bluuebunny/crossref_metadata_2025 huggingface dataset:
Neuronal Cyclin-Dependent Kinase 5 Activity Is Critical for Survival
Authors: Tanaka Teruyuki , Veeranna  , Ohshima Toshio , Rajan Prithi , Amin Niranjana D. , Cho Andrew , Sreenath Taduru , Pant Harish C. , Brady Roscoe O. , Kulkarni Ashok B.
Published: April 2018

Cyclin-dependent kinase 5 (Cdk5) null mice exhibit a unique phenotype characterized by perinatal mortality, disrupted cerebral cortical layering attributable to abnormal neuronal migration, lack of cerebellar foliation, and chromatolytic changes of neurons in the brainstem and the spinal cord. Because Cdk5 is expressed in both neurons and astrocytes, it has been unclear whether this phenotype is primarily attributable to defects in neurons or in astrocytes. Herein we report reconstitution of Cdk5 expression in neurons in Cdk5 null mice and its effect on the null phenotype. Unlike the Cdk5 null mice, the reconstituted Cdk5 null mice that express the Cdk5 transgene under the p35 promoter (TgKO mice) were viable and fertile. Because Cdk5 expression is mainly limited to neurons in these mice and rescues the defects in the nervous system of the Cdk5 null phenotype, it clearly demonstrates that Cdk5 activity is necessary for normal development and survival of p35-expressing neurons.
Intracellular Signals That Control Cell Proliferation in Mammalian Balance Epithelia: Key Roles for Phosphatidylinositol-3 Kinase, Mammalian Target of Rapamycin, and S6 Kinases in Preference to Calcium, Protein Kinase C, and Mitogen-Activated Protein Kinase
Authors: Montcouquiol Mireille , Corwin Jeffrey T.
Published: April 2018

In fish, amphibians, and birds, the loss of hair cells can evoke S-phase entry in supporting cells and the production of new cells that differentiate as replacement hair cells and supporting cells. Recent investigations have shown that supporting cells from mammalian vestibular epithelia will proliferate in limited numbers after hair cells have been killed. Exogenous growth factors such as glial growth factor 2 enhance this proliferation most potently when tested on vestibular epithelia from neonates. In this study, the intracellular signaling pathways that underlie the S-phase entry were surveyed by culturing epithelia in the presence of pharmacological inhibitors and activators. The results demonstrate that phosphatidylinositol 3-kinase is a key element in the signaling cascades that lead to the proliferation of cells in mammalian balance epitheliain vitro. Protein kinase C, mammalian target of rapamycin, mitogen-activated protein kinase, and calcium were also identified as elements in the signaling pathways that trigger supporting cell proliferation.
Adaptive Electric Field Control of Epileptic Seizures
Authors: Gluckman Bruce J. , Nguyen Hanh , Weinstein Steven L. , Schiff Steven J.
Published: April 2018

We describe a novel method of adaptively controlling epileptic seizure-like events in hippocampal brain slices using electric fields. Extracellular neuronal activity is continuously recorded during field application through differential extracellular recording techniques, and the applied electric field strength is continuously updated using a computer-controlled proportional feedback algorithm. This approach appears capable of sustained amelioration of seizure events in this preparation when used with negative feedback. Seizures can be induced or enhanced by using fields of opposite polarity through positive feedback. In negative feedback mode, such findings may offer a novel technology for seizure control. In positive feedback mode, adaptively applied electric fields may offer a more physiological means of neural modulation for prosthetic purposes than previously possible.
Structural Determinants of Fast Desensitization and Desensitization–Deactivation Coupling in GABAAReceptors
Authors: Bianchi Matt T. , Haas Kevin F. , Macdonald Robert L.
Published: April 2018

Fast IPSCs in the brain are predominantly caused by presynaptic release of GABA that activates GABAAreceptor (GABAAR) channels. The IPSCs are shaped by the gating and desensitization properties of postsynaptic GABAARs. Specifically, fast desensitization has been suggested to decrease IPSC amplitude and to increase IPSC duration by slowing deactivation; however, the mechanisms underlying desensitization, deactivation, and their coupling are poorly understood. Consistent with this suggestion, α1β3γ2L GABAARs desensitize with a prominent fast phase and deactivate slowly, whereas α1β3δ GABAARs desensitize without a fast phase and deactivate rapidly. Using the concentration-jump technique applied to excised patches, we studied GABAARs containing chimeras or exchange mutants between δ and γ2L subunits to gain insight into the structural bases for fast desensitization and its coupling to deactivation. We demonstrated that the N terminus and two adjacent residues (V233, Y234) in the first transmembrane domain (TM1) of the δ subunit were both required to abolish fast desensitization. Additionally, these residues in TM1 of the γ2L subunit (Y235, F236) were critical for desensitized states to prolong deactivation after removal of GABA, because mutations resulted in accelerated deactivation despite unaltered desensitization time course. Interestingly, control of desensitization and deactivation was independent of the identity (γ2L or δ subunit sequence) of TM2, indicating that structures related to the putative channel gate may play a less direct role in desensitization than previously suggested.
β-Amyloid Stimulation of Microglia and Monocytes Results in TNFα-Dependent Expression of Inducible Nitric Oxide Synthase and Neuronal Apoptosis
Authors: Combs Colin K. , Karlo J. Colleen , Kao Shih-Chu , Landreth Gary E.
Published: April 2018

