

Salter, M.W., Beggs, S., 2014. Sublime Microglia: Expanding Roles for the Guardians of the CNS. Cell 158, 15-24.
https://doi.org/10.1016/j.cell.2014.06.008

Sánchez-Zavaleta, R., Cortés, H., Avalos-Fuentes, J.A., García, U., Vila, J.S., Erlij, D., Florán, B., 2018. Presynaptic cannabinoid CB2 receptors modulate [3H]-Glutamate release at subthalamo-nigral terminals of the rat. Synapse 72, e22061. https://doi.org/10.1002/syn.22061

Schmitz, K., Mangels, N., Häussler, A., Ferreirós, N., Fleming, I., Tegeder, I., 2016. Pro-inflammatory obesity in aged cannabinoid-2 receptor-deficient mice. Int J Obes 40, 366-379. https://doi.org/10.1038/ijo.2015.169

Schmöle, A.-C., Lundt, R., Ternes, S., Albayram, Ö., Ulas, T., Schultze, J.L., Bano, D., Nicotera, P., Alferink, J., Zimmer, A., 2015. Cannabinoid receptor 2 deficiency results in reduced neuroinflammation in an Alzheimer's disease mouse model. Neurobiology of Aging 36, 710-719.
https://doi.org/10.1016/j.neurobiolaging.2014.09.019

Schmöle, A.-C., Lundt, R., Toporowski, G., Hansen, J.N., Beins, E., Halle, A., Zimmer, A., 2018. Cannabinoid Receptor 2-Deficiency Ameliorates Disease Symptoms in a Mouse Model with Alzheimer's Disease-Like Pathology. J Alzheimers Dis 64, 379-392. https://doi.org/10.3233/JAD-180230

Schurman, L.D., Lu, D., Kendall, D.A., Howlett, A.C., Lichtman, A.H., 2019. Molecular Mechanism and Cannabinoid Pharmacology, in: Nader, M.A., Hurd, Y.L. (Eds.), Substance Use Disorders, Handbook of Experimental Pharmacology. Springer International Publishing, Cham, pp. 323-353.
https://doi.org/10.1007/164_2019_298

Shao, B., Wei, W., Ke, P., Xu, Z., Zhou, J., Liu, C., 2014. Activating Cannabinoid Receptor 2 Alleviates Pathogenesis of Experimental Autoimmune Encephalomyelitis Via Activation of Autophagy and Inhibiting NLRP3 Inflammasome. CNS Neurosci Ther 20, 1021-1028.
https://doi.org/10.1111/cns.12349

Shapiro, L., Wong, J.C., Escayg, A., 2019. Reduced Cannabinoid 2 Receptor Activity Increases Susceptibility to Induced Seizures in Mice. Epilepsia 60, 2359-2369. https://doi.org/10.1111/epi.16388

Sheng, W.S., Chauhan, P., Hu, S., Prasad, S., Lokensgard, J.R., 2019. Antiallodynic Effects of Cannabinoid Receptor 2 (CB2R) Agonists on Retrovirus Infection-Induced Neuropathic Pain. Pain Res Manag 2019.
https://doi.org/10.1155/2019/1260353

Sheng, W.S., Hu, S., Min, X., Cabral, G.A., Lokensgard, J.R., Peterson, P.K., 2005. Synthetic cannabinoid WIN55,212-2 inhibits generation of inflammatory mediators by IL-1β-stimulated human astrocytes. Glia 49, 211-219. https://doi.org/10.1002/glia.20108

Shi, H.-K., Guo, H.-C., Liu, H.-Y., Zhang, Z.-L., Hu, M.-Y., Zhang, Y., Li, Q., 2020. Cannabinoid type 2 receptor agonist JWH133 decreases blood pressure of spontaneously hypertensive rats through relieving inflammation in the rostral ventrolateral medulla of the brain. Journal of Hypertension 38,
Whiting, Z.M., Yin, J., de la Harpe, S.M., Vernall, A.J., Grimsey, N.L., 2022. Developing the Cannabinoid Receptor 2 (CB2) pharmacopoeia: past, present, and future. *Trends in Pharmacological Sciences* 43, 754–771.
https://doi.org/10.1016/j.tips.2022.06.010

Wiley, J.L., Beletskaya, I.D., Ng, E.W., Dai, Z., Crocker, P.J., Mahadevan, A., Razdan, R.K., Martin, B.R., 2002. Resorcinol derivatives: a novel template for the development of cannabinoid CB(1)/CB(2) and CB(2)-selective agonists. *J. Pharmacol. Exp. Ther.* 301, 679–689.
https://doi.org/10.1124/jpet.301.2.679

Wu, J., Bie, B., Yang, H., Xu, J.J., Brown, D.L., Naguib, M., 2013. Activation of the CB2 receptor system reverses amyloid-induced memory deficiency. *Neurobiology of Aging* 34, 791–804.
https://doi.org/10.1016/j.neurobiolaging.2012.06.011

Wu, J., Hocevar, M., Bie, B., Foss, J.F., Naguib, M., 2019. Cannabinoid Type 2 Receptor System Modulates Paclitaxel-Induced Microglial Dysregulation and Central Sensitization in Rats. *J Pain* 20, 501–514.
https://doi.org/10.1016/j.jpain.2018.10.007

Wu, J., Hocevar, M., Foss, J.F., Bie, B., Naguib, M., 2017. Activation of CB2 receptor system restores cognitive capacity and hippocampal Sox2 expression in a transgenic mouse model of Alzheimer's disease. *European Journal of Pharmacology* 811, 12–20.
https://doi.org/10.1016/j.ejphar.2017.05.044

Xi, Z.-X., Peng, X.-Q., Li, X., Song, R., Zhang, H., Liu, Q.-R., Yang, H.-J., Bi, G.-H., Li, J., Gardner, E.L., 2011. Brain Cannabinoid CB2 Receptors Modulate Cocaine's Actions in Mice. *Nat Neurosci* 14, 1160–1166.
https://doi.org/10.1038/nn.2874

Xu, J., Tang, Y., Xie, M., Bie, B., Wu, J., Yang, H., Foss, J.F., Yang, B., Rosenquist, R.W., Naguib, M., 2016. Activation of cannabinoid receptor 2 attenuates mechanical allodynia and neuroinflammatory responses in a chronic post-ischemic pain model of complex regional pain syndrome type I in rats. *European Journal of Neuroscience* 44, 3046–3055.
https://doi.org/10.1111/ejn.13414

Yang, L., Li, Z., Xu, Z., Zhang, B., Liu, A., He, Q., Zheng, F., Zhan, J., 2022. Protective Effects of Cannabinoid Type 2 Receptor Activation Against Microglia Overactivation and Neuronal Pyroptosis in Sepsis-Associated Encephalopathy. *Neuroscience* 493, 99–108.
https://doi.org/10.1016/j.neuroscience.2022.04.011

Young, A.P., Denovan-Wright, E.M., 2022. Synthetic cannabinoids reduce the inflammatory activity of microglia and subsequently improve neuronal survival in vitro. *Brain, Behavior, and Immunity* 105, 29–43.
https://doi.org/10.1016/j.bbi.2022.06.011

Youssef, D.A., El-Fayoumi, H.M., Mahmoud, M.F., 2019. Beta-caryophyllene alleviates diet-induced neurobehavioral changes in rats: The role of CB2 and PPAR-γ receptors. *Biomedicine & Pharmacotherapy* 110, 145–154.
https://doi.org/10.1016/j.biopha.2018.11.039

Yu, H., Liu, X., Chen, B., Vickstrom, C.R., Friedman, V., Kelly, T.J., Bai, X., Zhao, L., Hillard, C.J., Liu, Q.-S., 2021. The Neuroprotective Effects of the CB2 Agonist GW842166x in the 6-OHDA Mouse Model of Parkinson's Disease. *Cells* 10, 3548.
https://doi.org/10.3390/cells10123548

Yu, S.-J., Reiner, D., Shen, H., Wu, K.-J., Liu, Q.-R., Wang, Y., 2015. Time-Dependent Protection of CB2 Receptor Agonist in Stroke. *PLoS One* 10.

https://doi.org/10.1371/journal.pone.0132487

Yu, Y., Li, L., Nguyen, D.T., Mustafa, S.M., Moore, B.M., Jiang, J., 2020. Inverse Agonism of Cannabinoid Receptor Type 2 Confers Anti-inflammatory and Neuroprotective Effects Following Status Epileptics. *Mol Neurobiol* 57, 2830–2845.
https://doi.org/10.1007/s12035-020-01923-4

Zarruk, J.G., Fernández-López, D., García-Yébenes, I., García-Gutiérrez, M.S., Vivancos, J., Nombela, F., Torres, M., Burguete, M.C., Manzanares, J., Lizasoain, I., Moro, M.A., 2012. Cannabinoid Type 2 Receptor Activation Downregulates Stroke-Induced Classic and Alternative Brain Macrophage/Microglial Activation Concomitant to Neuroprotection. *Stroke* 43, 211–219.
https://doi.org/10.1161/STROKEAHA.111.631044

Zhang, H., Shen, H., Jordan, C.J., Liu, Q., Gardner, E.L., Bonci, A., Xi, Z., 2019. CB2 receptor antibody signal specificity: correlations with the use of partial CB2-knockout mice and anti-rat CB2 receptor antibodies. *Acta Pharmacol Sin* 40, 398–409.
https://doi.org/10.1038/s41401-018-0037-3

