bluuebunny/crossref_metadata_2025 huggingface dataset:
TheseizureLocus Encodes theDrosophilaHomolog of the HERG Potassium Channel
Authors: Wang XinJing , Reynolds Elaine R. , Déak Péter , Hall Linda M.
Published: April 2018

Mutations in theseizure(sei) locus cause temperature-induced hyperactivity, followed by paralysis. Gene cloning studies have established that theseizuregene product is theDrosophilahomolog ofHERG, a member of theeagfamily of K+channels implicated in one form of hereditary long QT syndrome in humans. A series of five null alleles with premature stop codons are all recessive, but viable. A missense mutation in theseigene, which changes the charge at a conserved glutamate residue near the outer mouth of the pore, has a semidominant phenotype, suggesting that the mutant seizure protein acts as a poison in a multimeric complex. Transformation rescue of a null allele with a cDNA under the control of an inducible promoter demonstrates that induced expression of seizure potassium channels in adults rescues the paralytic phenotype. This rescue decays with at1/2of ∼1-1.5 d after gene induction is discontinued, providing the first estimate of ion channel stability in an intact, multicellular animal.
Reduced Acetylcholinesterase (AChE) Activity in Adrenal Medulla and Loss of Sympathetic Preganglionic Neurons in TrkA-Deficient, But Not TrkB-Deficient, Mice
Authors: Schober Andreas , Minichiello Liliana , Keller Markus , Huber Katrin , Layer Paul G. , Roig-López José L. , Garcı́a-Arrarás José E. , Klein Rüdiger , Unsicker Klaus
Published: April 2018

TrkA high-affinity receptors are essential for the normal development of sympathetic paravertebral neurons and subpopulations of sensory neurons. Paravertebral sympathetic neurons and chromaffin cells of the adrenal medulla share an ontogenetic origin, responsiveness to NGF, and expression of TrkA. Which aspects of development of the adrenal medulla might be regulated via TrkA are unknown. In the present study we demonstrate that mice deficient for TrkA, but not the neurotrophin receptor TrkB, show an early postnatal progressive reduction of acetylcholinesterase (AChE) enzymatic activity in the adrenal medulla and in preganglionic sympathetic neurons within the thoracic spinal cord, which are also significantly reduced in number. Quantitative determinations of specific AChE activity revealed a massive decrease (−62%) in the adrenal gland and a lesser, but still pronounced, reduction in the thoracic spinal cord (−40%). Other markers of the adrenal medulla and its innervation, including various neuropeptides, chromogranin B, secretogranin II, amine transporters, the catecholamine-synthesizing enzymes tyrosine hydroxylase and PNMT, synaptophysin, and L1, essentially were unchanged. Interestingly, AChE immunoreactivity appeared unaltered, too. Preganglionic sympathetic neurons, in contrast to adrenal medullary cells, do not express TrkA. They must, therefore, be affected indirectly by the TrkA knock-out, possibly via a retrograde signal from chromaffin cells. Our results suggest that signaling via TrkA, but not TrkB, may be involved in the postnatal regulation of AChE activity in the adrenal medulla and its preganglionic nerves.
Calpain Activation Contributes to Dendritic Remodeling after Brief Excitotoxic InjuryIn Vitro
Authors: Faddis Brian T. , Hasbani M. Josh , Goldberg Mark P.
Published: April 2018

The calcium-dependent protease calpain may contribute to neuronal death in acute neurological insults and may be activated very early in the neuronal injury cascade. We assessed the role of calpain in a model of rapid, reversible dendritic injury in murine cortical cultures. Brief sublethal NMDA exposure (10–30 μmfor 10 min) resulted in focal swellings, or varicosities, along the length of neuronal dendrites as visualized with the lipophilic membrane tracer DiI or with immunostaining using antibodies to the somatodendritic protein MAP2. These varicosities appeared within minutes of NMDA exposure and recovered spontaneously within 2 hr after NMDA removal. Addition of the calpain inhibitors MDL28,170, calpain inhibitors I and II, and leupeptin (all 1–100 μm) had little effect on the development of NMDA-induced dendrite injury. However, the resolution of varicosities was substantially delayed by addition of calpain inhibitors after sublethal excitotoxic exposure. Using Western blots and immunocytochemistry, we observed reactivity for a calpain-specific spectrin proteolytic fragment during the period of recovery from dendritic swelling, but not during its formation. Spectrin breakdown product immunoreactivity could be blocked by the calpain inhibitor MDL28,170 and appeared in neuronal cell bodies and neurites in a time course that paralleled dendritic recovery. These observations suggest that calcium-dependent proteolysis contributes to recovery of dendritic structure after NMDA exposure. Calpain activation is not necessarily detrimental and may play a role in dendritic remodeling after neuronal injury.
Tumor Necrosis Factor Enhances the Capsaicin Sensitivity of Rat Sensory Neurons
Authors: Nicol Grant D. , Lopshire John C. , Pafford Carl M.
Published: April 2018

