bluuebunny/crossref_metadata_2025_split huggingface dataset:
A loss of central serotonin in middle‐aged rats leads to increased blood pressure solely in males
Authors: Magnusson Jennifer , Li Aihua , Hodges Matthew , Cummings Kevin 
Published: November 2023

Serotonin (5‐Hydroxytryptamine, 5‐HT) neurons within the midline raphe project widely to autonomic regions in the brainstem, but the specific role of 5‐HT in arterial blood pressure (BP) regulation is unresolved. In addition, while the role of sex steroids in BP regulation has been extensively characterized, the extent to which they interact with 5‐HT signaling has not been investigated. Furthermore, 5‐HT neurons have a relatively high firing rate during wakefulness (QW) and non‐rapid eye movement sleep (NREM), but are virtually quiescent during rapid eye movement sleep (REM). Thus, 5‐HT's role in BP regulation may also be sleep‐state dependent. Therefore, this study seeks to clarify the specific role of central 5‐HT on BP regulation during sleep in young (3–4 months) and middle‐aged (12–13 months), male and female rats across vigilance states. We compared BP in wild‐type (WT; young: n=9 males, 9 females; middle aged: n=8 males, 7 females) and rats deficient in tryptophan hydroxylase 2 (TPH2−/−; young: n=8 males, 6 females; middle aged: n=5 males, 4 females), that lack central 5‐HT. Based on data from female adult TPH2−/− mice, we hypothesized that TPH2−/− rats would have lower BP than WT, and that the effects of 5‐HT deficiency on BP regulation would be most obvious in wakefulness and NREM sleep, when these neurons retain activity. Electroencephalography and electromyography were used to assess sleep state. Femoral arterial catheters were used to measure BP and heart rate. We also performed power spectral analysis of systolic BP and calculated the ratio of low to high frequency components (LF/HF) as a measure of sympathovagal balance to the vasculature. A loss of central 5‐HT had no effect on BP in young rats. However, middle‐aged male TPH2−/− rats had a significantly higher BP (128, 126, and 132 mmHg in wakefulness, NREM and REM, respectively) than male WT littermates (116, 117 and 122 mmHg) (p=0.015). In contrast, the BP of middle‐aged female TPH2−/− rats (102, 100, 105 mmHg in wakefulness, NREM and REM, respectively) was the same as female WT littermates (104, 105, 107 mmHg). The higher BP of middle‐aged male TPH2−/− rats was due to an increased diastolic BP (p=0.003). In addition, middle‐aged male TPH2−/− rats had an increased LF/HF ratio compared to WT controls (p<0.05). Overall, our data suggests that a loss of central 5‐HT leads to increased BP in male rats, possibly due to increased sympathetic tone to the vasculature. These findings also suggest that female sex steroids serve to protect against the deleterious effects of central 5‐HT deficiency on BP regulation.Support or Funding InformationFunding for this research was provided by an American Heart Association Scientist Development Grant (14SDG18560022; PI: KJC), and a University of Missouri, College of Veterinary Medicine Faculty Research Grant.
Acute Inhibition of HDACs after MI Leads to Low Inflammatory Profile: Implications for Post‐MI Angiogenesis
Authors: Veeranki Sudhakar , Tyagi Suresh C 
Published: November 2023

IntroductionAlthough the histone deacetylase inhibitors (HDACi) confer protection after myocardial injury (MI), the changes in the profiles of the angiogenic regulators and neoangiogenesis are unknown.HypothesisWe hypothesized that pan‐HDACi treatment confers positive angiogenic responses after MI, thereby improving the myocardial repair process.MethodsAll the mice underwent permanent LAD ligation. The mice were treated with PXD101, a HDACi, for a period of 3 days at a dose of 100 mg/kg, I.P., daily after the creation of MI. Mice were sacrificed after 72 hrs. of MI and the ischemic and non‐ischemic portions of heart tissue were collected. Relative levels of 53 angiogenesis‐related proteins were quantified by using angiogenic arrays. Heart function was evaluated before and after 72 hrs. of MI injury. Markers of inflammatory cells (macrophages and neutrophils) and endothelial progenitor cells (EPCs) were assayed in the ischemic tissues.ResultsPXD101 treatment did not ameliorate heart function after the 72 hrs. of treatment. There were specific changes in the levels of inflammatory markers such as CXCL1 and MCP‐1 after the treatment. Further assaying of various markers revealed that less inflammatory cell presence with moderate changes in the markers of EPCs in the ischemic tissues after the treatment. Deacetylation of non‐histone proteins were implicated in suppression of the inflammatory cell chemoattractants.ConclusionsOur study underscored the role of HDACi in regulation of myocardial angiogenic responses after MI injury. Acute inhibition of HDACs led to lower inflammatory profile. This study has pinpointed underlying epigenetic changes responsible for the neoangiogenesis and tissue regeneration after MI.Support or Funding InformationNIH grants: HL108621, HL074185 and DK104653.
Heterogeneous Stock Rats: A Model To Study The Genetics Of Despair And Fluoxetine Resistance in Adolescence
Authors: Woods Leah C Solberg , Holl Katie , He Hong , Wedemeyer Michael , Clopton Larissa , Wert Stephanie , Meckes Jeanie , Kastner Abigail , Palmer Abraham , Redei Eva 
Published: November 2023