Reactive microglia associated with the β-amyloid plaques in Alzheimer's disease (AD) brains initiate a sequence of inflammatory events integral to the disease process. We have observed that fibrillar β-amyloid peptides activate a tyrosine kinase-based signaling response in primary mouse microglia and the human monocytic cell line, THP-1, resulting in production of neurotoxic secretory products, proinflammatory cytokines, and reactive oxygen species. We report that most of the amyloid-induced tyrosine kinase activity was stimulated after activation of Src family members such as Lyn. However, transduction of the signaling response required for increased production of the cytokines TNFα and IL1-β was mediated by the nonreceptor tyrosine kinase, Syk. Additionally, β-amyloid stimulated an NFκB-dependent pathway in parallel that was required for cytokine production. Importantly, TNFα generated by the monocytes and microglia was responsible for the majority of the neuorotoxic activity secreted by these cells after β-amyloid stimulation but must act in concert with other factors elaborated by microglia to elicit neuronal death. Moreover, we observed that the neuronal loss was apoptotic in nature and involved increased neuronal expression of inducible nitric oxide synthase and subsequent peroxynitrite production. Selective inhibitors of inducible nitric oxide synthase effectively protected cells from toxicity associated with the microglial and monocytic secretory products. This study demonstrates a functional linkage between β-amyloid-dependent activation of microglia and several characteristic markers of neuronal death occurring in Alzheimer's disease brains.
Assembly with the NR1 Subunit Is Required for Surface Expression of NR3A-Containing NMDA Receptors
Authors: Pérez-Otaño Isabel , Schulteis Christine T. , Contractor Anis , Lipton Stuart A. , Trimmer James S. , Sucher Nikolaus J. , Heinemann Stephen F.
Published: April 2018

Functional NMDA receptors are heteromultimeric complexes of the NR1 subunit in combination with at least one of the four NR2 subunits (A–D). Coexpression of NR3A, an additional subunit of the NMDA receptor family, modifies NMDA-mediated responses. It is unclear whether NR3A interacts directly with NR1 and/or NR2 subunits and how such association might regulate the intracellular trafficking and membrane expression of NR3A. Here we show that NR3A coassembles with NR1-1a and NR2A to form a receptor complex with distinct single-channel properties and a reduced relative calcium permeability. NR3A associates independently with both NR1-1a and NR2A in the endoplasmic reticulum, but only heteromeric complexes containing the NR1-1a NMDA receptor subunit are targeted to the plasma membrane. Homomeric NR3A complexes or complexes composed of NR2A and NR3A were not detected on the cell surface and are retained in the endoplasmic reticulum. Coexpression of NR1-1a facilitates the surface expression of NR3A-containing receptors, reduces the accumulation of NR3A subunits in the endoplasmic reticulum, and induces the appearance of intracellular clusters where both subunits are colocalized. Our data demonstrate a role for subunit oligomerization and specifically assembly with the NR1 subunit in the trafficking and plasma membrane targeting of the receptor complex.
The Basic Helix-Loop-Helix Genehesr2Promotes Gliogenesis in Mouse Retina
Authors: Satow Tetsu , Bae Soo-Kyung , Inoue Tomoyuki , Inoue Chihiro , Miyoshi Goichi , Tomita Koichi , Bessho Yasumasa , Hashimoto Nobuo , Kageyama Ryoichiro
Published: April 2018

Members of a subclass ofhairy/Enhancer of split[E(spl)] homologs, calledhesrgenes, are structurally related to another subclass ofhairy/E(spl)homologs,Hesgenes, which play an important role in neural development. To characterize the roles ofhesrgenes in neural development, we used the retina as a model system.In situhybridization analysis indicated that allhesrgenes are expressed in the developing retina, but onlyhesr2expression is associated spatially with gliogenesis. Each member was then misexpressed with retrovirus in the retinal explant cultures prepared from mouse embryos or neonates, which well mimicin vivoretinal development. Interestingly,hesr2but nothesr1orhesr3promoted gliogenesis while inhibiting rod genesis without affecting cell proliferation or death, suggesting that the cells that normally differentiate into rods adopted the glial fate by misexpression ofhesr2. The gliogenic activity ofhesr2was more profound when it was misexpressed postnatally than prenatally. In addition, double mutation of the neuronal determination genesMash1andMath3, which increases Müller glia at the expense of bipolar cells, upregulatedhesr2expression. These results indicate that, among structurally relatedhesrgenes, onlyhesr2promotes glial versus neuronal cell fate specification in the retina and that antagonistic regulation betweenhesr2andMash1–Math3may determine the ratios of neurons and glia.
Cardiotrophin-1, a Muscle-Derived Cytokine, Is Required for the Survival of Subpopulations of Developing Motoneurons
Authors: Oppenheim Ronald W. , Wiese Stefan , Prevette David , Armanini Mark , Wang Siwei , Houenou Lucien J. , Holtmann Bettina , Götz Rudolf , Pennica Diane , Sendtner Michael
Published: April 2018