The capacity of the proinflammatory cytokines, tumor necrosis factor α (TNFα) and interleukin 1β (IL-1β), to modulate the sensitivity of isolated sensory neurons grown in culture to the excitatory chemical agent capsaicin was examined. Alterations in capsaicin sensitivity were assessed by quantifying the number of neurons labeled with cobalt after exposure to capsaicin and by recording the whole-cell response from a single neuron to the focal application of capsaicin. A 24 hr pretreatment of the neuronal cultures with TNFα (10 or 50 ng/ml), but not IL-1β (10 or 50 ng/ml), produced a concentration-dependent increase in the number of cobalt-labeled neurons after exposure to 100 nmcapsaicin. The peak increase in the number of labeled neurons was attained after a 4 hr treatment with 10 ng/ml TNFα. Similarly, pretreatment with TNFα (10 ng/ml for 4, 12, and 24 hr) produced a greater than twofold increase in the average peak amplitude of the inward current evoked by 100 nmcapsaicin. Both the TNFα-induced increase in labeling and current amplitude were blocked by treating the neuronal cultures with indomethacin before the addition of TNFα. Enhancement of the capsaicin-evoked current also was blocked by the specific cyclo-oxygenase-2 inhibitor SC-236. These results indicate that TNFα can enhance the sensitivity of sensory neurons to the excitation produced by capsaicin and that this enhancement likely is mediated by the neuronal production of prostaglandins. Isolated sensory neurons grown in culture may prove to be a useful model system in which to explore how prolonged exposure to mediators associated with chronic inflammation alter the regulatory pathways that modulate the excitability of the nervous system.
EpileptogenesisIn VivoEnhances the Sensitivity of Inhibitory Presynaptic Metabotropic Glutamate Receptors in Basolateral Amygdala NeuronsIn Vitro
Authors: Neugebauer Volker , Keele N. Bradley , Shinnick-Gallagher Patricia
Published: April 2018

Modulation of excitatory synaptic transmission by presynaptic metabotropic glutamate receptors (mGluRs) was examined in brain slices from control rats and rats with amygdala-kindled seizures. Using whole-cell voltage-clamp and current-clamp recordings, this study shows for the first time that in control and kindled basolateral amygdala neurons, two pharmacologically distinct presynaptic mGluRs mediate depression of synaptic transmission. Moreover, in kindled neurons, agonists at either group II- or group III-like mGluRs exhibit a 28- to 30-fold increase in potency and suppress synaptically evoked bursting. The group II mGluR agonist (2S,3S,4S)-2-(carboxycyclopropyl)glycine (l-CCG) dose-dependently depressed monosynaptic EPSCs evoked by stimulation in the lateral amygdala with EC50values of 36 nm(control) and 1.2 nm(kindled neurons). The group III mGluR agonistl-2-amino-4-phosphonobutyrate (l-AP4) was less potent, with EC50values of 297 nm(control) and 10.8 nm(kindled neurons). The effects ofl-CCG andl-AP4 were fully reversible. Neitherl-CCG (0.0001–10 μm) norl-AP4 (0.001–50 μm) caused membrane currents or changes in the current–voltage relationship. The novel mGluR antagonists (2S,3S,4S)-2-methyl-2-(carboxycyclopropyl)-glycine (MCCG; 100 μm) and (S)-2-methyl-2-amino-4-phos-phonobutyrate (MAP4; 100 μm) selectively reversed the inhibition byl-CCG andl-AP4 to 81.3 ± 12% and 65.3 ± 6.6% of predrug, respectively. MCCG and MAP4 (100–300 μm) themselves did not significantly affect synaptic transmission. The exquisite sensitivity of agonists in the kindling model of epilepsy and the lack of evidence for endogenous receptor activation suggest that presynaptic group II- and group III-like mGluRs might be useful targets for suppression of excessive synaptic activation in neurological disorders such as epilepsy.
Role of NMDA and Non-NMDA Ionotropic Glutamate Receptors in Traumatic Spinal Cord Axonal Injury
Authors: Agrawal Sandeep K. , Fehlings Michael G.
Published: April 2018

We examined the role of glutamatergic mechanisms in acute injury to rat spinal cord white matter. Compound action potentials (CAPs) were recorded from isolated dorsal column segmentsin vitro. Under control conditions (Ringer’s solution), the CAPs decreased to 71.4 ± 2.0% of preinjury values after compression injury with a clip exerting a closing force of 2g. The combination of the NMDA receptor blocker APV (50 μm) and the AMPA/kainate (KA) receptor blocker CNQX (10 μm) resulted in significantly improved recovery of CAP amplitude postinjury; however, the NMDA receptor antagonist APV alone did not enhance postinjury recovery, and infusion of NMDA (10 μm) did not affect recovery of the CAPs. In contrast, the AMPA/KA receptor blockers NBQX (10 μm) or CNQX (10 μm) significantly enhanced the recovery of CAP amplitude postinjury. The agonists AMPA (100 μm) or KA (100 μm) resulted in significant attenuation of CAP amplitude postinjury. Coapplication of AMPA/KA plus NBQX and CNQX was also associated with improved functional recovery. After incubation with AMPA and KA, Co2+-positive glia were visualized in spinal cord white matter. Similar results were seen after compressive injury but not in control cords. Immunohistochemistry and Western blot analysis demonstrated AMPA (GluR4)- and KA (GluR6/7 and KA2)-positive astrocytes in spinal cord white matter. In summary, non-NMDA ionotropic glutamate receptors seem to be involved in the pathophysiology of traumatic spinal cord injury. The presence of AMPA (GluR4) and KA (GluR6/7 and KA2) receptors on periaxonal astrocytes suggests a role for these cells in glutamatergic white matter injury.
Selective Degeneration of Purkinje Cells with Lewy Body-Like Inclusions in Aged NFHLACZ Transgenic Mice
Authors: Tu Pang-hsien , Robinson Kathryn A. , de Snoo Femke , Eyer Joel , Peterson Alan , Lee Virginia M.-Y. , Trojanowski John Q.
Published: April 2018