Major depressive disorder is a complex illness caused by both genetics and the environment. Antidepressant resistance also has a genetic component. To date, very few genetic variations have been identified for major depression or antidepressant resistance. In the current study, we investigated whether outbred heterogeneous stock (HS) rats would be a suitable model to uncover the genetics of depression and its connection to antidepressant resistance. The WKY rat, one of the founders of the HS colony, is a known animal model of depression, in both adults and adolescents, and has been found to be resistant to fluoxetine anti‐depressant treatment. We therefore hypothesized that adolescent HS rats would exhibit variation in both despair behavior and response to fluoxetine treatment. We assessed despair behavior using a modified Forced Swim Test (FST) in four‐week old male and female HS rats. A separate group of rats were given sub‐acute treatment with either saline or fluoxetine prior to the FST and immobility, climbing and swimming behaviors were scored. Finally, we tested whether or not blood transcript levels previously identified to be differentially expressed in depressed human and rat models correlate with despair behavior in the HS rats. We demonstrate that 4‐week old HS rats have large variation in despair behavior in the FST and that most HS rats are resistant to treatment with fluoxetine. We also demonstrate that four genes previously identified as differentially expressed in the blood of depressed rats and humans are also negatively correlated with despair behavior in the HS rats. Taken together, these data demonstrate that the HS rat is a suitable model for investigating the underlying genetic basis of despair behavior as well as resistance to fluoxetine and that these studies will likely translate to depression in humans.Support or Funding InformationP50 DA037844, Davee Foundation
Metabolomic Analysis of Microdialysis Samples from the Frontal Association Cortex (FrA) of C57BL/6J (B6) Mouse during Isoflurane Anesthesia
Authors: Baer Aaron , Bourdon Allen K , Spano Giovanna , Bellesi Michele , Bradley Abram , Tononi Giulio , Cirelli Chiara , Serra Pier Andrea , Baghdoyan Helen A. , Campagna Shawn R. , Lydic Ralph 
Published: November 2023

Preliminary data have begun to characterize the metabolome in mouse FrA during sleep (FASEB J 30.1, 2016). Anesthesia and sleep are different states of consciousness that share a number of trait similarities. This ongoing study is quantifying the similarities and differences in the FrA metabolome during sleep and anesthesia. Microdialysis samples have been collected from the FrA of adult, male B6 mice (n=12) anesthetized with isoflurane. All experiments have adhered to FASEB policies on humane use of animals in research. The samples are being analyzed using ultra‐performance liquid chromatography coupled with high‐resolution mass spectrometry on an Exactive Plus Orbitrap instrument. An untargeted metabolomics method is being used to identify brain metabolites (Anal Chem 82:3212, 2010). To date we have analyzed 15 samples from 3 mice during isoflurane (1.1%) anesthesia. The results revealed approximately 90 known molecules in each 25 μL microdialysis sample. In FrA, 44 analytes were common to states of sleep and anesthesia, including myo‐inositol, tryptophan, homovanillic acid, uracil, and pyruvate. State‐specific differences in metabolomic measures included 37 metabolites present during anesthesia that were not observed during sleep, and absence of 6 metabolites during anesthesia that were found during sleep. Known metabolites common to both sleep and anesthesia were analyzed by partial least squares discriminant analysis (PLS‐DA). Variable importance projection scores calculated from PLS‐DA analyses highlighted 20 of the 44 metabolites present during sleep and anesthesia that contributed significantly to the observed separation in the PLS‐DA. Histological comparisons of FrA microdialysis sites during sleep and anesthesia are pending. These emerging results support the interpretation that metabolomic analyses provide a powerful approach for developing a neurochemical cartography of lower level phenotypes putatively regulating states of consciousness.Support or Funding InformationNIH grants HL65252 & Kavli Foundation (RL and HAB); MH099231 & 1P01NS083514 (CC and GT)
Early Life Exposure to Intermittent Hypoxia Induces Long‐Lasting Changes in Microglial Inflammatory Responses
Authors: Kiernan Elizabeth A , Ewald Andrea , Watters Jyoti 
Published: November 2023