Developing motoneurons require trophic support from their target, the skeletal muscle. Despite a large number of neurotrophic molecules with survival-promoting activity for isolated embryonic motoneurons, those factors that are required for motoneuron survival during development are still not known. Cytokines of the ciliary neurotrophic factor (CNTF)–leukemia inhibitory factor (LIF) family have been shown to play a role in motoneuron (MN) survival. Importantly, in mice lacking the LIFRβ or the CNTFRα there is a significant loss of MNs during embryonic development. Because genetic deletion of either (or both) CNTF or LIF fails, by contrast, to perturb MN survival before birth, it was concluded that another ligand exists that is functionally inactivated in the receptor deleted mice, resulting in MN loss during development. One possible candidate for this ligand is the CNTF–LIF family member cardiotrophin-1 (CT-1). CT-1 is highly expressed in embryonic skeletal muscle, secreted by myotubes, and promotes the survival of cultured embryonic mouse and rat MNs. Here we show thatct-1deficiency causes increased motoneuron cell death in spinal cord and brainstem nuclei of mice during a period between embryonic day 14 and the first postnatal week. Interestingly, no further loss was detectable during the subsequent postnatal period, and nerve lesion in young adultct-1-deficient mice did not result in significant additional loss of motoneurons, as had been previously observed in mice lacking both CNTF and LIF. CT-1 is the first bona fide muscle-derived neurotrophic factor to be identified that is required for the survival of subgroups of developing motoneurons.
The Untranslated Region of μ-Opioid Receptor mRNA Contributes to Reduced Opioid Sensitivity in CXBK Mice
Authors: Ikeda Kazutaka , Kobayashi Toru , Ichikawa Tomio , Kumanishi Toshiro , Niki Hiroaki , Yano Ryoji
Published: April 2018

It is well known that there are individual differences in a sensitivity to analgesics. Several lines of evidence have suggested that the level of opioid-induced analgesia is dependent on the level of expression of the μ-opioid receptor (μ-OR). However, the molecular mechanisms underlying the diversity of the level of the opioid receptor and the opioid sensitivity among individuals remain to be elucidated. In the present study, we analyzed the opioid-receptor genes of CXBK recombinant-inbred mice, which show reduced sensitivity to opioids. Northern blotting, nucleotide sequencing, andin situhybridization histochemical analyses demonstrated that CXBK mice possessed μ-OR mRNA with a normal coding region but an abnormally long untranslated region (UTR). In addition, the μ-OR mRNA level in CXBK mice was less than in the control mice. Next, we produced littermate mice that had inherited two copies of the wild-type μ-OR gene, had inherited two copies of the CXBK μ-OR gene, and had inherited both copies of the μ-OR genes. In these mice, inheritance of the CXBK μ-OR gene was well correlated with less μ-OR mRNA and reduced opioid effects on nociception and locomotor activity. We conclude that the CXBK μ-OR gene is responsible for the CXBK phenotypes. Because UTR differences are known to affect the level of the corresponding mRNA and protein and because UTRs are more divergent among individuals than coding regions, the present findings suggest that opioid sensitivity may vary, depending on different μ-OR levels attributable to divergent UTR of μ-OR mRNA.
Long-Range Temporal Correlations and Scaling Behavior in Human Brain Oscillations
Authors: Linkenkaer-Hansen Klaus , Nikouline Vadim V. , Palva J. Matias , Ilmoniemi Risto J.
Published: April 2018

The human brain spontaneously generates neural oscillations with a large variability in frequency, amplitude, duration, and recurrence. Little, however, is known about the long-term spatiotemporal structure of the complex patterns of ongoing activity. A central unresolved issue is whether fluctuations in oscillatory activity reflect a memory of the dynamics of the system for more than a few seconds.

We investigated the temporal correlations of network oscillations in the normal human brain at time scales ranging from a few seconds to several minutes. Ongoing activity during eyes-open and eyes-closed conditions was recorded with simultaneous magnetoencephalography and electroencephalography. Here we show that amplitude fluctuations of 10 and 20 Hz oscillations are correlated over thousands of oscillation cycles. Our analyses also indicated that these amplitude fluctuations obey power-law scaling behavior. The scaling exponents were highly invariant across subjects. We propose that the large variability, the long-range correlations, and the power-law scaling behavior of spontaneous oscillations find a unifying explanation within the theory of self-organized criticality, which offers a general mechanism for the emergence of correlations and complex dynamics in stochastic multiunit systems. The demonstrated scaling laws pose novel quantitative constraints on computational models of network oscillations. We argue that critical-state dynamics of spontaneous oscillations may lend neural networks capable of quick reorganization during processing demands.
Long-Lasting Cholinergic Modulation Underlies Rule Learning in Rats
Authors: Saar Drorit , Grossman Yoram , Barkai Edi
Published: April 2018

We studied the role of acetylcholine (ACh) in creating learning-related long-lasting modifications in the rat cortex. Rats were trained to discriminate positive and negative cues in pairs of odors, until they demonstrated rule learning and entered a mode of high capability for learning of additional odors. We have previously reported that pyramidal neurons in olfactory (piriform) cortex from trained rats had reduced spike afterhyperpolarization (AHP) for 3 d after rule learning. In the present study we examined the mechanism underlying this long-lasting modification. The cholinergic agonist carbachol reduced both slow AHP and firing adaptation in neurons from pseudotrained rats, but had no effect on neurons from trained rats, suggesting pre-existing cholinergic effect. Intracellular application of the calcium chelator BAPTA abolished the difference in slow AHP and in adaptation between groups, suggesting that the difference resulted from reduction in the ACh-sensitive, Ca2+-dependent potassium current,IAHP. At the behavioral level, application of the muscarinic blocker scopolamine before each training session delayed rule learning but had no effect on further acquisition of odor memory. We suggest that intense ACh activity during rule learning enhances neuronal excitability in the piriform cortex by reducingIAHPand that the effect outlasts the stage of rule learning, so that ACh activity is not crucial for further odor learning.
Stimulation of Nitric Oxide–cGMP Pathway Excites Striatal Cholinergic Interneurons via Protein Kinase G Activation
Authors: Centonze Diego , Pisani Antonio , Bonsi Paola , Giacomini Patrizia , Bernardi Giorgio , Calabresi Paolo
Published: April 2018