Transgenic (NFHLacZ) mice expressing a fusion protein composed of a truncated high-molecular-weight mouse neurofilament (NF) protein (NFH) fused to β-galactosidase (LacZ) develop inclusions in neurons throughout the CNS. These inclusions persist from birth to advanced age and contain massive filamentous aggregates including all three endogenous NF proteins and the NFHLacZ fusion protein. Further, the levels of endogenous NF proteins are selectively reduced in NFHLacZ mice. Because these inclusions resemble NF-rich Lewy bodies (LBs) in Parkinson’s disease and LB dementia, we asked whether these lesions compromised the viability of affected neurons during aging. We studied hippocampal CA1 neurons, nearly all of which harbored inclusions (type I) devoid of cellular organelles, and cerebellar Purkinje cells, nearly all of which accumulated inclusions (type II) containing numerous entrapped organelles. Purkinje cells with type II inclusions began to degenerate in the NFHLacZ mice at ∼1 year of age, and most were eliminated by 18 months of age. In contrast, there was no significant loss of type I inclusion-bearing CA1 neurons with age. These data suggest that the sequestration of cellular organelles in type II inclusions may isolate and impair the function of these organelles, thereby rendering Purkinje cells selectively vulnerable to degeneration with age as in neurodegenerative diseases of the elderly characterized by accumulation of LBs.
Developmental Regulation of Synaptotagmin I, II, III, and IV mRNAs in the Rat CNS
Authors: Berton Frédérique , Iborra Cécile , Boudier Jeanne-Andrée , Seagar Michael J. , Marquèze Béatrice
Published: April 2018

Synaptotagmin I is an abundant synaptic vesicle protein that has an essential function in mediating Ca2+-triggered neurotransmitter release. We have analyzed the distribution of four neural synaptotagmin isoforms during postnatal development of the rat CNS byin situhybridization. Synaptotagmin I, II, III, and IV genes have distinct patterns of spatiotemporal expression except in cerebellum granule cells, where the four transcripts were detected during the formation of parallel fiber/Purkinje cell synapses. Throughout development synaptotagmin I mRNAs were widely expressed in brain, whereas synaptotagmin II transcripts were predominant in spinal cord. At all stages synaptotagmin III mRNAs were expressed uniformly in most neurons examined, although at a low level. Synaptotagmin I, II, and III gene expressions mainly increased during development and persisted in adulthood, mirroring neuronal differentiation. Conversely, synaptotagmin IV transcripts were predominant during perinatal development in a heterogeneous population of neurons and subsequently were expressed uniformly at a low level. Intense labeling was observed in the hippocampal CA3 field and in the subiculum, but not in the CA1 field, of the newborn rat. In cerebral cortex, lamina-specific labeling was detected with a high expression in cell layer V. Only a small number of Purkinje cell clusters were labeled in the flocculus and paraflocculus of the cerebellum. Heterogeneous sets of neurons expressing synaptotagmin IV gene also were observed in spinal cord. We thus speculate that synaptotagmin IV may a play a role in the development of the mammalian nervous system.
Disruption of the Metallothionein-III Gene in Mice: Analysis of Brain Zinc, Behavior, and Neuron Vulnerability to Metals, Aging, and Seizures
Authors: Erickson Jay C. , Hollopeter Gunther , Thomas Steven A. , Froelick Glenda J. , Palmiter Richard D.
Published: April 2018

Metallothionein-III (MT-III), a brain-specific member of the metallothionein family of metal-binding proteins, is abundant in glutamatergic neurons that release zinc from their synaptic terminals, such as hippocampal pyramidal neurons and dentate granule cells. MT-III may be an important regulator of zinc in the nervous system, and its absence has been implicated in the development of Alzheimer’s disease. However, the roles of MT-III in brain physiology and pathophysiology have not been elucidated. Mice lacking MT-III because of targeted gene inactivation were generated to evaluate the neurobiological significance of MT-III. MT-III-deficient mice had decreased concentrations of zinc in several brain regions, including hippocampus, but the pool of histochemically reactive zinc was not disturbed. Mutant mice exhibited normal spatial learning in the Morris water maze and were not sensitive to systemic zinc or cadmium exposure. No neuropathology or behavioral deficits were detected in 2-year-old MT-III-deficient mice, but the age-related increase in glial fibrillary acidic protein expression was more pronounced in mutant brain. MT-III-deficient mice were more susceptible to seizures induced by kainic acid and subsequently exhibited greater neuron injury in the CA3 field of hippocampus. Conversely, transgenic mice containing elevated levels of MT-III were more resistant to CA3 neuron injury induced by seizures. These observations suggest a potential role for MT-III in zinc regulation during neural stimulation.
Chronic Alcohol Consumption and Withdrawal Do Not Induce Cell Death in the Suprachiasmatic Nucleus, But Lead to Irreversible Depression of Peptide Immunoreactivity and mRNA Levels
Authors: Madeira M. D. , Andrade J. P. , Lieberman A. R. , Sousa N. , Almeida O. F. X. , Paula-Barbosa M. M.
Published: April 2018