Zhang, H.-Y., Bi, G.-H., Li, X., Li, J., Qu, H., Zhang, S.-J., Li, C.-Y., Onaivi, E.S., Gardner, E.L., Xi, Z.-X., Liu, Q.-R., 2015. Species Differences in Cannabinoid Receptor 2 and Receptor Responses to Cocaine Self-Administration in Mice and Rats. *Neuropsychopharmacology* 40, 1037–1051.
https://doi.org/10.1038/npp.2014.297

Zhang, H.-Y., Gao, M., Liu, Q.-R., Bi, G.-H., Li, X., Yang, H.-J., Gardner, E.L., Wu, J., Xi, Z.-X., 2014. Cannabinoid CB2 receptors modulate midbrain dopamine neuronal activity and dopamine-related behavior in mice. *Proc Natl Acad Sci U S A* 111, E5007–E5015.
https://doi.org/10.1073/pnas.1413210111

Zhang, H.-Y., Gao, M., Shen, H., Bi, G.-H., Yang, H.-J., Liu, Q.-R., Wu, J., Gardner, E.L., Bonci, A., Xi, Z.-X., 2017. Expression of Functional Cannabinoid CB2 Receptor in VTA Dopamine Neurons in Rats. *Addict Biol* 22, 752–765.
https://doi.org/10.1111/adb.12367

Zhang, H.-Y., Shen, H., Gao, M., Ma, Z., Hempel, B.J., Bi, G.-H., Gardner, E.L., Wu, J., Xi, Z.-X., 2021. Cannabinoid CB2 receptors are expressed in glutamate neurons in the red nucleus and functionally modulate motor behavior in mice. *Neuropharmacology* 189, 108538.
https://doi.org/10.1016/j.neuropharm.2021.108538

Zhang, M., Martin, B.R., Adler, M.W., Razdan, R.K., Jallo, J.I., Tuma, R.F., 2007. Cannabinoid CB2 receptor activation decreases cerebral infarction in a mouse focal ischemia/reperfusion model. *J Cereb Blood Flow Metab* 27, 1387–1396.
https://doi.org/10.1038/sj.jcbfm.9600447

Zheng, L., Wu, X., Dong, X., Ding, X., Song, C., 2015. Effects of Chronic Alcohol Exposure on the Modulation of Ischemia-Induced Glutamate Release via Cannabinoid Receptors in the Dorsal Hippocampus. *Alcoholism: Clinical and Experimental Research* 39, 1908–1916.
https://doi.org/10.1111/acer.12845

Zoppi, S., Madrigal, J.L., Caso, J.R., García-Gutiérrez, M.S., Manzanares, J., Leza, J.C., García-Bueno, B., 2014. Regulatory role of the cannabinoid CB2 receptor in stress-induced neuroinflammation in mice. *Br J Pharmacol* 171, 2814–2826.
https://doi.org/10.1111/bph.12607

Zou, S., Kumar, U., 2018. Cannabinoid Receptors and the Endocannabinoid System: Signaling and Function in the Central Nervous System. *Int J Mol Sci* 19, 833.
https://doi.org/10.3390/ijms19030833
CB2 receptor in the CNS:
From immune and neuronal modulation to behavior

Wanda Grabon<fnref n="1" />,<fnref n="2" />*, Sylvain Rheims<fnref n="1" />,<fnref n="2" />,<fnref n="3" />, Jonathon Smith<fnref n="1" />,<fnref n="2" />, Jacques Bodennec<fnref n="1" />,<fnref n="2" />, Amor Belmeguenai<fnref n="1" />,<fnref n="2" />, Laurent Bezin<fnref n="1" />,<fnref n="2" />*

AB, LB: contributed equally.

PMID: 37164044, DOI: 10.1016/j.neubiorev.2023.105226

Number of supplementary tables: 12

<table><caption>**Supplementary Table 1 – In vitro evidence of CB2R neuroimmunomodulatory functions.** Details on CB2R ligand treatments are provided, such as the concentration of the ligand, the time it was applied to the cell culture with respect to inflammatory challenge. Outcome is either enhanced (↗) or decreased (↘).</caption><thead><tr><th>CHALLENGE</th><th>CELL LINE & MODEL</th><th>TREATMENT</th><th>OUTCOME</th><th>OTHER</th><th>REF</th></tr></thead><tbody><tr><td colspan="6"><strong>CB2R AGONISTS APPLIED BEFORE CHALLENGE</strong></td></tr><tr><td>CCL2</td><td>Primary monocytes (Human)</td><td>GP1a, JWH-133, O-1966 (10 µM), AM1241 (2 µM)</td><td>↗ Adhesion to BMVEC, ↗ monocytes migration, ↗ lamellipodia formation, ↗ BMVEC monolayer disruption</td><td>-</td><td>(Rom et al., 2013)</td></tr><tr><td>CCL2 or CCL3</td><td>Primary monocytes (Human)</td><td>JWH-015 (5-20 µM, 12h pre-challenge)</td><td>↗ monocytes migration (dose-dep), ↗ CCR2, CCR1</td><td>Reversed with SR144528 (1 µM)</td><td>(Montecucco et al., 2008)</td></tr><tr><td>IFN-γ</td><td>Primary cultured microglia (Mouse)</td><td>JWH-015, JWH-133, GP1a (10, 10 and 3 µM respectively, 2h pre-challenge)</td><td>↗ phosphorylation of STAT3/STAT1</td><td>-</td><td>(Sheng et al., 2019)</td></tr><tr><td>IL-1β</td><td>Cultured astrocytes (Human fetus)</td><td>WIN55,212-2 (0.3-10 µM, 24h pre-challenge)</td><td>↗ iNOS, NO, TNF-α, IL-1β, IL-6, CXCL10, MCP1, CCL5, CXCL8</td><td>Reversed with SR144528 (10 µM)</td><td>(Sheng et al., 2005)</td></tr><tr><td rowspan="5">LPS</td><td>BV2 (Microglia, Mouse)</td><td>AM1241, JWH-015 (0.1 µM, 30 min pre-challenge)</td><td>↗ NF-κB, iNOS, ROS</td><td>-</td><td>(Ribeiro et al., 2013)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>Anandamide (1 µM, 30 min pre-challenge)</td><td>↗ IL-6, NO, ↗ IL-10</td><td>-</td><td>(Malek et al., 2015)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>JWH-015 (0.01-1 µM, 2h pre-challenge)</td><td>↗ ADP-induced microglia migration (dose-dep)</td><td>Reversed with AM630 (1 µM)</td><td>(Romero-Sandoval et al., 2009)</td></tr><tr><td>THP-1 (Monocytes, Human)</td><td>JWH-015 (1-10 µM, 30 min pre-challenge)</td><td>↗ IL-1β, TNF-α</td><td>-</td><td>(Klegeris et al., 2003)</td></tr><tr><td>OLN-93, (Oligodendrocytes, Rat)</td><td>β-caryophyllene (0.1-50 µM) JWH-133 (1 µM, 24h pre-challenge)</td><td>↗ NO, ROS, TNF-α</td><td>Reversed with AM630 (1 µM)</td><td>(Askari and Shafiee-Nick, 2019a)</td></tr><tr><td>LPS/IFN-γ</td><td>Primary cultured Macrophages (Mouse)</td><td>JWH-133 (10 nM-5 µM, 5 min pre-challenge)</td><td>↗ IL-12 (dose-dep), ↗ IL-10</td><td>Reversed with SR144528 (1 µM)</td><td>(Correa et al., 2005)</td></tr><tr><td>MPP+</td><td>Primary cultured astrocytes (Rat)</td><td>JWH-133 (1 µM for 40 min pre-challenge)</td><td>↗ iNOS, COX2, TNF-α, IL-1β, DMT1</td><td>Reversed with AM630 (1 µM)</td><td>(Jia et al., 2020)</td></tr><tr><td>TNF-α</td><td>Primary monocytes (Human)</td><td>GP1a, JWH-133, O-1966 (10 µM), AM1241 (2 µM)</td><td>↗ Adhesion to BMVEC, ↗ monocytes migration, ↗ lamellipodia formation, ↗ BMVEC monolayer disruption</td><td>-</td><td>(Rom et al., 2013)</td></tr><tr><td colspan="6"><strong>CB2R AGONISTS APPLIED DURING CHALLENGE</strong></td></tr><tr><td>Αβ<sub>1-40</sub></td><td>Primary cultured microglia (mouse)</td><td>HU210, JWH-133, WIN55,212-2 (0.1 µM, 4h of co-treatment)</td><td>↗ microglial activation: ↗ rod-like morphology, ↗ TNF-α</td><td>-</td><td>(Ramírez et al., 2005)</td></tr><tr><td rowspan="2">Αβ<sub>1-42</sub></td><td>Primary cultured microglia (Rat)</td><td>JWH-133 (0.1 µM, 4h of co-treatment)</td><td>↗ TNF-α</td><td>-</td><td>(Ramírez et al., 2012)</td></tr><tr><td>C6 Glioma cell (Rat)</td><td>JWH-015 (1 µM, 24h of co-treatment)</td><td>↗ GFAP</td><td>-</td><td>(Esposito et al., 2007)</td></tr><tr><td>Αβ<sub>1-42</sub>/CD40</td><td>Primary cultured microglia (Mouse)</td><td>JWH-015 (5 µM, 24h of co-treatment)</td><td>↗ TNF-α, NO</td><td>-</td><td>(Ehrhart et al., 2005)</td></tr><tr><td>sCD40L</td><td>BMVEC (endothelial cells, Human)</td><td>JWH-133, O-1966 (10 µM, 24h of co-treatment)</td><td>↗ “BBB” model permeability</td><td>-</td><td>(Persidsky et al., 2015)</td></tr><tr><td>CCL2</td><td>BMVEC (endothelial cells, Human)</td><td>JWH-133, O-1966 (10 µM, 24h of co-treatment)</td><td>↗ monocyte migration</td><td>-</td><td>(Persidsky et al., 2015)</td></tr><tr><td rowspan="2">CXCL2</td><td>Cultured neutrophils (Mouse)</td><td>JWH-133 (1 µM for 30 min co-treatment)</td><td>↗ chemotaxis</td><td>No effect in CB2R KO cells</td><td>(Murikinata et al., 2010)</td></tr><tr><td>Primary cultured neutrophils (Mouse)</td><td>RO6871304 (0.01-1 µM, 30 min of co-treatment)</td><td>↗ neutrophil migration</td><td>No effect in CB2R KO neutrophils</td><td>(Porter et al., 2019)</td></tr></tbody></table>