Conflicting data have been collected so far on the action of nitric oxide (NO) on cholinergic interneurons of the striatum. In the presentin vitroelectrophysiological study, we reported that intracellularly recorded striatal cholinergic interneurons are excited by both hydroxylamine andS-nitroso-N-acetylpenicillamine, two NO donors. This excitation persisted unchanged in the presence of glutamate, dopamine, and substance P receptor antagonists as well as after blockade of tetrodotoxin (TTX)- and calcium channel-sensitive transmitter release, suggesting that NO produces its effects by modulating directly resting ion conductances in the somatodendritic region of striatal cholinergic cells. The depolarizing effect of hydroxylamine was greatly reduced by lowering external concentrations of sodium ions (from 126 to 38 mm) and did not reverse polarity in the voltage range from −120 to −40 mV. The sodium transporter blockers bepridil and 3′,4′-dichlorobenzamil were conversely ineffective in preventing NO-induced membrane depolarization. Intracellular cGMP elevation is required for the action of hydroxylamine on striatal cholinergic cells, as demonstrated by the findings that the membrane depolarization produced by this pharmacological agent was prevented by bath and intracellular application of two inhibitors of soluble guanylyl cyclase and was mimicked and occluded by zaprinast, a cGMP phosphodiesterase inhibitor. Finally, intracellular Rp-8-Br-cGMPS, a protein kinase G (PKG) inhibitor, blocked the hydroxylamine-induced membrane depolarization of cholinergic interneurons, whereas both okadaic acid and calyculin A, two protein phosphatase inhibitors, enhanced it, indicating that intracellular PKG and phosphatases oppositely regulate the sensitivity of striatal cholinergic interneurons to NO. The characterization of the cellular mechanisms involved in the regulation of striatal interneuron activity is a key step for the understanding of the role of these cells in striatal microcircuitry.
Non-Associative Learning and Serotonin Induce Similar Bi-Directional Changes in Excitability of a Neuron Critical for Learning in the Medicinal Leech
Authors: Burrell Brian D. , Sahley Christie L. , Muller Kenneth J.
Published: April 2018

In studies of the cellular basis of learning, much attention has focused on plasticity in synaptic transmission in terms of transmitter release and the number or responsiveness of neurotransmitter receptors. However, changes in postsynaptic excitability independent of receptors may also play an important role. Changes in excitability of a single interneuron in the leech, the S-cell, were measured during non-associative learning of the whole-body shortening reflex. This interneuron was chosen because it is known to be necessary for sensitization and full dishabituation of the shortening response. During sensitization, S-cell excitability increased, and this enhancement corresponded to facilitation of the shortening reflex and increased S-cell activity during the elicited response. During habituation training, there was a decrement in both the shortening reflex and the elicited S-cell activity, along with decreased S-cell excitability. Conversely, dishabituation facilitated both the shortening response and S-cell activity during shortening, with an accompanying increase in S-cell excitability. Bath application of 1–10 μmserotonin (5HT), a modulatory neurotransmitter that is critical for sensitization, for full dishabituation, and for associative learning, increased S-cell excitability. S-cell excitability also increased after stimulation of the serotonergic Retzius cells. However, focal application of serotonin onto the S-cell soma hyperpolarized the interneuron, and bath application of a lower dose of serotonin (0.1 μm) decreased excitability. The observed changes in postsynaptic excitability appear to contribute to non-associative learning, and modulatory neurotransmitters, such as serotonin, evidently help regulate excitability. Such changes in S-cell excitability may also be relevant for more complex, associative forms of learning.
Semaphorin 3A–Vascular Endothelial Growth Factor-165 Balance Mediates Migration and Apoptosis of Neural Progenitor Cells by the Recruitment of Shared Receptor
Authors: Bagnard Dominique , Vaillant Catherine , Khuth Seng-Thuon , Dufay Nathalie , Lohrum Marion , Püschel Andreas W. , Belin Marie-Françoise , Bolz Jürgen , Thomasset Nicole
Published: April 2018

The dynamic and coordinated interaction between cells and their microenvironment controls cell migration, proliferation, and apoptosis, mediated by different cell surface molecules. We have studied the response of a neuroectodermal progenitor cell line, Dev, to a guidance molecule, semaphorin 3A (Sema3A), described previously as a repellent–collapsing signal for axons, and we have shown that Sema3A acts as a repellent guidance cue for migrating progenitor cells and, on prolonged application, induces apoptosis. Both repulsion and induction of cell death are mediated by neuropilin-1, the ligand-binding component of the Sema3A receptor. The vascular endothelial growth factor, VEGF165, antagonizes Sema3A-induced apoptosis and promotes cell survival, migration, and proliferation. Surprisingly, repulsion by Sema3A also depends on expression of VEGFR1, a VEGF165 receptor, expressed in Dev cells. Moreover, we found that these repulsive effects of Sema3A require tyrosine kinase activity, which can be attributed to VEGFR1. These results indicate that the balance between guidance molecules and angiogenic factors can modulate the migration, apoptosis (or survival), and proliferation of neural progenitor cells through shared receptors.
The NeuronalMicrotubule-Associated Protein 1BIs under Homeoprotein Transcriptional Control
Authors: Montesinos Marı́a Luz , Foucher Isabelle , Conradt Marcus , Mainguy Gaëll , Robel Laurence , Prochiantz Alain , Volovitch Michel
Published: April 2018