There is evidence that chronic ethanol treatment (CET) disrupts the biological rhythms of various brain functions and behaviors. Because the suprachiasmatic nucleus (SCN) is widely recognized as the dominant pacemaker of the circadian system, we have examined the effects of CET and withdrawal on the main morphological features and chemoarchitecture of this hypothalamic nucleus. Groups of rats ethanol-treated for 6 and 12 months were compared with withdrawn rats (ethanol-treated for 6 months and then switched to a normal diet for an additional 6 months) and with groups of age-matched control and pair-fed control rats. The volume and the total number of neurons of the SCN were estimated from conventionally stained material, whereas the total number of astrocytes and of neurons containing vasopressin (AVP), vasoactive intestinal polypeptide (VIP), gastrin-releasing peptide (GRP), and somatostatin (SS) were estimated from immunostained sections. The estimates were obtained using unbiased stereological methods, based on Cavalieri’s principle and the optical fractionator. The volume of the SCN and the total number of SCN neurons and astrocytes did not vary among groups. We found, however, that CET induced a significant reduction in the total number of AVP-, VIP-, GRP-, and SS-containing neurons. Withdrawal from alcohol did not reduce but rather augmented the loss of VIP- and GRP-immunoreactive neurons. The CET-induced neurochemical alterations seem to result from a decrease in neuropeptide synthesis, as revealed by the reduction in AVP and VIP mRNA levels demonstrated byin situhybridization with radioactively labeled 48-mer AVP and 30-mer VIP probes. It is thus possible to conclude that the irreversible CET-induced changes in the neurochemistry of the SCN might underpin the disturbances in circadian rhythms observed after long-term alcohol consumption.
The SolubleN-Ethylmaleimide-Sensitive Factor Attached Protein Receptor Complex in Growth Cones: Molecular Aspects of the Axon Terminal Development
Authors: Igarashi Michihiro , Tagaya Mitsuo , Komiya Yoshiaki
Published: April 2018

SolubleN-ethylmaleimide-sensitive factor attached protein (SNAP) receptor (SNARE) mechanisms are thought to be involved in two important processes in axonal growth cones: (1) membrane expansion for axonal growth and (2) vesicular membrane fusion for mature synaptic transmission. We investigated the localization and interactions among the proteins involved in SNARE complex formation in isolated growth cone particles (GCP) from forebrain. We demonstrated that the SNARE complex is present in GCPs morphologically without synaptic vesicles (SVs) and associated with growth cone vesicles. However, the apparently SV-free GCP was lacking in the regulatory mechanisms inhibiting SNARE complex formation proposed in SV fusion, i.e., the association of synaptotagmin with the SNARE complex, and vesicle-associated membrane protein (VAMP)-synaptophysin complex formation. The core components of the SNARE complex (syntaxin, SNAP-25, and VAMP) accumulated for several days before postnatal day 7, when SVs first appeared, and preceded the accumulation of marker proteins such as synaptophysin, SV2, and V-ATPase. Our present results suggest that the SNARE mechanism for vesicular transmitter release is not fully functional in growth cones before the appearance of SVs, but the SNARE mechanism is working for membrane expansion in growth cones, which supports our recent report. We concluded that the regulation of the SNARE complex in growth cones is different from that in mature presynaptic terminals and that this switching may be one of the key steps in development from the growth cone to the presynaptic terminal.
Partition of Transient and Sustained Inhibitory Glycinergic Input to Retinal Ganglion Cells
Authors: Han Yi , Zhang Jian , Slaughter Malcolm M.
Published: April 2018

Physiological and pharmacological properties of possible subtypes of the native glycine receptor were investigated in retinal neurons using whole-cell voltage-clamp techniques. Two discrete inhibitory glycine responses were identified in ganglion cells. The responses could be distinguished pharmacologically: one was sensitive to strychnine and the other to 5,7-dichlorokynurenic acid. The two responses had different kinetics: the former had a fast onset and fast desensitization, whereas the latter had a slower onset and was much more sustained. The physiological and pharmacological distinctions suggest that the responses are mediated by different receptors. These receptors transduce glycinergic synaptic signals to ganglion cells, where they serve as low- and high-pass filters, respectively, of EPSPs.
Deactivation Retards Recovery from Inactivation in Shaker K+Channels
Authors: Kuo Chung-Chin
Published: April 2018