Chronic intermittent hypoxia (IH), a hallmark of sleep‐disordered breathing, is prevalent in pediatric and pre‐term infant populations. IH increases reactive oxygen species formation, causes neuronal apoptosis, promotes peripheral and CNS inflammation, and results in long‐term neurocognitive deficits. However, little is known about cellular mechanisms underlying long‐term alterations in CNS function following early life IH exposure, nor if the activities of adult microglia, CNS resident innate immune cells, are also affected by early‐life IH. Previously, we observed that early life IH increased the microglial inflammatory response to a peripheral immune challenge the day following the last IH exposure, and that this occurred without changes in basal levels of microglial inflammation, suggesting that IH primes microglial responses acutely. It is unknown if these effects persist into adulthood, or if they are sex‐specific. To test the hypothesis that early life IH exposure will exert long‐lasting, sex‐specific effects on microglial inflammatory gene responses to an immune challenge in adulthood, we exposed male and female rat pups to 8 nights of IH (2 min cycles of 10.5% O2 for 8hrs/day) beginning on the first postnatal day (P1). We assessed microglial responses to a low‐dose peripheral lipopolysaccharide challenge (LPS; 0.1 mg/kg for 3hrs) either the day after the last IH exposure, or 6 weeks after. Using RNA sequencing (RNA‐Seq) and qRT‐PCR, we found in microglia from 9 day‐old neonates that IH alone downregulated multiple genes involved in metabolism, RNA processing and chromosome regulation, without changing basal expression of inflammatory genes such as IL‐1b and iNOS. Interestingly, microglial responses to acute LPS challenge in neonates and adults were different; microglial responses to an LPS challenge at 6 weeks of age were impaired in males. To begin to dissect potential chromatin changes (i.e. histone modifications) that may contribute to hypoxia‐induced alterations in microglial immune responses, we performed chromatin immunoprecipitation‐sequencing (ChIP‐Seq) in immortalized male microglia to evaluate genome‐wide histone 3 lysine 4 trimethylation (H3K4me3) changes, a mark of active gene transcription. We found increases in this mark at multiple genes involved in hypoxic and inflammatory responses, and decreases at genes downregulated by IH in vivo. Ongoing studies are aimed at resolving mechanisms whereby early life exposure to hypoxic challenges can exert epigenetic alterations that modulate the adult microglial inflammatory response. Understanding these mechanisms may enable us to identify and harness cellular mechanisms that can restore proper microglial immune responses in adult CNS disorders, many of which display sexual dimorphisms.Support or Funding InformationThis work was supported by NIH R01 NS085226
Characterization of Mago Homology B (Magohb) in Skeletal Muscle
Authors: Zubair Samira , Waddell David 
Published: November 2023

Skeletal muscle atrophy is a physiological condition that occurs due to immobilization, denervation, spinal cord transection and aging. Muscle wasting causes significant loss of muscle size and measurable reduction is strength. In order to better characterize the molecular genetic events of neurogenic atrophy, the gastrocnemius muscle was isolated from mice following 3 days and 14 days of sciatic nerve denervation. The gene expression profile in the denervated muscle tissue was then analyzed by microarray and compared to control muscle tissue in order to identify novel neurogenic atrophy‐induced genes. The microarray data revealed for the first time that Mago Homology B (Magohb), an exon junction core complex component involved in splicing and mRNA surveillance, is expressed in skeletal muscle and is significantly induced in response to denervation. In order to characterize the transcriptional regulation of Magohb, fragments of the proximal promoter located immediately upstream from the start of transcription were cloned and fused to a reporter gene. The reporter plasmids were then transfected into C2C12 mouse muscle cells alone or in combination with MyoD1 or myogenin expression plasmids, resulting in a differential effect on reporter activity. Furthermore, analysis of the promoter region of Magohb has revealed canonical E‐box elements, which are known myogenic regulatory factor (MRF) binding sites. The discovery that Magohb is expressed in skeletal muscle combined with the observation that this gene, a potential modulator of mRNA processing, is induced in response to neurogenic atrophy helps further our understanding of the molecular genetic events of skeletal muscle wasting.Support or Funding InformationThis work was supported by University of North Florida Transformational Learning Opportunity grants to D.W.
Preference, dependence, and pharmacokinetic outcomes in an opioid two bottle choice paradigm
Authors: Grim Travis Wiliam , Schmid Cullen L. , LaPrairie Robert B. , Park Jin‐Hong , Lin Li , Cameron Michael D. , Bohn Laura M. 
Published: November 2023