To identify genes regulated by homeoprotein transcription factors in postnatal neurons, the DNA-binding domain (homeodomain) of Engrailed homeoprotein was internalized into rat cerebellum neurons. The internalized homeodomain (EnHD) acts as a competitive inhibitor of Engrailed and of several homeoproteins (Mainguy et al., 2000). Analysis by differential display revealed thatmicrotubule-associated protein 1B(MAP1B) mRNA is upregulated by EnHD. This upregulation does not require protein synthesis, suggesting a direct effect of the homeodomain onMAP1Btranscription. The promoter region ofMAP1Bwas cut into several subdomains, and each subdomain was tested for its ability to bind Engrailed and EnHD and to associate with Engrailed-containing cerebellum nuclear extracts. In addition, the activity, and regulation by Engrailed, of each subdomain and of the entire promoter were evaluatedin vivoby electroporation in the chick embryo neural tube. These experiments demonstrate thatMAP1Bpromoter is regulated by Engrailedin vivo. Moreover, they show that one promoter domain that contains all ATTA homeoprotein cognate binding sites common to the rat and human genes is an essential element of this regulation. It is thus proposed that MAP1B, a cytoskeleton protein involved in neuronal growth and regeneration, is under homeoprotein transcriptional regulation.
Ezrin-Dependent Promotion of Glioma Cell Clonogenicity, Motility, and Invasion Mediated by BCL-2 and Transforming Growth Factor-β2
Authors: Wick Wolfgang , Grimmel Cornelia , Wild-Bode Christine , Platten Michael , Arpin Monique , Weller Michael
Published: April 2018

Ezrin belongs to the ezrin–radixin–moesin family proteins, which cross-link actin cytoskeleton and plasma membrane. Malignant glioma cells are paradigmatic for their strong migratory and invasive properties. Here, we report that the expression of dominant-negative ezrins inhibits clonogenicity, migration, and invasiveness of human malignant glioma cells. Furthermore, dominant-negative ezrins block hepatocyte growth factor (HGF)-mediated stimulation of clonogenicity and migration, without altering HGF-induced protein kinase B/Akt and focal adhesion kinase phosphorylation. Glioma cells expressing dominant-negative ezrins exhibit a shift of the BCL-2/BAX rheostat toward apoptosis, reduced αVβ3integrin expression and reduced matrix metalloproteinase (MMP) expression and activity. These changes are associated with a dramatic loss of transforming growth factor β2(TGF-β2) release. Exogenous supplementation of TGF-β2overcomes the inhibitory effects of dominant-negative ezrins on migration and clonogenicity. A neutralizing TGF-β2antibody mimics the effects of dominant-negative ezrins on clonogenicity and migration. Exogenous HGF markedly induces TGF-β2protein levels, and a neutralizing TGF-β2antibody abolishes the HGF-mediated increase in glioma cell motility. Finally, TGF-β2does not modulate BCL-2 or BAX expression, but BCL-2 gene transfer increases the levels of latent and active TGF-β2. Intracranial xenografts of U87MG glioma cells transfected with the dominant-negative ezrins in athymic mice grow to significantly smaller volumes, and the median survival of these mice is 50 d compared with 28 d in the control group. These data define a novel pathway for HGF-induced glioma cell migration and invasion, which requires ezrin, changes in the BCL-2/BAX rheostat, and the induction of TGF-β2expressionin vitro,and underscore the important role of HGF signalingin vivo.
Proliferation of NG2-Positive Cells and Altered Oligodendrocyte Numbers in the Contused Rat Spinal Cord
Authors: McTigue Dana M. , Wei Ping , Stokes Bradford T.
Published: April 2018

Given the numerous reparative roles glia may play after spinal cord injury (SCI), glial proliferation and cell number were examined in a model of traumatic SCI. Emphasis was placed on analysis of oligodendrocytes and NG2-positive (NG2+) cells, an endogenous cell population that may be involved in oligodendrocyte replacement. Overall, proliferation (assessed by bromodeoxyuridine incorporation) was markedly elevated during the first 2 weeks after injury and declined thereafter; a large portion of these dividing cells likely consisted of microglia–macrophages. Although the total number of NG2+ cells in the epicenter was reduced by half, we noted protracted proliferation in surviving NG2+ cells, with values sevenfold greater than in uninjured controls. Elevated proliferation of NG2+ cells persisted throughout the first 4 weeks after injury. However, the absolute number of NG2+ cells was not increased over controls, suggesting that the daughter cells either did not survive or they differentiated into other cell types. As expected, oligodendrocyte numbers were drastically altered after SCI. By 7 d after injury, the number of oligodendrocytes at the impact site was reduced by 93%. Despite ongoing tissue loss, the number of oligodendrocytes in spared tissue rose threefold at 14 d after injury. Although the function of NG2+ cells within the spinal cord is not completely understood, several studies suggest that they may differentiate into oligodendrocytes. Thus, proliferating NG2+ cells may contribute to the increased oligodendrocyte number observed at 2 weeks after injury. Future studies are required, however, to definitively determine the role NG2+ cells play in oligodendrocyte genesis, remyelination, and other post-injury events.
Loss of the Major GABAAReceptor Subtype in the Brain Is Not Lethal in Mice
Authors: Sur Cyrille , Wafford Keith A. , Reynolds David S. , Hadingham Karen L. , Bromidge Frances , Macaulay Alison , Collinson Neil , O'Meara Gillian , Howell Owain , Newman Richard , Myers Janice , Atack John R. , Dawson Gerard R. , McKernan Ruth M. , Whiting Paul J. , Rosahl Thomas W.
Published: April 2018