<footnote n="1">¹ Université Claude Bernard Lyon 1, CNRS, Inserm, Centre de Recherche en Neurosciences de Lyon, U10208 UMR5292, TIGER Team – F-69500 Bron, France</footnote>

<footnote n="2">² Epilepsy Institute IDEE, 59 boulevard Pinel – F-69500 Bron, France</footnote>

<footnote n="3">³ Department of Functional Neurology and Epileptology, Hospices Civils de Lyon – France</footnote>
<table><tbody><tr><td rowspan="2">IFN-γ</td><td>Primary cultured microglia (Mouse)</td><td>JWH-015 (5 µM, 12h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (1 µM)</td><td>(Ehrhart et al., 2005)</td></tr><tr><td>Primary monocytes (Human)</td><td>JWH-015, JWH-133 (20 µM, 10 µM respectively, 24h co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (1 µM)</td><td>(Montecucco et al., 2008)</td></tr><tr><td rowspan="2">LPS, IFN-γ</td><td>SIM-A9 (Microglia, mouse)</td><td>HU308 (2.5 µM, 30 min of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (1 µM)</td><td>(Young and Donovan-Wright, 2022)</td></tr><tr><td>Primary cultured microglia (Mouse)</td><td>JWH-133 (10-100 nM, 18h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (1 µM)</td><td>(Correa et al., 2010)</td></tr><tr><td rowspan="6">LPS</td><td>Mixed glial cells (Microglia and astrocytes, rat)</td><td>JWH-133 (1 µM, 48h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (1 µM)</td><td>(Pan et al., 2020)</td></tr><tr><td>Primary cultured microglia (Mouse)</td><td>Anandamide (5 µM, 12h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Partially reversed with SR144528 (1 µM)</td><td>(Ortega-Gutiérrez et al., 2005)</td></tr><tr><td>Primary cultured microglia (Mouse)</td><td>β-caryophyllene, JWH-133 (0.2-25, 1 µM respectively, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (0.1 µM)</td><td>(Askari and Shafiee-Nick, 2019b)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>HU210, WIN55,212-2 (1-10 µM, 2h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (10 µM)</td><td>(Facchinetti et al., 2003)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>JWH-015 (1 µM, 1-2h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (10 µM)</td><td>(Romero-Sandoval et al., 2009)</td></tr><tr><td>MDM (Monocyte-derived macrophages, Human)</td><td>JWH-133 (10 µM, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (10 µM)</td><td>(Persidsky et al., 2015)</td></tr><tr><td rowspan="2">MPP+</td><td>SH-SY5Y (neuroblastoma cells, Human)</td><td>β-caryophyllene (1-2.5 µM, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (10 µM)</td><td>(G. Wang et al., 2018)</td></tr><tr><td>S. pneumoniae serotype 3</td><td>Mixed glial cells (Microglia and astrocytes, rat)</td><td>JWH-133 (1 µM, 48h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with SR144528 (10 µM)</td><td>(Pan et al., 2020)</td></tr><tr><td rowspan="3">TNF-α</td><td>Endothelium (Endothelial cells, mouse)</td><td>JWH-133 (1 µM for 20 min co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>No effect in CB2R KO cells</td><td>(Murikinata et al., 2010)</td></tr><tr><td>BMVEC (endothelial cells, Human)</td><td>JWH-133, O-1966 (10 µM, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>No effect in CB2R KO cells</td><td>(Persidsky et al., 2015)</td></tr><tr><td>HBMEC (endothelial cells, Human)</td><td>JWH-133, O-1966 (10 µM, 4-24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>No effect in CB2R KO cells</td><td>(Ramirez et al., 2012)</td></tr><tr><td colspan="6">CB2R AGONISTS APPLIED AFTER CHALLENGE</td></tr><tr><td>LPS/IFN-γ</td><td>N9 (Microglia, mouse)</td><td>AM1241 (5 µM, 24h treatment, 1h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Ma et al., 2015)</td></tr><tr><td>LPS</td><td>C8B4 (Microglia, mouse)</td><td>HU308, JWH-133 (24h treatment, 1h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Presley et al., 2015)</td></tr><tr><td rowspan="2">Thrombin</td><td>Primary cultured microglia (Rat)</td><td>JWH-133 (4 µM, 24h treatment, 24h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (1 µM)</td><td>(Tang et al., 2015)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>JWH-133 (4 µM, 24h treatment, 24h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (1 µM)</td><td>(Tao et al., 2016)</td></tr><tr><td colspan="6">CB2R INVERSE AGONISTS APPLIED BEFORE CHALLENGE</td></tr><tr><td rowspan="3">IL-4 + IL-13</td><td>Primary cultured microglia (Rat)</td><td>AM630 (1 µM, 30 min pre-challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Mecha et al., 2015)</td></tr><tr><td>BV2 (Microglia, mouse)</td><td>AM630, SR144528 (0.1 µM, 30 min pre-challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Ribeiro et al., 2013)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>AM630 (0.5 µM, 24h treatment 30 min pre-challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Malek et al., 2015)</td></tr><tr><td rowspan="2">LPS</td><td>Primary cultured microglia (Rat)</td><td>SMM-189 (10-40 µM, 15 min pre-challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Yu et al., 2020)</td></tr><tr><td>Primary cultured microglia (Rat)</td><td>SMM-189 (10-40 µM, 15 min pre-challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Yu et al., 2020)</td></tr><tr><td colspan="6">CB2R INVERSE AGONISTS APPLIED DURING CHALLENGE</td></tr><tr><td>Aβ1-42</td><td>C6 Glioma cell (Rat)</td><td>SR144528 (1 µM 12-24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Esposito et al., 2007)</td></tr><tr><td>IL-4</td><td>Immortalized microglia (Human)</td><td>SMM-189 (13.4 µM, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Reiner et al., 2014)</td></tr><tr><td>LPS</td><td>Immortalized microglia (Human)</td><td>SMM-189 (13.4 µM, 24h of co-treatment)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Reiner et al., 2014)</td></tr><tr><td colspan="6">CB2R INVERSE AGONISTS APPLIED AFTER CHALLENGE</td></tr><tr><td rowspan="3">LPS</td><td>C8B4 (Microglia, mouse)</td><td>SMM-189, SR144528 (0.15 µM, 24h of treatment 1h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Presley et al., 2015)</td></tr><tr><td>Primary cultured microglia (Human)</td><td>SMM-189 (0.15 µM, 24h of treatment 1h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Presley et al., 2015)</td></tr><tr><td>Primary cultured microglia (Human)</td><td>SMM-189 (9.8 µM, 18h of treatment 1h post challenge)</td><td>IL-1β, TNF-α, NO, CD40, JAK/STAT phosphorylation</td><td>Reversed with AM630 (10 µM)</td><td>(Reiner et al., 2014)</td></tr></tbody></table>