Non‐oral routes of administration are most common for illicit opioid abusers, while oral consumption remains the predominant method of intake for most prescription opioid abusers. Accordingly, preclinical oral self‐administration of opioids by rodents should be investigated to discern abuse liability in addition to the classical intravenous models. Two bottle choice (TBC) represents an assay which can quantify preference for one solution over another, and is one possible option to investigate oral abuse liability of opiates. Historically, C57BL6 and DBA strains have been compared against one another to assess the genetic determinants of oral self‐administration of morphine. Between strains, C57BL6J mice have been shown to prefer saccharin‐morphine solutions to tap water, and morphine solutions to quinine solutions, while DBA mice display the opposite phenotype. Additionally, C57BL6 mice can consume over 150 mg/kg/day of morphine versus quinine and naloxone (i.p.) following TBC can precipitate withdrawal. Here, we hypothesized that C57BL6 mice would prefer opioid‐saccharin to quinine‐saccharin solutions, and that naloxone would precipitate withdrawal after TBC. The potent opioid analgesic fentanyl was included to control for taste effects, and the non‐psychoactive, brain‐penetrant non‐steroidal anti‐inflammatory drug (NSAID) ketoprofen was included as a control analgesic which should lack reinforcing properties. To facilitate solubility of potential test compounds, a 1% DMSO and 0.5% Captisol solution was used in addition to other solutes. Using naloxone, withdrawal was precipitated, and plasma and brain levels of each test drug was quantified via mass spectrometry.Under these conditions, and similar to previous results, C57BL6 mice preferred morphine to quinine in a saccharin vehicle, but developed no preference when the concentration of quinine was lowered or included in both bottles. Similarly, fentanyl in the absence of quinine did not induce preference, and ketoprofen was preferred at the low dose but not the high dose. Naloxone precipitated withdrawal in morphine and fentanyl groups, but not ketoprofen, and all drugs were detected in plasma and brain. Together, these data demonstrate a reliable method to induce dependence via the oral self‐administration, and indicate the novel vehicle is amenable to TBC procedures. The preference of C57BL6 mice for morphine is likely driven by the enhancing palatability via saccharin in the morphine solution or via quinine in the opposite solution. Further optimization could reveal conditions in which morphine and/or fentanyl is preferred without enhancing palatability.Support or Funding InformationThis research was supported by NIH grant R01‐DA038964.
The Catalytic α Subunit of cAMP‐Dependent Protein Kinase Regulates Formoterol‐Induced Gene Expression in BEAS‐2B Human Bronchial Epithelial Cells
Authors: Hamed Omar , Yan Dong , Newton Robert , Giembycz Mark Anthony 
Published: November 2023