The α1β2γ2 is the most abundant subtype of the GABAAreceptor and is localized in many regions of the brain. To gain more insight into the role of this receptor subtype in the modulation of inhibitory neurotransmission, we generated mice lacking either the α1 or β2 subunit. In agreement with the reported abundance of this subtype, >50% of total GABAAreceptors are lost in both α1−/− and β2−/− mice. Surprisingly, homozygotes of both mouse lines are viable, fertile, and show no spontaneous seizures. Initially half of the α1−/− mice died prenatally or perinatally, but they exhibited a lower mortality rate in subsequent generations, suggesting some phenotypic drift and adaptive changes. Both adult α1−/− and β2−/− mice demonstrate normal performances on the rotarod, but β2−/− mice displayed increased locomotor activity. Purkinje cells of the cerebellum primarily express α1β2γ2 receptors, and in electrophysiological recordings from α1−/− mice GABA currents in these neurons are dramatically reduced, and residual currents have a benzodiazepine pharmacology characteristic of α2- or α3-containing receptors. In contrast, the cerebellar Purkinje neurons from β2−/− mice have only a relatively small reduction of GABA currents. In β2−/− mice expression levels of all six α subunits are reduced by ∼50%, suggesting that the β2 subunit can coassemble with α subunits other than just α1. Our data confirm that α1β2γ2 is the major GABAAreceptor subtype in the murine brain and demonstrate that, surprisingly, the loss of this receptor subtype is not lethal.
Neurotrophic Factors and Receptors in the Immature and Adult Spinal Cord after Mechanical Injury or Kainic Acid
Authors: Widenfalk Johan , Lundströmer Karin , Jubran Marie , Brené Stefan , Olson Lars
Published: April 2018

Delivery of neurotrophic factors to the injured spinal cord has been shown to stimulate neuronal survival and regeneration. This indicates that a lack of sufficient trophic support is one factor contributing to the absence of spontaneous regeneration in the mammalian spinal cord. Regulation of the expression of neurotrophic factors and receptors after spinal cord injury has not been studied in detail. We investigated levels of mRNA-encoding neurotrophins, glial cell line-derived neurotrophic factor (GDNF) family members and related receptors, ciliary neurotrophic factor (CNTF), and c-fos in normal and injured spinal cord. Injuries in adult rats included weight-drop, transection, and excitotoxic kainic acid delivery; in newborn rats, partial transection was performed. The regulation of expression patterns in the adult spinal cord was compared with that in the PNS and the neonate spinal cord. After mechanical injury of the adult rat spinal cord, upregulations of NGF and GDNF mRNA occurred in meningeal cells adjacent to the lesion. BDNF and p75 mRNA increased in neurons, GDNF mRNA increased in astrocytes close to the lesion, and GFRα-1 and truncated TrkB mRNA increased in astrocytes of degenerating white matter. The relatively limited upregulation of neurotrophic factors in the spinal cord contrasted with the response of affected nerve roots, in which marked increases of NGF and GDNF mRNA levels were observed in Schwann cells. The difference between the ability of the PNS and CNS to provide trophic support correlates with their different abilities to regenerate. Kainic acid delivery led to only weak upregulations of BDNF and CNTF mRNA. Compared with several brain regions, the overall response of the spinal cord tissue to kainic acid was weak. The relative sparseness of upregulations of endogenous neurotrophic factors after injury strengthens the hypothesis that lack of regeneration in the spinal cord is attributable at least partly to lack of trophic support.
Endogenous Brain-Derived Neurotrophic Factor and Neurotrophin-3 Antagonistically Regulate Survival of Axotomized Corticospinal NeuronsIn Vivo
Authors: Giehl Klaus M. , Röhrig Stephan , Bonatz Henk , Gutjahr Martin , Leiner Britta , Bartke Ilse , Yan Qiao , Reichardt Louis F. , Backus Carey , Welcher Andrew A. , Dethleffsen Kathrin , Mestres Pedro , Meyer Michael
Published: April 2018

Neuronal growth factors regulate the survival of neurons by their survival and death-promoting activity on distinct populations of neurons. The neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) promote neuronal survival via tyrosine kinase (Trk) receptors, whereas NGF and BDNF can also induce apoptosis in developing neurons through p75NTRreceptors in the absence of their respective Trk receptors. Using mutant mice and inactivation of neurotrophins and their receptors with antibodies in rats, we show that endogenous NT-3 induces death of adult BDNF-dependent, axotomized corticospinal neurons (CSNs). When NT-3 is neutralized, the neurons survive even without BDNF, suggesting complete antagonism. Whereas virtually all unlesioned and axotomized CSNs express both trkB and trkC mRNA, p75 is barely detectable in unlesioned CSNs but strongly upregulated in axotomized CSNs by day 3 after lesion, the time point when cell death occurs. Blocking either cortical TrkC or p75NTRreceptors alone prevents death, indicating that the opposing actions of NT-3 and BDNF require their respective Trk receptors, but induction of death depends on p75NTRcosignaling. The results show that neuronal survival can be regulated antagonistically by neurotrophins and that neurotrophins can induce neuronal death in the adult mammalian CNS. We further present evidence that signaling of tyrosine kinase receptors of thetrkfamily can be crucially involved in the promotion of neuronal deathin vivo.
Enhancement of Sensorimotor Behavioral Recovery in Hemiparkinsonian Rats with Intrastriatal, Intranigral, and Intrasubthalamic Nucleus Dopaminergic Transplants
Authors: Mukhida K. , Baker K. A. , Sadi D. , Mendez I.
Published: April 2018