**Abbreviations:** Aβ, amyloid beta; ADP, adenosine diphosphate; Arg-1, arginase 1; BBB, blood-brain barrier; BDNF, brain-derived neurotrophic factor; BMVEC, brain microvascular endothelial cells; cAMP, cyclic adenosine monophosphate; CB2R, cannabinoid receptor type 2; CCL, Chemokine (C-C motif) ligand; CCR, C-C chemokine receptor; CD, cluster of differentiation; COX2, cyclooxygenase 2; CXCL, chemokine (C-X-C motif) ligand; DMT1, divalent metal transporter 1; Epac1, Exchange Protein Directly Activated by cAMP Type 1; ERK, Extracellular signal-regulated kinases; GDNF, Glial cell line-derived neurotrophic factor; GFAP, Glial fibrillary acidic protein; GSH, reduced glutathione; HBMEC, human brain microvascular endothelial cells; Iba1, ionized calcium-binding adapter molecule 1; ICAM, intercellular adhesion molecule; IFN, interferon; IL, interleukin; iNOS, Inducible nitric oxide synthase; JAK, janus kinase; LPS, lipopolysaccharide; MAPK, mitogen-activated protein kinase; MCP1, monocyte chemoattractant protein 1; MCSF-1, macrophage colony-stimulating factor 1; MDM, monocyte-derived macrophage; MPP+, 1-methyl-4-phenylpyridinium; NF-κB, nuclear factor-kappa B; PGE2, prostaglandin E2; PKA, Protein kinase A; Ref, reference; ROS, Reactive oxygen species; STAT, signal transducer and activator of transcription; THP-1, Tohoku Hospital Pediatrics-1 Cells; TNF, tumor necrosis factor; VCAM, vascular cell adhesion molecule.
<table><caption>**Supplementary Table 2 – In vivo evidence of CB2R neuro-immunomodulatory functions.** Details on CB2R ligand treatments are provided, such as the dose of the ligand, treatment initiation and duration, administration route, with respect to inflammatory insult. Outcome is either enhanced (>) decreased (>) or unchanged (=). In some studies, the specificity of the ligand used has been challenged by the use of CBR2 KO mice (the “Deltagen strain”, with a deletion of the sequence encoding the N-terminal part of the protein (The Jackson Laboratory, Cnr2tm10gen/], #005786), or the “Zimmer strain” with a deletion of the sequence encoding the C-terminal part of the protein (Buckley et al., 2000)).</caption><thead><tr><th>MODEL</th><th>TREATMENT</th><th>TISSUE</th><th>OUTCOME</th><th>OTHER</th><th>REF</th></tr></thead><tbody><tr><td colspan="6"><strong>CB2R AGONISTS GIVEN BEFORE CHALLENGE</strong></td></tr><tr><td>Cerebral ischemia (MCAO, Mouse)</td><td>JWH-133 (1 mg/kg/day, IP microosmotic pump, 4h pre-insult, for >3 days)</td><td>Ischemic hemisphere</td><td>✓ neutrophils recruitment, ✓ MPO activity, = TNF-α, IL-1β, CXCL2</td><td>-</td><td>(Murikinata et al., 2010)</td></tr><tr><td>Chronic post-ischemia pain (Rat)</td><td>MDA7 (15 mg/kg, IP, 30 min pre-insult + daily for 14 days)</td><td>Spinal cord dorsal horn</td><td>✓ CD11b, CXCR3</td><td>Reversed with AM630 (5 mg/kg, IP)</td><td>(Xu et al., 2016)</td></tr><tr><td>Focal ischemia/reperfusion (Mouse)</td><td>O-1966, O-3853 (1 mg/kg, IV, 1h pre-insult)</td><td>Brain</td><td>✓ leukocyte/endothelium interaction</td><td>-</td><td>(Zhang et al., 2007)</td></tr><tr><td>HIV-1 glycoprotein 120 induced neural injury (Rat)</td><td>WIN55,212-2 (3 mg/kg/day, IP, for 3 days pre-insult)</td><td>Hippocampus</td><td>✓ IL-1β, TNF-α, IL-6, CXCL10, ↗ IL-10</td><td>Reversed with AM630 (1.5 mg/kg/day, IP)</td><td>(Wang et al., 2022)</td></tr><tr><td>Huntington's disease (Rat)</td><td>HU308 (5 mg/kg, IP, 20 min pre- and 2h post insult)</td><td>Striatum</td><td>✓ TNF-α, = SOD1, SOD2</td><td>-</td><td>(Sagredo et al., 2009)</td></tr><tr><td>LPS-induced inflammation (Mouse)</td><td>1-phenylisatin (20 mg/kg, PO, 1h pre-insult)</td><td>Brain</td><td>✓ TNF-α, ↗ GSH, catalase, SOD</td><td>-</td><td>(Sahu et al., 2019)</td></tr><tr><td rowspan="2">MDMA (ecstasy) induced inflammation (Rat)</td><td>JWH-015 (2.4 mg/kg, IP, 2 days, 1 day and 30 min pre-insult)</td><td>Hypothalamus & PFC</td><td>✓ IL-1β, = CD11b</td><td>Reversed with AM630 (3 mg/kg, IP)</td><td>(Torres et al., 2010)</td></tr><tr><td>JWH-015 (2.4 mg/kg, IP, 2 days, 1 day and 30 pre+ 6h post-insult)</td><td>Hypothalamus & PFC</td><td>✓ CD11b</td><td>Reversed with AM630 (3mg/kg, IP)</td><td>(Torres et al., 2010)</td></tr><tr><td>Neonatal stroke (Rat)</td><td>JWH-133 (1.5 mg/kg, IP, 1h pre-insult)</td><td>Peri-hematoma brain</td><td>✓ TNF-α, ✓ number of Iba1+ cells</td><td>-</td><td>(Tang et al., 2016)</td></tr><tr><td>Paclitaxel-induced inflammation (Rat)</td><td>MDA7 (15 mg/kg, IP, 15 min pre-insult and for 4 days)</td><td>Spinal dorsal horn (L4-L5)</td><td>✓ Iba1 immunostaining intensity, ↗ IL-6, ↗ IL-10</td><td>-</td><td>(Wu et al., 2019)</td></tr><tr><td rowspan="2">Parkinson's disease (Mouse)</td><td>JWH-133, WIN55,212 (10 µg/kg/day, IP, 2 days pre-insult + for 8 days post-insult)</td><td>Substansia Nigra</td><td>✓ IL-1β, TNF-α, iNOS, ↗ CCL3, CCL4, MCP-1, CXCL10, CCL5, ↗ T-cells and macrophages infiltration, ✓ number of MAC1+ cells, ↗ GFAP, MPO, ↗ BBB permeability</td><td>Reversed with AM630 (20 µg/kg/day, IP)</td><td>(Chung et al., 2016)</td></tr><tr><td>β-caryophyllene (10 mg/kg, IP, 1/day for 5 days, 24h pre-insult)</td><td>Substansia Nigra and striatum</td><td>= TNF-α, ↗ IL-6, IL-1β, ↗ number of Iba1+ and GFAP+ cells</td><td>Reversed with AM630 (3 mg/kg, IP)</td><td>(Viveros-Paredes et al., 2017)</td></tr><tr><td>Parkinson's disease (Rat)</td><td>β-caryophyllene (50 mg/kg/day, IP, for 4 weeks and 30 min pre-insult)</td><td>Midbrain</td><td>✓ TNF-α, IL-6, IL-1β, NF-κB p65, iNOS, COX-2, ↗ GSH, catalase, SOD, ↗ Iba1, ↗ GFAP</td><td>Partially reversed with AM630 (1 mg/kg, IP)</td><td>(Javed et al., 2016)</td></tr><tr><td>PTZ-induced seizure (Rat)</td><td>HU308 (3 mg/kg, IP, 30 min pre-insult)</td><td>Dorsal hippocampus and neocortex</td><td>✓ TNF-α, IL-6 (hippocampus only)</td><td>-</td><td>(Karan et al., 2021)</td></tr><tr><td>Spinal cord ischemia (Rat)</td><td>JWH-133 (1 mg/kg, IP, 1h pre-insult)</td><td>Spinal cord</td><td>✓ TLR4, MMP9, MyD88, NF-κB p65, ↗ GFAP, ↗ BBB permeability, ↗ Zonula occludens-1</td><td>Reversed with AM630 (1 mg/kg, IP)</td><td>(Jing et al., 2020)</td></tr><tr><td>Theiler's virus induced demyelination disease (Mouse)</td><td>2-AG (5 mg/kg, IP, 48h and 24h pre-insult)</td><td>Cortex</td><td>✓ Iba1+ area, ↗ CSF1R, TNF-α, iNOS</td><td>Reversed with AM630 (2 mg/kg, IP)</td><td>(Mecha et al., 2018)</td></tr><tr><td colspan="6"><strong>CB2R AGONSISTS GIVEN DURING CHALLENGE</strong></td></tr><tr><td>AMPA-induced retina inflammation (Rat)</td><td>WIN55,212 (10<sup>-7</sup> to 10<sup>-4</sup> M, intravitreal)</td><td>Retina</td><td>✓ number of Iba1+ cells</td><td>Reversed with AM630 (0.1 mM, intraviteal)</td><td>(Spyridakos et al., 2021)</td></tr><tr><td>Chronic restraint stress (Rat)</td><td>β-Caryophyllene (25-100 mg/kg, IP, daily for 28 days)</td><td>Hippocampus</td><td>✓ COX-2, ↗ BDNF</td><td>-</td><td>(Hwang et al., 2020)</td></tr><tr><td>EAE (Mouse)</td><td>β-Caryophyllene (50 mg/kg, PO, twice a