RationaleThe β2‐adrenoceptor is a GPCR that classically couples to Gsα, the activation of adenylyl cyclases and the formation of cAMP. Through distinct signalling complexes often containing cAMP‐dependent protein kinase (PKA), cAMP mediates a myriad of functional responses including the regulation of gene expression. We have shown previously that in the BEAS‐2B human airway epithelial cell line, long acting β2‐adrenoceptor agonists (LABAs) induce cAMP response element (CRE) dependent transcription, which we suggest contributes to their therapeutic activity in obstructive lung diseases. In the current study we have extended those findings by identifying the PKA catalytic subunit responsible for mediating gene expression induced by the LABA, formoterol.MethodsMicroarray, RT‐PCR, western blotting and gene silencing were used to identify and implicate PKA catalytic subunits in the regulation of LABA‐induced gene expression. A CRE reporter construct stably transfected into BEAS‐2B cells was used for this purpose together with an assessment of CREB phosphorylation and an analysis of a panel of cAMP‐inducible genes (NR4A3, CRISPLD2, RGS2, NR4A2, CD200 and IL‐6).ResultsMicroarray‐based gene expression profiling indicated that BEAS‐2B cells express mRNA transcripts for PRKACA (Cα), PRKACB (Cβ) and PRKX (Cx) whereas PRKACG (Cg) was absent. These data were confirmed at the protein level by western blotting. siRNAs were optimized to achieve 80 to 90% knockdown of PRKACA, PRKACB and PRKX, which was reproduced at the protein level. Under these conditions, silencing PRKACA was associated with a significant (~50%) abrogation of formoterol‐induced CRE reporter activation, CREB phosphorylation and gene expression. No additional effect was seen when siRNAs that target PRKACB or PRKX were used in combination with those directed towards PRKACA.ConclusionsThese data show that Cα is an essential cAMP signalling component controlling formoterol‐induced gene transcription in BEAS‐2B cells. The incomplete inhibition produced under conditions of 80–90% Cα knockdown implies that this PKA catalytic subunit is present in significant excess of that required to maximally mediate cAMP‐induced gene expression. This is an initial first step at identifying cAMP signalling components (adenylyl cyclases, scaffold proteins, phosphodiesterases and effectors) that control LABA‐induced functional responses in airway structural cells. We propose that identifying these components and their assembly into multiprotein signalling complexes could provide new opportunities for the development of novel therapies for asthma and chronic obstructive pulmonary disease.Support or Funding InformationAlberta Lung Association (ALA)
Cellular Compensation for Chronic Decreases in Cyclic AMP Concentration: Gαs‐Deficient Dendritic Cells as a Model and Implications for Therapeutics for Allergic Disorders
Authors: Chinn Amy M. , Lee Jihyung , Raz Eyal , Insel Paul A. 
Published: November 2023

Dendritic cells (DCs), the main antigen‐presenting cells, are involved in the induction of type II helper T cell (Th2) immune responses, but the mechanism by which this occurs is not fully elucidated. Mice that have a CD11c (DC)‐specific deletion of the gene Gnas (CD11cΔGnas), which encodes the α subunit of the heterotrimeric (αβγ) G protein Gαs, display very low basal cyclic AMP (cAMP) concentrations and an allergic phenotype; the decrease in cAMP and protein kinase A (PKA) activation biases DCs to provoke a Th2 response ex vivo and in vivo (Lee et al PNAS 112: 1529–34 [2015]). By comparing the mRNA expression of cAMP pathway components in WT and CD11cΔGnas bone marrow‐derived DCs (BMDCs), we assessed how cells compensate for a chronic decrease in cAMP/PKA signaling. Using quantitative real‐time PCR (qPCR), we found that BMDCs from CD11cΔGnas mice have >2‐fold increased expression of adenylyl cyclase (AC) 3, decreased expression of numerous cyclic nucleotide phosphodiesterase (PDE) isoforms that degrade cAMP, but no overall increase in Gαi proteins or the cAMP transporter MRP4/ABCC4. We also assessed the expression of G protein‐coupled receptors (GPCRs; assayed by qPCR‐based Taqman GPCR arrays), including those that regulate cAMP formation. A greater number and larger increase in expression occurs for GPCRs that couple to Gαi (rather than other Gα proteins) in CD11cΔGnas BMDCs compared to BMDCs from WT mice. Additionally, expression of multiple PKA subunits (Cα, RIα, RIIα, RIIβ) is decreased at least 30% in CD11cΔGnas BMDCs. A‐kinase anchor protein (AKAP)6 and AKAP9 have an increase and decrease, respectively, in those BMDCs. Together, these data indicate that DCs with lower cAMP levels have altered expression of “upstream” cAMP signaling components (GPCRs and AC3) and of “downstream” components that regulate cAMP levels (PDE isoforms) and actions (PKA subunits, AKAPs). Such changes in expression of GPCRs and post‐G protein components may reflect withdrawal of “basal”, tonic effects of cAMP, may contribute to the greater Th2 responses and allergic asthma of CD11cΔGnas mice, and may be novel therapeutic targets for allergic asthma.Support or Funding InformationSupported by NIH grants R56AI110505 and T32GM007752.