One of the critical variables that influences the efficacy of clinical neural transplantation for Parkinson's disease (PD) is optimal graft placement. The current transplantation paradigm that focuses on ectopic placement of fetal grafts in the striatum (ST) fails to reconstruct the basal ganglia circuitry or normalize neuronal activity in important basal ganglia structures, such as the substantia nigra (SN) and the subthalamic nucleus (STN). The aim of this study was to investigate a multitarget neural transplantation strategy for PD by assessing whether simultaneous dopaminergic transplants in the ST, SN, and STN induce functional recovery in hemiparkinsonian rats. Forty-six female Wistar rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal pathway were randomly divided into eight groups and received lesions only or injections of 900,000 embryonic rat ventral mesencephalic cells in the (1) ST, (2) SN, (3) STN, (4) ST and SN, (5) ST, SN, and STN, (6) ST and STN, or (7) SN and STN. The number of cells transplanted was equally divided among grafting sites. Animals with two grafts received 450,000 cells in each structure, and animals with three grafts received 300,000 cells per structure. Recovery was assessed by amphetamine-induced rotations and the stepping tests. Graft survival was assessed using tyrosine hydroxylase immunohistochemistry. At 8 weeks after transplantation, simultaneous dopaminergic transplants in the ST, SN, and STN induced significant improvement in rotational behavior and stepping test scores. Intrastriatal transplants were associated with significant recovery of rotational asymmetry, whereas SN and STN transplants were associated with improved forelimb function scores. These results suggest that restoration of dopaminergic activity to multiple basal ganglia targets, such as the ST and SN, or the ST and STN, promotes a more complete functional recovery of complex sensorimotor behaviors. A multitarget transplant strategy aimed at optimizing dopaminergic reinnervation of the basal ganglia may be crucial in improving clinical outcomes in PD patients.
Analysis of the Activity-Deprived Zebrafish MutantmachoReveals an Essential Requirement of Neuronal Activity for the Development of a Fine-Grained Visuotopic Map
Authors: Gnuegge Lara , Schmid Susanne , Neuhauss Stephan C. F.
Published: April 2018

The formation of a retinotopic map is thought to involve an activity-independent molecular phase for early steps of both axon pathfinding and projection and a later phase in which cross talk between retinal ganglion cells (RGCs) and tectal neurons modifies and refines the neuronal connections. We report that the maturation of the retinotopic map in the zebrafish tectum involves activity-dependent processes. Zebrafish larvae mutant for the genemacho(mao) lack neuronal activity in RGCs and also display an enlarged retinotectal projection field but no significant increase in single axon length. This morphological defect can be phenocopied by raising larvae under TTX-induced neural impulse blockade. The effect of activity deprivation is dependent on the developmental stage. The projection phenotype inmaoas well as in the TTX-treated larvae develops between 4 and 6 d post-fertilization (dpf), after complete tectal coverage is first achieved. Electrophysiological recordings of RGCs in wild-type andmaozebrafish larvae reveal a temporally regulated reduction of sodium current in the mutant between 5 and 6 dpf. This coincides with the time of the axonal projection shifting on the tectum to compensate for the disparate growth patterns of the retina and the tectum. Our genetic and physiological analyses suggest a model in which neuronal activity in RGCs is needed for the establishment of morphological plasticity.
Neurotoxic Lesions of the Lateral Nucleus of the Amygdala Decrease Conditioned Fear But Not Unconditioned Fear of a Predator Odor: Comparison with Electrolytic Lesions
Authors: Wallace Karin J. , Rosen Jeffrey B.
Published: April 2018

Considerable evidence suggests that the lateral (LA) and basal (BA) nuclei of the amygdala are sites of plasticity and storage of emotional memory. Recent arguments, however, have seriously challenged this view, suggesting that the effects of amygdala lesions are attributable to interference with performance of fear behavior and not learning and memory. One way to address this controversy is to measure the same behavioral response during both conditioned and unconditioned fear. This is done in the present study by measuring fear-related freezing behavior after electrolytic and neurotoxic lesions of the LA or LA/BA nuclei in rats in a contextual fear conditioning paradigm and unconditioned fear to a predator odor. Electrolytic LA lesions attenuated post-shock freezing, retention test freezing, and freezing to the predator odor trimethylthiazoline (TMT). In contrast, excitotoxic NMDA lesions of the LA had no effect on post-shock freezing but significantly attenuated retention test freezing. Furthermore, excitotoxic LA lesions did not diminish freezing to TMT. Larger excitotoxic lesions that included the BA significantly reduced freezing in both the post-shock and retention tests but did not appreciably decrease freezing to TMT. The results suggest that the LA is important for memory of learned fear but not for generation of freezing behavior. In addition, the BA plays a role in freezing in conditioned fear situations but not in unconditioned fear. The studies suggest that the LA and BA play different roles in fear conditioning, but neither of them has a significant role in unconditioned freezing to a predator odor.
A Peripheral Mechanism for Behavioral Adaptation to Specific “Bitter” Taste Stimuli in an Insect
Authors: Glendinning John I. , Brown Hannah , Capoor Maya , Davis Adrienne , Gbedemah Amakoe , Long Eliza
Published: April 2018