day, from day 0 to 30 post-insult)</td><td>Lumbar spinal cord</td><td>✓ Iba1+ area, ↗ iNOS</td><td>-</td><td>(Alberti et al., 2017)</td></tr><tr><td>High-Fat Diet (Mouse)</td><td>HU308 (5 µg/day for 2 weeks, icv)</td><td>Hypothalamus</td><td>✓ Iba1+ cells in the hypothalamic arcuate, ↗ Iba1+ cells and GFAP+ in the VMH, = STAT3 and ERK phosphorylation</td><td>-</td><td>(Rorato et al., 2022)</td></tr><tr><td>LPS-induced inflammation (Mouse)</td><td>JWH-133, O-1966 (10 and 5 mg/kg respectively, IP)</td><td>Brain</td><td>✓ BBB permeability, ↗ leukocyte/endothelium interaction, ↗ ICAM-1</td><td>Reversed with SR144528 (10 mg/kg, IP) and in KO mice (Deltagen)</td><td>(Ramirez et al., 2012)</td></tr><tr><td>Morphine-mediated hyperalgesia (Rat)</td><td>AM1241 (0.1-2.5 mg/kg, IP, 2/day for 6 days)</td><td>Spinal Cord</td><td>✓ IL-1β, TNF-α, ↗ CD11b, ↗ GFAP (dose dep)</td><td>-</td><td>(Tumati et al., 2012)</td></tr><tr><td colspan="6"><strong>CB2R AGONISTS GIVEN AFTER CHALLENGE</strong></td></tr><tr><td>Alzheimer's disease (icv STZ, Mouse)</td><td>1-phenylisatin (10 mg/kg/day, PO, from day 1 for 30 days post-insult)</td><td>Brain</td><td>✓ MPO, ↗ AchE</td><td>-</td><td>(Jayant et al., 2016)</td></tr><tr><td>Alzheimer's disease (intracortical Aβ<sub>1-42</sub>, Rat)</td><td>JWH-015 (5mg/kg, IP, 3-, 5- and 7-days post-insult)</td><td>Hippocampus</td><td> ↗ GFAP</td><td>-</td><td>(Esposito et al., 2007)</td></tr><tr><td>Alzheimer's disease (intrahippocampal Aβ<sub>1-40</sub>, Rat)</td><td>MDA7 (15 mg/kg/day, IP, from day 1 and for 14 days post-insult)</td><td>Hippocampus</td><td>✓ IL-1β, ↗ CD11b, ↗ GFAP</td><td>-</td><td>(Wu et al., 2013)</td></tr><tr><td>Alzheimer's disease (intrahippocampal Aβ<sub>1-42</sub>, Rat)</td><td>WIN55,212 (10 µg, intra hippocampal, 1h post-insult and repeated day 3, 5 and 7)</td><td>Hippocampus</td><td>✓ TNF-α, NF-κB</td><td>Partially reversed with SR144528 (70 µg, intra hippocampal)</td><td>(Fakhfouri et al., 2012)</td></tr></tbody></table>
<table><tbody><tr><td>CO-induced cognitive memory (Rat)</td><td>WIN55,212-2 (1 mg/kg IP, 2h, 12h, and 24h post-insult)</td><td>Hippocampus</td><td>▲ number of Iba1+ cells, ↗ CD206+ cells, ↗ IL1R1, IL-33</td><td>Reversed with AM630 (1 mg/kg IP)</td><td>(Du et al., 2020)</td></tr><tr><td>Diet-induced inflammation (Rat)</td><td>β-Caryophyllene (30 mg/kg/D, PO, 9h post-insult for 4 weeks)</td><td>Prefrontal cortex</td><td>▲ TNF-α, iNOS, NF-κB</td><td>Partially reversed by AM630 (1 mg/kg, IP)</td><td>(Youssef et al., 2019)</td></tr><tr><td>Hemicerebellectomy (Rat)</td><td>JWH-015 (3 mg/kg, IP, daily for 7 days post-insult)</td><td>Pontine nuclei</td><td>▲ iNOS, ▲ number of CD11b+ and GFAP+ cells, ▲ ROS</td><td>Reversed with SR144528 (3 mg/kg, IP)</td><td>(Oddi et al., 2012)</td></tr><tr><td>HIV-1 encephalitis (Mouse)</td><td>GP1a (1 mg/kg/day, food, 1 day post-insult for 14 days)</td><td>Brain</td><td>▲ CD11b, TNF-α, = iNOS, IL-6, IL-1β, GFAP</td><td>-</td><td>(Gorantla et al., 2010)</td></tr><tr><td rowspan="2">Intracerebral Hemorrhage (Rat)</td><td>JWH-133 (1.5 mg/kg, IP, 1h post-insult)</td><td>Peri hematoma-brain</td><td>▲ IL-1β, TNF-α, iNOS, ↗ M2 polarization</td><td>-</td><td>(Lin et al., 2017)</td></tr><tr><td>JWH-133 (1.5 mg/kg, IP, 1h post-insult)</td><td>Peri hematoma-brain</td><td>▲ TNF-α, IL-6, IL-1β, p-ERK, p38, = MCP1, ▲ number of Iba1+ cells, ▲ BBB permeability, ↗ Zonula occludens-1, claudin 5, ▲ MMP9</td><td>Reversed with SR144528 (3 mg/kg, IP)</td><td>(Li et al., 2018)</td></tr><tr><td rowspan="6">Multiple sclerosis (Mouse)</td><td>WIN55,212 (5 mg/kg/day, IP, 0-5 days post-insult)</td><td>Spinal cord</td><td>▲ IFN-α, IFN-β</td><td>-</td><td>(Croxford and Miller, 2003)</td></tr><tr><td>WIN55,212 (5 mg/kg/day, IP, 26-31 days post-insult)</td><td>Spinal cord</td><td>▲ IL-1β, IL-6, TNF-α, IFN-γ</td><td>-</td><td>(Ortega-Gutiérrez et al., 2005)</td></tr><tr><td>UCM707 (5 mg/kg/day, IP, 90 days post-insult, for 12 days)</td><td>Spinal cord</td><td>▲ microglial reactive morphology, CD11b</td><td>-</td><td>(Kong et al., 2014b)</td></tr><tr><td>GP1a (5 mg/kg, IV, twice/week from day 0)</td><td>Spinal cord</td><td>▲ T cell infiltration (T<sub>H1</sub>, T<sub>H17</sub>), ▲ number of Iba1+ cells</td><td>-</td><td>(Kong et al., 2014b)</td></tr><tr><td>GP1a (5 mg/kg, IV, twice/week from day 7)</td><td>Spinal cord</td><td>▲ TNF-α, IL-1β, IL-12p40, IL23p19, IFN-γ, IL-17, VCAM1, iNOS, CCL2, CCL5, CXCL10, ▲ T cell infiltration, ▲ number of Iba1+ cells</td><td>-</td><td>(Kong et al., 2014a)</td></tr><tr><td>β-caryophyllene (0.1-10 mg/kg daily, PO, starting 1-day post-insult)</td><td>Spinal cord</td><td>▲ Iba1, ▲ GFAP</td><td>-</td><td>(Klauke et al., 2014)</td></tr><tr><td>Okadaic acid-induced neurodegeneration (Rat)</td><td>JWH-133 (0.2 mg/kg, IP, for 13 days post-insult)</td><td>Hippocampus</td><td>▲ TNF-α, IL-1β</td><td>-</td><td>(Cakur et al., 2019)</td></tr><tr><td>Parkinson's disease (Intrastriatal LPS Injection, Mouse)</td><td>HU308 (5 mg/kg/day, IP, 16h post-insult and for 14 days)</td><td>Striatum</td><td>= TNF-α, IL-1β, ▲ iNOS, ▲ CD68+ cells</td><td>No effect in KO mice (Zimmer)</td><td>(Gómez-Gálvez et al., 2016)</td></tr><tr><td>Parkinson's disease (6-OHDA and L-DOPA, Mouse)</td><td>HU308 (1-5 mg/kg, IP, daily beginning 3 weeks post-insult)</td><td>Striatum</td><td>▲ Iba1+ cells, ▲ GFAP+ cells, ▲ TNF-α, IL-1β</td><td>-</td><td>(Rentsch et al., 2020)</td></tr><tr><td>Pneumococcal Meningitis Model (Rat)</td><td>JWH-133 (1 mg/kg, 18h post-insult)</td><td>Cortex</td><td>▲ number of reactive microglial cells, ↗ resting microglia, ▲ neutrophil chemoattractant chemokines expression, ▲ IL-6, CCL3, CCL20</td><td>-</td><td>(Pan et al., 2020)</td></tr><tr><td>Retrovirus Infection-Induced Neuropathic Pain (Mouse)</td><td>JWH-015, JWH-133, GP1a, HU308 (5mg/kg/week, 5week post-insult)</td><td>Dorsal root ganglia & lumbar spinal cord</td><td>▲ macrophages and T infiltration (GP1a only, NS)</td><td>-</td><td>(Sheng et al., 2019)</td></tr><tr><td>Sepsis-induced encephalopathy (Mouse)</td><td>HU308 (2.5 mg/kg, IP, 30 post-insult and 1/day for 3 days)</td><td>Hippocampus</td><td>▲ IL-1β, CD68</td><td>-</td><td>(Yang et al., 2022)</td></tr><tr><td>Sepsis-induced encephalopathy (Rat)</td><td>JWH-133 (0.2-5 mg/kg, IP, immediately after insult)</td><td>Cortex and hippocampus</td><td>▲ TNF-α, IL-1β, IL-6, caspase-3, p-NF-κB</td><td>-</td><td>(Cakur et al., 2020)</td></tr><tr><td>Spinal cord injury (Mouse)</td><td>O-1966 (5 mg/kg, IP, 1h+24h post-insult)</td><td>Spinal cord</td><td>= TNF-α, IL-1β, ▲ IL-23p19, IL-23R, TLR1, TLR4, TLR6, TLR7, CXCL9, CXCL11, ↗ IL-10, CCL2, CXCL1, ▲ Neutrophil and monocytes infiltration (but NS), ▲ number of Iba1+ cells</td><td>-</td><td>(Adhikary et al., 2011)</td></tr><tr><td>Stroke (Mouse)</td><td>JWH-133 (1.5 mg/kg, IP, 10 min post-insult)</td><td>Peri infarct-brain</td><td>= IL-1β, ▲ TNF-α, IL-6, IL-12, iNOS, TGF β, Ym1, IL-10, CCL5, MCP1, MIP1 