In Na+channels, recovery from inactivation begins with a delay, followed by an exponential course, and hyperpolarization shortens the delay as well as hastens the entire exponential phase. These findings have been taken to indicate that Na+channels must deactivate to recover from inactivation, and deactivation facilitates the unbinding of the inactivating particle. In contrast, it is demonstrated in this study that recovery from inactivation in Shaker K+channels begins with no delay on repolarization. Moreover, hyperpolarization hastens only the initial phase (fast component) of recovery yet retards the later phases of recovery by increasing the proportion of slow components. The time course of slow inward “tail” K+currents, which presumably result from the open state(s) traversed by the recovering inactivated channel, always matches the fast, but not the slow, components of recovery, suggesting that the fast and the slow components primarily correspond to recovery via the open state (unblocking of the inactivating particle before channel deactivation) and via the closed state (deactivation before unblocking), respectively. Besides, changing external K+concentration effectively alters the absolute value of the initial recovery speed, but not its voltage dependence. It seems that Shaker K+channel deactivation hinders, rather than facilitates, the unbinding of the inactivating particle and therefore retards recovery from inactivation, whereas external K+may enhance unbinding of the inactivating particle by binding to a site located near the external entrance of the pore.
Glutamate-Dependent Phosphorylation of Elongation Factor-2 and Inhibition of Protein Synthesis in Neurons
Authors: Marin Philippe , Nastiuk Kent L. , Daniel Nadine , Girault Jean-Antoine , Czernik Andrew J. , Glowinski Jacques , Nairn Angus C. , Prémont Joël
Published: April 2018

Postischemic delayed neuronal death is attributed to excitotoxic activation of glutamate receptors. It is preceded by a persistent inhibition of protein synthesis, the molecular basis of which is not known. Here we have examined in cortical neurons in culture the regulation by glutamate of phosphorylation of eukaryotic elongation factor-2 (eEF-2) by eEF-2 kinase, a Ca2+/calmodulin-dependent enzyme. Using a phosphorylation state-specific antibody, we show that glutamate, which triggers a large influx of Ca2+, enhances dramatically the phosphorylation of eEF-2. On the basis of kinetic and pharmacological analysis, we demonstrate a close correlation among the increase in cytosolic Ca2+concentration, the degree of eEF-2 phosphorylation, and the inhibition of protein synthesis. A 30 min treatment with NMDA induced a transient phosphorylation of eEF-2 and delayed neuronal death. However, pharmacological inhibition of protein translation was not neurotoxic by itself and protected neurons against the toxicity evoked by low concentrations of NMDA. Thus, phosphorylation of eEF-2 and the resulting depression of protein translation may have protective effects against excitotoxicity and open new perspectives for understanding long-term effects of glutamate.
Silent GABAASynapses during Flurazepam Withdrawal Are Region-Specific in the Hippocampal Formation
Authors: Poisbeau Pierrick , Williams Stephen R. , Mody Istvan
Published: April 2018

Whole-cell patch-clamp recordings were made from CA1 pyramidal and dentate gyrus granule cells (GCs) in hippocampal slices to assess the effects of withdrawal from chronic flurazepam (FRZ) treatment on the function of synaptic GABAAreceptors. In slices from control rats, acute perfusion of FRZ (30 μm) increased the monoexponential decay time constant of miniature IPSCs (mIPSCs) in CA1 and GCs (from 3.4 ± 0.6 to 7.6 ± 2.1 msec and from 4.2 ± 0.6 to 7.1 ± 1.8 msec, respectively) but did not change their mean conductance, 10–90% rise time, or frequency of occurrence. Withdrawal (2–5 d) from chronicin vivoFRZ treatment (40–110 mg/kg per day, per os) resulted in a dramatic loss of mIPSCs in CA1 neurons. On day 5 of withdrawal, no mIPSCs could be recorded in 40% of CA1 pyramidal cells. In the remaining 60% of the neurons, mIPSCs had a reduced mean conductance (from 0.78 ± 0.12 nS in vehicle-treated controls to 0.31 ± 0.05 nS) and a diminished frequency of occurrence (from 20.7 ± 7.9 to 4.1 ± 0.6 Hz). We have estimated that >80% of GABAAsynapses on CA1 pyramidal cells had become silent, whereas at still-active synapses the number of functional GABAAreceptor channels decreased by 60%. This reduction rapidly reverted to control levels on day 6 of withdrawal. FRZ withdrawal did not alter mIPSC properties in GCs. Our results are consistent with the hypothesis that chronic benzodiazepine treatment leads to a reduced number of functional synaptic GABAAreceptors in a region-specific manner that may stem from differences in the subunit composition of synaptic GABAAreceptors.
Downregulation of Tetrodotoxin-Resistant Sodium Currents and Upregulation of a Rapidly Repriming Tetrodotoxin-Sensitive Sodium Current in Small Spinal Sensory Neurons after Nerve Injury
Authors: Cummins Theodore R. , Waxman Stephen G.
Published: April 2018