Animals have evolved several chemosensory systems for detecting potentially dangerous foods in the environment. Activation of specific sensory cells within these chemosensory systems usually elicits an aversive behavioral response, leading to avoidance of the noxious foods. Although this aversive behavioral response can be adaptive, there are many instances in which it generates “false alarms,” causing animals to reject harmless foods. To minimize the number of false alarms, animals have evolved a variety of physiological mechanisms for selectively adapting their aversive behavioral response to harmless noxious compounds. We examined the mechanisms underlying exposure-induced adaptation to specific “bitter” compounds inManduca sextacaterpillars.M. sextaexhibits an aversive behavioral response to many plant-derived compounds that taste bitter to humans, including caffeine and aristolochic acid. This aversive behavioral response is mediated by three pairs of bitter-sensitive taste cells: one responds vigorously to aristolochic acid alone, and the other two respond vigorously to both caffeine and aristolochic acid. We found that 24 hr of exposure to a caffeinated diet desensitized all of the caffeine-responsive taste cells to caffeine but not to aristolochic acid. In addition, we found that dietary exposure to caffeine adapted the aversive behavioral response of the caterpillar to caffeine, but not to aristolochic acid. We propose that the adapted aversive response to caffeine was mediated directly by the desensitized taste cells and that the adapted aversive response did not generalize to aristolochic acid because the signaling pathway for this compound was insulated from that for caffeine.
Involvement of Spinal Protein Kinase Cγ in the Attenuation of Opioid μ-Receptor-Mediated G-Protein Activation after Chronic Intrathecal Administration of [d-Ala2,N-MePhe4,Gly-Ol5]Enkephalin
Authors: Narita Minoru , Mizoguchi Hirokazu , Narita Michiko , Nagase Hiroshi , Suzuki Tsutomu , Tseng Leon F.
Published: April 2018

The present study was designed to investigate the role of a protein kinase C (PKC) isoform in the uncoupling of the μ-opioid receptor from G-proteins after repeated intrathecal injection of a selective μ-receptor agonist, [d-Ala2,N-MePhe4,Gly-ol5]enkephalin (DAMGO), in the spinal cord of mice. The activation of G-proteins by opioids was measured by monitoring the guanosine-5′-o-(3-[35S]thio)triphosphate ([35S]GTPγS) binding. Mice were injected intrathecally with saline or DAMGO once a day for 1–7 d. At 24 hr after every injection the spinal cord membranes were prepared for the assay. The enhanced [35S]GTPγS binding by μ-agonists DAMGO, endomorphin-1, or endomorphin-2 was attenuated clearly in spinal cord membranes obtained from mice that were treated intrathecally with DAMGO for 5 and 7 d, but not for 1 or 3 d. By contrast, no change in levels of [35S]GTPγS binding induced by the δ-receptor agonist SNC-80 or κ-receptor agonist U-50,488H was noted in membranes obtained from mice that were treated with DAMGO. Concomitant intrathecal administration of a specific PKC inhibitor Ro-32-0432 with DAMGO blocked the attenuation of DAMGO-induced G-protein activation that was caused by chronic DAMGO treatment. Western blotting analysis showed that chronic DAMGO treatment increased the levels of PKCγ, but not PKCα, PKCβI, and PKCβII isoforms, in spinal cord membranes. Furthermore, mice lacking PKCγ failed to exhibit the desensitization of the DAMGO-stimulated [35S]GTPγS binding after repeated DAMGO injection. These findings indicate that repeated intrathecal administration of DAMGO may activate the PKCγ isoform and in turn cause a desensitization of μ-receptor-mediated G-protein activation in the mouse spinal cord.
Direct Visualization of the Movement of the Monomeric Axonal Transport Motor UNC-104 along Neuronal Processes in LivingCaenorhabditis elegans
Authors: Zhou H. Mimi , Brust-Mascher Ingrid , Scholey Jonathan M.
Published: April 2018

The formation and function of axons depends on the microtubule-based transport of cellular components from their sites of synthesis in the neuronal cell body to their sites of utilization at the axon terminus. To directly visualize this axonal transport in a living organism, we constructed transgenic lines ofCaenorhabditis elegansthat express green fluorescent protein fused to the monomeric synaptic vesicle transport motor, UNC-104. This UNC-104:: GFP construct rescued the Unc-104 mutant phenotype and was expressed throughout the nervous system. Using time-lapse confocal fluorescence microscopy, we were able to visualize fluorescent motor proteins moving in both directions along neuronal processes, some of which were identified definitely as axons and others as dendrites. Using kymograph analysis, we followed the movement of >900 particles. Most of them moved in one direction, but not necessarily at the same velocity. Ten percent of the observed particles reversed direction of movement during the period of observation, and 10% exhibited periods of movement interspersed with pauses. During episodes of persistent movement, particles moved at an average velocity of 1.02 μm/sec, which is close to thein vitrovelocity of microtubule gliding driven by purified monomeric kinesin at high motor density. To our knowledge, this is the first direct visualization and analysis of the movement of specifically labeled microtubule motor proteins along axonsin vivo.
A Common Exocytotic Mechanism Mediates Axonal and Dendritic Outgrowth
Authors: Martinez-Arca Sonia , Coco Silvia , Mainguy Gaëll , Schenk Ursula , Alberts Philipp , Bouillé Pascale , Mezzina Mauro , Prochiantz Alain , Matteoli Michela , Louvard Daniel , Galli Thierry
Published: April 2018

Outgrowth of the dendrites and the axon is the basis of the establishment of the neuronal shape, and it requires addition of new membrane to both growing processes. It is not yet clear whether one or two exocytotic pathways are responsible for the respective outgrowth of axons and dendrites. We have previously shown that tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP) defines a novel network of tubulovesicular structures present both at the leading edge of elongating dendrites and axons of immature hippocampal neurons developing in primary culture and that TI-VAMP is an essential protein for neurite outgrowth in PC12 cells. Here we show that the expression of the N-terminal domain of TI-VAMP inhibits the outgrowth of both dendrites and axons in neurons in primary culture. This effect is more prominent at the earliest stages of the development of neuronsin vitro. Expression of the N-terminal domain deleted form of TI-VAMP has the opposite effect. This constitutively active form of TI-VAMP localizes as the endogenous protein, particularly concentrating at the leading edge of growing axons. Our results suggest that a common exocytotic mechanism that relies on TI-VAMP mediates both axonal and dendritic outgrowth in developing neurons.