α, ▲ Iba1+</td><td>-</td><td>(Zarruk et al., 2012)</td></tr><tr><td>Stroke (Rat)</td><td>AM1241 (2.5 mg/kg, IP, 2 days and 5 days post insult)</td><td>Ischemic cortex</td><td>▲ T-cell infiltration (CD4, CD8), ▲ Iba1+</td><td>-</td><td>(Yu et al., 2015)</td></tr><tr><td rowspan="2">Stroke (Neonatal, Rat)</td><td>JWH-133 (1.5 mg/kg, IP, 6h post-insult)</td><td>Peri hematoma-brain</td><td>▲ TNF-α, iNOS, p-ERK, ▲ number of Iba1+ cells</td><td>Reversed with AM630 (1 mg/kg, IP)</td><td>(Tang et al., 2015)</td></tr><tr><td>JWH-133 (1 mg/kg, IP, 1h post-insult)</td><td>Peri hematoma-brain</td><td>▲ IL-1β, TNF-α, INF-γ, ↗ IL-4, IL-10, BDNF, ▲ Iba1+CD68+ cells, ↗ M2 polarization</td><td>Reversed with AM630 (1 mg/kg, IP)</td><td>(Tao et al., 2016)</td></tr><tr><td>Surgery-induced cognitive impairment (Mouse)</td><td>JWH-133 (2 mg/kg/day, IP, immediately after insult)</td><td>Hippocampus, PFC</td><td>▲ IL-1β, TNF-α, ▲ MCP1, ▲ CD11b</td><td>-</td><td>(Sun et al., 2017)</td></tr><tr><td rowspan="2">Traumatic Brain Injury (CCI, Mouse)</td><td>O-1966 (5 mg/kg, IP, 1h+24h post-insult)</td><td>Cortex</td><td>▲ number of Iba1+ activated cells</td><td>-</td><td>(Elliott et al., 2011)</td></tr><tr><td>O-1966, JWH-133 (5 and 1 mg/kg IP, respectively, 2h + 18h post-insult)</td><td>Cortex</td><td>▲ iNOS, TNF-α, ▲ BBB permeability</td><td>-</td><td>(Amenta et al., 2014)</td></tr><tr><td></td><td>GP1a (1-5 mg/kg, IP, 10 min post-insult)</td><td>Cortex</td><td>▲ iNOS, TNF-α, IL-1β, IL-6, ▲ CCL2, CXCL10, ↗ IL-10, Arg-1, ▲ macrophage infiltration, ↗ M2 polarization (dose dep)</td><td>-</td><td>(Braun et al., 2018)</td></tr><tr><td>Traumatic Brain Injury (CCI, Rat)</td><td>JWH-133 (1.5 mg/kg, IP, 1h post-insult)</td><td>Cortex</td><td>▲ M2 microglial polarization</td><td>Reversed with SR144528 (3 mg/kg, IP)</td><td>(Li et al., 2022)</td></tr><tr><td>Traumatic brain injury (CHI, Mouse)</td><td>HU914 (0.1-10 mg/kg, IP, 1h post-insult)</td><td>Cortex and hippocampus</td><td>▲ TNF-α (hippocampus only)</td><td>-</td><td>(Magid et al., 2019)</td></tr><tr><td rowspan="2">Uveitis (Mouse)</td><td>HU308 (1.5%, topic, 15 min post-insult)</td><td>Eye</td><td>▲ neutrophil infiltration, ▲ endothelium adhering leukocytes</td><td>No effect in KO mice (Deltagen)</td><td>(Toguri et al., 2018)</td></tr><tr><td>RO6871304, RO6871085 (1.5%, topic), HU910 (3 mg/kg, IV) immediately after insult</td><td>Eye</td><td>▲ endothelium adhering leukocytes</td><td>No effect in KO mice (Deltagen)</td><td>(Porter et al., 2019)</td></tr></tbody></table>
<table><thead><tr><th></th><th></th><th></th><th></th><th></th><th></th><th></th></tr></thead><tbody><tr><td>Uveitis (Rat)</td><td>HU308 (1.5%, topic, 15 min post-insult)</td><td>Eye</td><td>∠ TNF-α, IL-1β, IL-6, IFN-γ, IL-10, NF-κB, AP-1, ∠ CCL5, CXCL2, ∠ endothelium adhering leukocytes, = VCAM, ICAM</td><td>Reversed with AM630 (2.5 mg/kg, IV)</td><td>(Toguri et al., 2014)</td></tr><tr><td>Vascular dementia (Rat)</td><td>Paeoniflorin (40 mg/kg/day, IP, from day 15 post-insult for 25 days)</td><td>Hippocampus</td><td>∠ p-mTOR, p-IκBα, pNF-κB p-p65, ∠ number of Iba1+ cells, ∠ M2 polarization</td><td>Reversed with AM630 (3 mg/kg, IP)</td><td>(Luo et al., 2018)</td></tr><tr><td colspan="6"><strong>CHRONIC CB2R ACTIVATION</strong></td></tr><tr><td>Alzheimer's disease (APP2576 Tg Mouse)</td><td>JWH-133 (0.2 mg/kg/day, drinking water from 7 months)</td><td>Cortex</td><td>∠ TNF-α, COX2, = IL-6, ∠ number of Iba1+ cells, = GFAP</td><td>-</td><td>(Martín-Moreno et al., 2012)</td></tr><tr><td rowspan="3">Alzheimer's disease (APP/PS1 Tg Mouse)</td><td>JWH-133 (0.2 mg/kg/day for 5 weeks, drinking water from 3 months)</td><td>Cortex</td><td>∠ IL-1β, TNF-α, IL-6, IL-10, SOD1, SOD2, ∠ number of Iba1+ cells, =GFAP</td><td>-</td><td>(Aso et al., 2013)</td></tr><tr><td>JWH-015 (0.15 mg/kg/day, IP, from 8 months, for 8 weeks)</td><td>Cortex</td><td>∠ Iba1+, ∠ M2 polarization</td><td>-</td><td>(Li et al., 2019)</td></tr><tr><td>MDA7 (14 mg/kg, IP, ½ day, from 3- to 8-month-old)</td><td>Hippocampus, entorhinal cortex</td><td>∠ Iba1</td><td>-</td><td>(Wu et al., 2017)</td></tr><tr><td>Amyotrophic lateral sclerosis (Mouse)</td><td>HU-308 (5 mg/kg, IP, daily from 65 to 90 days old)</td><td>Ventral and dorsal horn</td><td>∠ activated GFAP+ cells, ∠ Iba1+ cells</td><td>-</td><td>(Espejo-Porras et al., 2019)</td></tr><tr><td>Sub-chronic stress-induced inflammation (Mouse)</td><td>JWH-133 (2 mg/kg, IP, daily)</td><td>Frontal cortex</td><td>∠ TNF-α, iNOS, PGE2</td><td>No effect in KO mice (Zimmer)</td><td>(Zoppi et al., 2014)</td></tr><tr><td>Spontaneous hypertension-induced inflammation (Rat)</td><td>JWH-133 (1 mM, 10µL, icv, daily for 28 days)</td><td>Rostral ventrolateral medulla</td><td>∠ TNF-α, IL-6, IL-1β</td><td>Reversed with AAV2-r-CB2shRNA infusion</td><td>(Shi et al., 2020)</td></tr><tr><td colspan="6"><strong>PRE-INDUCTION CB2R BLOCKADE</strong></td></tr><tr><td>Stroke (Neonatal, Rat)</td><td>AM630 (1 mg/kg, IP, 1h pre-insult)</td><td>Peri hematoma brain</td><td>= IL-6, TNF-α, TGF-β, = number of Iba1+ cells</td><td>-</td><td>(Tang et al., 2016)</td></tr><tr><td colspan="6"><strong>POST-INDUCTION CB2R BLOCKADE</strong></td></tr><tr><td>Alzheimer's disease (intracortical Aβ<sub>1-42</sub>, Rat)</td><td>SR144528 (5 mg/kg, IP, 3, 5 and 7D post-insult)</td><td>Hippocampus</td><td>∠ GFAP</td><td>-</td><td>(Esposito et al., 2007)</td></tr><tr><td>Surgery-induced cognitive impairment (Mouse)</td><td>AM630 (3 mg/kg/day, IP, immediately post-insult for 7 days)</td><td>Hippocampus & PFC</td><td>∠ TNF-α, IL-1β, ∠ MCP1 (PFC only), ∠ CD11b</td><td>-</td><td>(Sun et al., 2017)</td></tr><tr><td>Status Epilepticus (Kainate, Mouse)</td><td>SMM-189 (6 mg/kg, IP, 1h post-insult)</td><td>Hippocampus & cortex</td><td>∠ IL-1β, IL-6, TNF-α, CCL2, CCL3, CCL4, ∠ Iba1, GFAP</td><td>-</td><td>(Yu et al., 2020)</td></tr><tr><td>Traumatic Brain Injury (air blast injury, Mouse)</td><td>SMM-189 (6 mg/kg, IP, 2h post-insult and daily for 14 days)</td><td>Optic tract</td><td>∠ MHC class II</td><td>-</td><td>(Reiner et al., 2014)</td></tr><tr><td rowspan="2">Traumatic Brain Injury (CCI, Mouse)</td><td>SR144528 (5 mg/kg, IP, 2h and 18h post-insult)</td><td>Cortex</td><td>∠ TNF-α, = ICAM</td><td>-</td><td>(Amenta et al., 2014)</td></tr><tr><td>AM630 (1-5 mg/kg, IP, 10 post-insult)</td><td>Cortex</td><td>= iNOS, TNF-α, IL-1β, IL-6, CCL2, CXCL10, = monocyte infiltration, no effect on polarization</td><td>-</td><td>(Braun et al., 2018)</td></tr><tr><td>Uveitis (Rat)</td><td>AM630 (2.5 mg/kg, IV, 15 min post-insult)</td><td>Eye</td><td>∠ NF-κB, = AP-1, ∠ TNF-α, IL-10, CXCL2, ∠ endothelium adhering leukocytes</td><td>-</td><td>(Toguri et al., 2014)</td></tr><tr><td>Vascular dementia (Rat)</td><td>AM630 (3 mg/kg/day, IP, day 15 post insult for 27 days)</td><td>Hippocampus</td><td>∠ p-mTOR, p-IκBα, pNF-κB p-p65, ∠ M1 polarization</td><td>-</td><td>(Luo et al., 2018)</td></tr></tbody></table>