Clinical and experimental studies have shown that spinal sensory neurons become hyperexcitable after axonal injury, and electrophysiological changes have suggested that this may be attributable to changes in sodium current expression. We have demonstrated previously that sodium channel α-III mRNA levels are elevated and sodium channel α-SNS mRNA levels are reduced in rat spinal sensory neurons after axotomy. In this study we show that small (C-type) rat spinal sensory neurons express sodium currents with dramatically different kinetics after axotomy produced by sciatic nerve ligation. Uninjured C-type neurons express both slowly inactivating tetrodotoxin-resistant (TTX-R) sodium current and a fast-inactivating tetrodotoxin-sensitive (TTX-S) current that reprimes (recovers from inactivation) slowly. After axotomy, the TTX-R current density was greatly reduced. No difference was observed in the density of TTX-S currents after axotomy, and their voltage dependence was not different from controls. However, TTX-S currents in axotomized neurons reprimed four times faster than control TTX-S currents. These data indicate that axotomy of spinal neurons is followed by downregulation of TTX-R current and by the emergence of a rapidly repriming TTX-S current and suggest that this may be attributable to the upregulation of a sodium channel isoform that was unexpressed previously in these cells. These axotomy-induced changes in sodium currents are expected to alter excitability substantially and could underlie the molecular pathogenesis of some chronic pain syndromes associated with injury to the axons of spinal sensory neurons.
Relationship between the Development of Outer Hair Cell Electromotility and Efferent Innervation: A Study in Cultured Organ of Corti of Neonatal Gerbils
Authors: He David Z. Z.
Published: April 2018

Outer hair cell (OHC) electromotility, which powers the cochlear amplifier, develops at a later stage of hearing ontogeny. There has been speculation whether efferents play a necessary role in directing or achieving OHC maturation in mammals. In this study, we examine whether the development of OHC motility depends on the establishment of efferent innervation of the cells’ synaptic pole by measuring electromotility of OHCs grown in cultures, deprived of efferent innervation. Tissue cultures of the organ of Corti were prepared from the cochleas of newborn gerbils. Solitary OHCs were obtained from 4- to 15-d-old cultures by enzymatic digestion and mechanical trituration. Length changes evoked by transcellular electrical stimulation were detected and measured with a photodiode sensor. Results show that OHCs develop electromotility between 6 and 13 d in culture without the presence of efferent innervation. The timetable for the onset of OHC electromotility is comparable with thatin vivo. This demonstrates that the ontogeny of OHC electromotility is an intrinsic process that does not require the influence of efferent innervation.
Stop and Branch Behaviors of Geniculocortical Axons: A Time-Lapse Study in Organotypic Cocultures
Authors: Yamamoto Nobuhiko , Higashi Shuji , Toyama Keisuke
Published: April 2018

The behavior of growing thalamic axons was studied in an organotypic coculture of the lateral geniculate nucleus (LGN) with the visual cortex (VC) to reveal cellular interactions that underlie the formation of lamina-specific thalamocortical connections. The LGN explant was placed at the ventral side, pial surface, or lateral edge of the VC explant, and fluorescent dye-labeled LGN axons were observed by confocal microscopy in fixed and living tissue. The axonal projection pattern in fixed cocultures after 1 weekin vitrodemonstrated that, in all three configurations, LGN axons formed primitive branches mainly in layer 4. A time-lapse study further examined axonal growth and branch formation in the living cortical explant. The majority of branches emerged within layer 4 behind the axonal tip, regardless of the direction of axonal entry. In addition, most axons entering from the ventral or pial side of the VC exhibited a transient or persistent stop of axonal growth in and around layer 4, whereas those entering from the lateral edge of the VC traveled along layer 4 without exhibiting stop behavior. The axonal stop often was accompanied by growth cone collapse and a slight retraction. These results suggest the existence of branch and stop cues in layer 4 of the cortex that are recognized by LGN axons.
Dentate Granule Cell Neurogenesis Is Increased by Seizures and Contributes to Aberrant Network Reorganization in the Adult Rat Hippocampus
Authors: Parent Jack M. , Yu Timothy W. , Leibowitz Rebecca T. , Geschwind Daniel H. , Sloviter Robert S. , Lowenstein Daniel H.
Published: April 2018

The dentate granule cell layer of the rodent hippocampal formation has the distinctive property of ongoing neurogenesis that continues throughout adult life. In both human temporal lobe epilepsy and rodent models of limbic epilepsy, this same neuronal population undergoes extensive remodeling, including reorganization of mossy fibers, dispersion of the granule cell layer, and the appearance of granule cells in ectopic locations within the dentate gyrus. The mechanistic basis of these abnormalities, as well as their potential relationship to dentate granule cell neurogenesis, is unknown. We used a systemic chemoconvulsant model of temporal lobe epilepsy and bromodeoxyuridine (BrdU) labeling to investigate the effects of prolonged seizures on dentate granule cell neurogenesis in adult rats, and to examine the contribution of newly differentiated dentate granule cells to the network changes seen in this model. Pilocarpine-induced status epilepticus caused a dramatic and prolonged increase in cell proliferation in the dentate subgranular proliferative zone (SGZ), an area known to contain neuronal precursor cells. Colocalization of BrdU-immunolabeled cells with the neuron-specific markers turned on after division, 64 kDa, class III β-tubulin, or microtubule-associated protein-2 showed that the vast majority of these mitotically active cells differentiated into neurons in the granule cell layer. Newly generated dentate granule cells also appeared in ectopic locations in the hilus and inner molecular layer of the dentate gyrus. Furthermore, developing granule cells projected axons aberrantly to both the CA3 pyramidal cell region and the dentate inner molecular layer. Induction of hippocampal seizure activity by perforant path stimulation resulted in an increase in SGZ mitotic activity similar to that seen with pilocarpine administration. These observations indicate that prolonged seizure discharges stimulate dentate granule cell neurogenesis, and that hippocampal network plasticity associated with epileptogenesis may arise from aberrant connections formed by newly born dentate granule cells.
Distribution of Fos-Like Immunoreactivity in the Medullary Reticular Formation of the Rat after Gustatory Elicited Ingestion and Rejection Behaviors
Authors: DiNardo Lisa A. , Travers Joseph B.
Published: April 2018