**Abbreviations**: AAV2-r-CB2shRNA, adeno-associated virus expressing CB2-specific shRNA; Aβ, amyloid beta; AchE, Acetylcholinesterase; AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; AP-1, Activator protein 1; Arg-1, arginase 1; BBB, blood-brain barrier; BDNF, brain-derived neurotrophic factor; CB2R, cannabinoid receptor type 2; CCI, controlled cortical impact; CCL, Chemokine (C-C motif) ligand; CD, cluster of differentiation; CHI, closed-head injury; COX2, cyclooxygenase 2; CSF1R, Colony stimulating factor 1 receptor; CXCL, chemokine (C-X-C motif) ligand; CXCR, C-X-C chemokine receptor; EAE, Experimental autoimmune encephalomyelitis; ERK, Extracellular signal-regulated kinases; GFAP, Glial fibrillary acidic protein; GSH, reduced glutathione; HIV, human immunodeficiency virus; Iba1, ionized calcium-binding adapter molecule 1; ICAM, intercellular adhesion molecule; ICV, intracerebroventricular; IFN, interferon; IL, interleukin; IL1R1, Interleukin 1 receptor-like 1; iNOS, Inducible nitric oxide synthase; IP, intra-peritoneal; IV, intra-veinous; KO, knock-out; LPS, lipopolysaccharide; Mac1, Macrophage-1 antigen; MCAO, middle cerebral artery occlusion; MCP1, monocyte chemoattractant protein 1; MDA7, melanoma differentiation associated gene-7; MDMA, 3,4-methylenedioxy-N-methylamphetamine; MHC, major histocompatibility complex; MMP9, Matrix metallopeptidase 9; MPO, Myeloperoxidase; mTOR, mammalian Target of Rapamycin; MyD88, myeloid differentiation primary response 88; NF-κB, nuclear factor-kappa B; NS, non-significant; PGE2, prostaglandin E2; PFC, prefrontal cortex; PO, per os; PTZ, pentylenetetrazole; Ref, reference; SOD, superoxide dismutase; STAT, signal transducer and activator of transcription; STZ, Streptozotocin; Tg, transgenic; TGF, transforming growth factor; TLR, Toll-like receptor; TNF, tumor necrosis factor; VCAM, vascular cell adhesion molecule; VMH, ventromedial hypothalamus.
<table><caption>**Supplementary Table 3 – CB2R KO mice.** Outcome from CB2R KO mice (the "Deltagen strain", with a deletion of the sequence encoding the N-terminal part of the protein (The Jackson Laboratory, Cnr2<sup>tm1Dgen</sup>/J, #005786), the "Zimmer strain" with a deletion of the sequence encoding the C-terminal part of the protein (Buckley et al., 2000), and the "Romero strain" generated with the cre-lox method (López et al., 2018)) is either enhanced (↗), decreased (↘) or unchanged (=) when compared to wild-type littermates.</caption><thead><tr><th>CONDITION</th><th>ANIMAL MODEL</th><th>TISSUE</th><th>OUTCOME</th><th>REF</th></tr></thead><tbody><tr><td>α-Syn fibril induced inflammation</td><td>Deltagen CB2R KO mice + Intra NAc Fibrillar α-Syn</td><td>Nucleus Accumbens</td><td>↗ microglial activation, ↗CD68, C3aR, ↗M1 polarization</td><td>(Feng et al., 2022)</td></tr><tr><td>Amyotrophic lateral sclerosis</td><td>Romero CB2R KO mice x A315T</td><td>Spinal cord</td><td>↗ GFAP, Iba1 intensity signal, = TNF-α, IL-1β</td><td>(Rodríguez-Cueto et al., 2021)</td></tr><tr><td rowspan="5">Alzheimer's disease</td><td>Deltagen CB2R KO mice x J20 APP mice</td><td>Hippocampus</td><td>= Iba1</td><td>(Koppel et al., 2014)</td></tr><tr><td>Zimmer CB2R KO mice x APP/PS1 mice</td><td>Brain</td><td>↗ TNF-α, CCL2, Iba1, = Arg-1, ↘ Macrophage infiltration</td><td>(Schmôle et al., 2015)</td></tr><tr><td>Romero CB2R KO mice x 5xFAD mice</td><td>Hippocampus</td><td>= IL-1β, Iba1</td><td>(López et al., 2018)</td></tr><tr><td>Zimmer CB2R KO mice x APP/PS1 mice</td><td>Peri-plaques areas</td><td>↗ microglial activation (morphological complexity)</td><td>(Schmôle et al., 2018)</td></tr><tr><td>Romero CB2R KO mice x 5xFAD mice</td><td>Cortex</td><td>↗ Iba1</td><td>(Ruiz de Martín Esteban et al., 2022)</td></tr><tr><td>Age-related cognitive decline</td><td>Zimmer CB2R KO mice</td><td>Hippocampus</td><td>↗ Iba1, CD68, ↘ microglia soma size (18 months)</td><td>(Komorowska-Müller et al., 2021a)</td></tr><tr><td>Hypoxia-ischemia</td><td>Zimmer CB2R KO mice + carotid ligation</td><td>Brain</td><td>↗ GFAP, Iba1</td><td>(Kossatz et al., 2016)</td></tr><tr><td>Isoflurane-induced inflammation</td><td>Deltagen CB2R KO mice + Isoflurane exposure</td><td>Hippocampus</td><td>↗ Iba1, microglial branches, ↗ IL-6, TNF-α, iNOS, ↘ Ym1/2</td><td>(Li et al., 2021)</td></tr><tr><td>LPS-induced inflammation</td><td>Deltagen CB2R KO mice + LPS</td><td>Brain</td><td>= leukocyte/endothelium interaction</td><td>(Ramirez et al., 2012)</td></tr><tr><td>LPS+IFN γ-induced inflammation</td><td>Zimmer CB2R KO mice</td><td>Organotypic hippocampal slice cultures</td><td>↗ Iba1, CD68</td><td>(Reusch et al., 2022)</td></tr><tr><td>Multiple Sclerosis</td><td>Deltagen CB2R KO mice + experimental autoimmune encephalomyelitis</td><td>Spinal cord</td><td>↗ IL-1β</td><td>(Shao et al., 2014)</td></tr><tr><td>Neuropathic pain</td><td>Zimmer CB2R KO mice + sciatic nerve injury</td><td>Spinal cord</td><td>↗ GFAP, ↗ Iba1, activated phenotype</td><td>(Racz et al., 2008)</td></tr><tr><td>Parkinson's disease</td><td>Zimmer CB2R KO mice + intrastriatal LPS injection</td><td>Substansia Nigra</td><td>↗ CD68</td><td>(Gómez-Gálvez et al., 2016)</td></tr><tr><td>Sub-chronic stress-induced inflammation</td><td>Zimmer CB2R KO mice</td><td>Frontal cortex</td><td>↗ TNF-α, CCL2, iNOS, PGE<sub>2</sub></td><td>(Zoppi et al., 2014)</td></tr><tr><td>Traumatic Brain Injury</td><td>Deltagen CB2R KO mice + CCI injury</td><td>Cortex</td><td>↗ TNF-α, ICAM, ↗ iNOS, ↗ BBB permeability</td><td>(Amenta et al., 2014)</td></tr><tr><td rowspan="2">Uveitis</td><td>Deltagen CB2R KO mice + endotoxin induced uveitis</td><td>Eye</td><td>= neutrophil infiltration, ↗ endothelium adhering leukocytes</td><td>(Toguri et al., 2018)</td></tr><tr><td>Deltagen CB2R KO mice + endotoxin induced uveitis</td><td>Eye</td><td>↗ endothelium adhering leukocytes</td><td>(Porter et al., 2019)</td></tr></tbody></table>