The distribution of neurons in the medullary reticular formation (RF) activated by the ingestion of sucrose or rejection of quinine was examined using standard immunohistochemical techniques to detect the expression of the Fos protein product of the immediate-early gene c-fos. Double-labeling techniques were used to gain further insight into the possible functional significance of RF neurons exhibiting Fos-like immunoreactivity (FLI). Compared with sucrose and unstimulated controls, quinine elicited significantly more FLI neurons in three specific RF subdivisions: parvocellular reticular nucleus (PCRt), intermediate reticular nucleus (IRt), and dorsal medullary reticular nucleus (MdD). Moreover, the number of FLI neurons in the RF of quinine-stimulated animals was significantly correlated with the degree of oromotor activity. Thus, the distinct distribution of FLI neurons throughout the RF after quinine may reflect the activation of a specific oral rejection circuit. The double-labeling results indicated a high degree of segregation between FLI neurons and premotor projection neurons to the hypoglossal nucleus (mXII) retrogradely labeled with Fluorogold. Thus, although there were a significant number of double-labeled neurons in the RF, the major concentration of premotor projection neurons to mXII in IRt were medial to the preponderance of FLI neurons in the PCRt. In contrast, there was substantial overlap between FLI neurons in the RF and labeled fibers after injections of the anterograde tracer, biotinylated dextran into the rostral (gustatory) portion of the nucleus of the solitary tract. These results support a medial (premotor)/lateral (sensory) functional topography of the medullary RF.
Dissociation of Tolerance and Dependence for Opioid Peripheral Antinociception in Rats
Authors: Aley K. O. , Levine J. D.
Published: April 2018

Repeated peripheral administration of the μ-opioid agonist [d-Ala2,N-Me-Phe4,gly5-ol] enkephalin (DAMGO) produces acute tolerance and dependence on its peripheral antinociceptive effect against prostaglandin E2(PGE2)-induced mechanical hyperalgesia. In this study we evaluated the roles of protein kinase C (PKC) and nitric oxide (NO) in the development of this tolerance and dependence. Repeated administration of PKC inhibitors chelerythrine and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride with DAMGO did not alter the tolerance to DAMGO; however, dependence (defined as naloxone-induced withdrawal hyperalgesia) was blocked. Repeated administration ofN-(n-heptyl)-5-chloro-1-naphthalenesulfonamide, a PKC activator, which alone did not produce tolerance, mimicked the dependence produced by DAMGO. Repeated administration of the NO synthase inhibitorNG-methyl-l-arginine with DAMGO blocked the development of tolerance to DAMGO but had no effect on the development of dependence. Repeated administration ofl-arginine, a NO precursor, mimicked tolerance produced by repeated administration of DAMGO (i.e., the antinociceptive effect of DAMGO was lost); however,l-arginine did not mimic dependence. These findings suggest that the development of acute tolerance and dependence on the peripheral antinociceptive effects of DAMGO have different, dissociable mechanisms. Specifically, PKC is involved in development of μ-opioid dependence, whereas the NO signaling system is involved in the development of μ-opioid tolerance.
Learning and Generalization of Auditory Temporal–Interval Discrimination in Humans
Authors: Wright Beverly A. , Buonomano Dean V. , Mahncke Henry W. , Merzenich Michael M.
Published: April 2018

The sensory encoding of the duration, interval, and order of different stimulus features provides vital information to the nervous system. The present study focuses on the influence of practice on auditory temporal–interval discrimination. The goals of the experiment were to determine (1) whether practice improved the ability to discriminate a standard interval of 100 msec bounded by brief 1 kHz tones from longer intervals, and, if so, (2) whether this improvement generalized to different tonal frequencies or temporal intervals. Learning was examined in 14 human subjects using an adaptive, two-alternative, forced-choice procedure. One hour of training per day for 10 d led to marked improvements in the ability to discriminate between the standard and longer intervals. The generalization of learning was evaluated by independently varying the spectral (tonal frequency) and temporal (interval) components of the stimuli in four conditions tested both before and after the training phase. Remarkably, there was complete generalization to the trained interval of 100 msec bounded by tones at the untrained frequency of 4 kHz, but no generalization to the untrained intervals of 50, 200, or 500 msec bounded by tones at the trained frequency of 1 kHz. Thus, these data show that (1) temporal–interval discrimination using a 100-msec standard undergoes perceptual learning, and (2) the neural mechanisms underlying this learning are temporally, but not spectrally, specific. These results are compared with those from previous investigations of learning in visual spatial tasks, and are discussed in relation to biologically plausible models of temporal processing.
5HT4Receptors Couple Positively to Tetrodotoxin-Insensitive Sodium Channels in a Subpopulation of Capsaicin-Sensitive Rat Sensory Neurons
Authors: Cardenas Carla G. , Del Mar Lucinda P. , Cooper Brian Y. , Scroggs Reese S.
Published: April 2018

The distribution of tetrodotoxin (TTX)-sensitive and -insensitive Na+currents and their modulation by serotonin (5HT) and prostaglandin E2(PGE2) was studied in four different types of dorsal root ganglion (DRG) cell bodies (types 1, 2, 3, and 4), which were previously identified on the basis of differences in membrane properties (Cardenas et al., 1995). Types 1 and 2 DRG cells expressed TTX-insensitive Na+currents, whereas types 3 and 4 DRG cells exclusively expressed TTX-sensitive Na+currents.