**Abbreviations**: α-Syn, α-synuclein; Arg-1, arginase 1; BBB, blood-brain barrier; C3aR, complement component 3a receptor; CB2R, cannabinoid receptor type 2; CCI, controlled cortical impact; CCL, Chemokine (C-C motif) ligand; CD, cluster of differentiation; GFAP, Glial fibrillary acidic protein; Iba1, ionized calcium-binding adapter molecule 1; ICAM, intercellular adhesion molecule; IFN, interferon; IL, interleukin; iNOS, Inducible nitric oxide synthase; KO, knock-out; LPS, lipopolysaccharide; NAc, nucleus accumbens; PGE2, prostaglandin E2; Ref, reference; TNF, tumor necrosis factor.
<table><caption>**Supplementary Table 4. In vitro effects of CB2R ligands.** CB2R agonists are presented in green, CB2R inverse agonists are presented in red. Outcome following CB2R activation is either enhanced (▲), decreased (▼) or unchanged (=).</caption><thead><tr><th>TISSUE/CELLS</th><th>NEURONAL SUBTYPE</th><th>CB2R LIGAND (bath application)</th><th>OUTCOME</th><th>OTHER</th><th>REF</th></tr></thead><tbody><tr><td colspan="6"><strong>NEUROTRANSMITTERS RELEASE</strong></td></tr><tr><td>Hippocampus (Mouse)</td><td>GABA neurons (hippocampal synaptosomes)</td><td>WIN55,212-2, HU210 (0.3 µM, 9 min)</td><td>▲ evoked GABA release</td><td>-</td><td>(Andó et al., 2012)</td></tr><tr><td>Spinal Cord (Rat)</td><td>CGRP neurons (spinal cord slices)</td><td>AM1241 (1-300 nM, 10 min)</td><td>▲ capsaicin induced CGRP release (dose dep, up to 50% reduction)</td><td>Blocked by SR144528 (60 nM)</td><td>(Beltramo et al., 2006)</td></tr><tr><td>Substantia Nigra (Rat)</td><td>Glutamatergic neurons (substansia nigra slices)</td><td>GW833972A, GW405833, JWH-133 (1nM-1µM)</td><td>▲ K+-stimulated [³H]-Glutamate release (dose-dep)</td><td>Blocked by JTE-907 (100nM) and AM630 (200 nM)</td><td>(Sánchez-Zavaleta et al., 2018)</td></tr><tr><td>Dorsal striatum (Rat)</td><td>DA neurons (dorsal striatum slices)</td><td>GW833972A (100 nM)</td><td>▲ K+-stimulated [³H]-dopamine release (when D2 autoreceptors were blocked)</td><td>Blocked by AM630 (100 nM)</td><td>(López-Ramírez et al., 2020)</td></tr><tr><td colspan="6"><strong>NEUROTRANSMITTER RECEPTORS EXPRESSION</strong></td></tr><tr><td>Neuronal cell culture</td><td>CLU213</td><td>GP1a (1 nM), JWH-133 (30 nM) for 24h</td><td>▲5-HT2A expression</td><td>Blocked by AM630 (1µM)</td><td>(Franklin and Carrasco, 2012)</td></tr><tr><td colspan="6"><strong>IONIC CURRENTS</strong></td></tr><tr><td rowspan="2">mPFC (Rat)</td><td>Layer II/III pyramidal neurons (mPFC slices)</td><td>JWH-133 (1µM, 2-6 min)</td><td>Cl- current evokation</td><td>-</td><td>(den Boon et al., 2014)</td></tr><tr><td>Layer II/III pyramidal neurons (mPFC slices)</td><td>JWH-133 (1-5 µM, 15 min)</td><td>▲ intracellular [Ca²⁺]</td><td>-</td><td>(den Boon et al., 2012)</td></tr><tr><td rowspan="2">Retina (Rat)</td><td>Retinal ganglion cells (isolated neurons)</td><td>CB65 (500 nM), HU-308 (300nM)</td><td>= L-type Ca²⁺ current amplitudes. ▲ T-type Ca²⁺ current amplitude</td><td>Blocked by AM630 (100 nM)</td><td>(Qian et al., 2017)</td></tr><tr><td>Retinal ganglion cells (isolated neurons)</td><td>AM630 (100 nM)</td><td>No effect on [Ca²⁺] currents.</td><td>-</td><td>(Qian et al., 2017)</td></tr><tr><td>VTA (Mouse)</td><td>DA neurons (VTA slices)</td><td>JWH-133 (1µM, 10 min)</td><td>▲ M-type K⁺ current</td><td>Blocked by AM630 (10 µM), No effect in KO mice (Zimmer strain)</td><td>(Ma et al., 2019)</td></tr><tr><td colspan="6"><strong>MEMBRANE POTENTIAL</strong></td></tr><tr><td>mPFC (Rat)</td><td>Layer II/III pyramidal neurons (mPFC slices)</td><td>JWH-133 (1-5 µM), HU310 (1µM, 15 min)</td><td>Transient delayed depolarization</td><td>Blocked by SCH.356036 (5µM). No effect in KO mice (Deltagen strain)</td><td>(den Boon et al., 2012)</td></tr><tr><td>VTA (Mouse)</td><td>DA neurons (single isolated neurons)</td><td>JWH-133 (1µM)</td><td>Hyperpolarization</td><td>Blocked by AM630 (1 µM). No effect in KO mice (Zimmer strain)</td><td>(Zhang et al., 2014)</td></tr><tr><td>VTA (Rat)</td><td>DA neurons (single isolated neurons)</td><td>JWH-133 (1µM, 2 min)</td><td>Hyperpolarization</td><td>-</td><td>(Zhang et al., 2017)</td></tr><tr><td colspan="6"><strong>EXCITABILITY</strong></td></tr><tr><td rowspan="2">mPFC (Rat)</td><td>Layer II/III pyramidal neurons (mPFC slices)</td><td>JWH-133 (1-5 µM, 15 min)</td><td>▲ Excitability</td><td>-</td><td>(den Boon et al., 2012)</td></tr><tr><td>Layer II/III pyramidal neurons (mPFC slices)</td><td>SCH.356036 (5 µM, 15 min)</td><td>▲ Excitability</td><td>No effect in KO mice (Deltagen strain)</td><td>(den Boon et al., 2012)</td></tr><tr><td rowspan="2">Red nucleus (Mouse)</td><td>RN glutamatergic magnocellular neurons (midbrain slices)</td><td>JWH-133 (0.01-1 µM, 10 min)</td><td>▲ Firing rate (dose dep)</td><td>Blocked by AM630 (1 µM). No effect in KO mice (Zimmer strain)</td><td>(Zhang et al., 2021)</td></tr><tr><td>RN GABAergic neurons (midbrain slices)</td><td>JWH-133 (0.01-1 µM, 10 min)</td><td>= Firing rate</td><td>-</td><td>(Zhang et al., 2021)</td></tr><tr><td rowspan="2">SNc</td><td>DA neurons (SNc slices)</td><td>GW842166 (1 µM)</td><td>▲ autonomous firing rate</td><td>Blocked by AM630 pre incubation (10µM)</td><td>(Yu et al., 2021)</td></tr><tr><td>DA neurons (VTA slices)</td><td>JWH-133 (1-100µM, 10 min)</td><td>▲ Firing rate (dose dep)</td><td>-</td><td>(Ma et al., 2019)</td></tr><tr><td rowspan="2">VTA (Mouse)</td><td>DA neurons (single isolated neurons)</td><td>JWH-133 (1-10µM)</td><td>▲ AP firing rate, ▲ AP initiation, ▲ AP duration and ▲ AHP = ▲ excitability</td><td>Blocked by AM630 (1 µM). No effect in KO mice (Zimmer strain)</td><td>(Zhang et al., 2014)</td></tr><tr><td>DA neurons (VTA slices)</td><td>GW405833 (10µM)</td><td>▲ AP firing rate</td><td>Blocked by AM630 (1 µM). No effect in KO mice (Zimmer strain)</td><td>(Zhang et al., 2014)</td></tr><tr><td>VTA (Rat)</td><td>DA neurons (single isolated neurons)</td><td>JWH-133 (1µM, 2 min)</td><td>▲ AP firing rate, ▲ AHP</td><td>-</td><td>(Zhang et al., 2017)</td></tr><tr><td colspan="6"><strong>SYNAPTIC TRANSMISSION</strong></td></tr><tr><td>Cerebellum (Mouse)</td><td>Purkinje cells (cerebellum slices)</td><td>CB65, JWH-133 (0.1 µM, 5 min)</td><td>= eEPSC amplitude, ▲IPSC amplitude, ▲ post-synaptic GABA<sub>A</sub> receptor-mediated currents</td><td>Blocked by SR144528 (0.5 µM)</td><td>(Sadanandan et al., 2020)</td></tr><tr><td rowspan="2">Entorhinal cortex (Rat)</td><td>Layer II-V neurons (EC-hippocampal slices)</td><td>JWH-133 (50 nM)</td><td>▲ sIPSC amplitude</td><td>Blocked by AM630 (50 nM)</td><td>(Morgan et al., 2009)</td></tr><tr><td>Layer II-V neurons (EC-hippocampal slices)</td><td>JTE-907 (0.5-1 nM)</td><td>▲ sIPSC amplitude and frequency</td><td>-</td><td>(Morgan et al., 2009)</td></tr><tr><td rowspan="2">Hippocampus (Mouse)</td><td>CA3 pyramidal cells (hippocampal slices)</td><td>HU-308 (1µM, 10 min)</td><td>= eIPSC amplitude of DSI+ neurons, = fEPSPs</td><td>-</td><td>(Stempel et al., 2016)</td></tr><tr><td>CA1 pyramidal cells (hippocampal slices)</td><td>SR144528 (0.2 µM, 40-45 min)</td><td>No effect on fEPSPs amplitude</td><td>-</td><td>(Li and Kim, 2016a)</td></tr><tr><td rowspan="2">Hippocampus (Rat)</td><td>Hippocampal slices</td><td>β-Caryophyllene (50 µM 100 min)</td><td>▲ fEPSPs, ▲ LPS-induced intensification of LTD</td><td>-</td><td>(Hwang et al., 2020)</td></tr><tr><td>CA1 pyramidal cells (hippocampal slices)</td><td>JWH-133 (0.2 µM, 7-10 days)</td><td>= mIPSCs and eIPSC amplitude and frequency, = eEPSC amplitude, ▲ mEPSC frequency</td><td>Blocked by SR144528 (100 nM). No effect in KO mice (Deltagen strain)</td><td>(Kim and Li, 2015)</td></tr><tr><td></td><td>CA1 pyramidal cells (hippocampal slices)</td><td>GP1a (10 nM)</td><td>▲ mEPSC frequency</td><td>-</td><td>(Kim and Li, 2015)</td></tr><tr><td colspan="6"><strong>SYNAPTIC PLASTICITY</strong></td></tr><tr><td>Hippocampus (Mouse)</td><td>CA3 pyramidal cells (hippocampal slices)</td><td>HU-308 (1µM, 10 min)</td><td>Mimic AP-Driven long-lasting Hyperpolarization (= slow self-inhibition)</td><td>No effect in KO mice (Zimmer strain) nor in (Syn-)CB2R custom cKO mice.</td><td>(Stempel et al., 2016)</td></tr></tbody></table>