Application of 5HT (1–10 μm) increased TTX-insensitive Na+currents in type 2 DRG cells but did not affect Na+currents in type 1, 3, or 4 DRG cells. The 5HT receptor involved resembled the 5HT4subtype. It was activated by 5-methoxy-N,N-dimethyltryptamine (10 μm) but not by 5-carboxyamidotryptamine (1 μm), (+)-8-hydroxydipropylaminotetralin (10 μm), or 2-methyl-5HT (10 μm), and was blocked by ICS 205-930 with an EC50of ∼2 μmbut not by ketanserin (1 μm). PGE2(4 or 10 μm) also increased Na+currents in varying portions of cells in all four groups.

The effect of 5HT and PGE2on Na+currents was delayed for 20–30 sec after exposure to 5HT, suggesting the involvement of a cytosolic diffusible component in the signaling pathway. The agonist-mediated increase in Na+current, however, was not mimicked by 8-chlorophenylthio-cAMP (200 μm), suggesting the possibility that cAMP was not involved.

The data suggest that the 5HT- and PGE2-mediated increase in Na+current may be involved in hyperesthesia in different but overlapping subpopulations of nociceptors.
Molecular Mechanism of the Inactivation of Tryptophan Hydroxylase by Nitric Oxide: Attack on Critical Sulfhydryls that Spare the Enzyme Iron Center
Authors: Kuhn Donald M. , Arthur Robert
Published: April 2018

Tryptophan hydroxylase (TPH), the initial and rate-limiting enzyme in the biosynthesis of the neurotransmitter serotonin (5-HT), is irreversibly inactivated by nitric oxide (NO). We have expressed brain TPH as a recombinant glutathione-S-transferase fusion protein and delineated the catalytic domain of the enzyme as the region spanning amino acids 99–444. Highly purified TPH catalytic core, like the native enzyme from brain, is inactivated by NO in a concentration-dependent manner. Removal of iron from TPH produces an apoenzyme with low activity that can be reconverted to its highly active holo-form by the addition of ferrous iron. Apo-TPH exposed to NO cannot be reactivated by iron. Treatment of holo-TPH (iron-loaded) with the disulfide 5,5′-dithio-bis (2-nitrobenzoic acid) (DTNB) causes an inactivation of TPH that is readily reversed by dithiothreitol (DTT). DTNB-treated TPH [sulfhydryl (SH)-protected] exposed to NO is returned to full activity by thiol reduction with DTT. The inactivation of native TPH by NO cannot be reversed by either iron or DTT. These data indicate that NO inactivates TPH by selective action on critical SH groups (i.e., cysteine residues) while sparing catalytic iron sites within the enzyme. The results are interpreted with reference to the substituted amphetamines, which are neurotoxic to 5-HT neurons, that inactivate TPHin vivoand are now known to produce NO and other reactive oxygen speciesin vivo.
Redistribution and Stabilization of Cell Surface Glutamate Receptors during Synapse Formation
Authors: Mammen Andrew L. , Huganir Richard L. , O’Brien Richard J.
Published: April 2018

Although the regulation of neurotransmitter receptors during synaptogenesis has been studied extensively at the neuromuscular junction, little is known about the control of excitatory neurotransmitter receptors during synapse formation in central neurons. Using antibodies against extracellular N-terminal (N-GluR1) and intracellular C-terminal (C-GluR1) domains of the AMPA receptor subunit GluR1, combined with surface biotinylation and metabolic labeling studies, we have characterized the redistribution and metabolic stabilization of the AMPA receptor subunit GluR1 during synapse formation in culture. Before synapse formation, GluR1 is distributed widely, both on the surface and within the dendritic cytoplasm of these neurons. The diffuse cell surface pool of receptor appears to be mobile within the membrane and can be induced to cluster by the addition of N-GluR1 to live neurons. As cultures mature and synapses form, there is a redistribution of surface GluR1 into clusters at excitatory synapses where it appears to be immobilized. The change in the distribution of GluR1 is accompanied by an increase in both the half-life of the receptor and the percentage of the total pool of GluR1 that is present on the cell surface. Blockade of postsynaptic AMPA and NMDA receptors had no effect on the redistribution of GluR1. These results begin to characterize the events regulating the distribution of AMPA receptors and demonstrate similarities between synapse formation at the neuromuscular junction and at excitatory synapses in cultured neurons.
TheSwiss CheeseMutant Causes Glial Hyperwrapping and Brain Degeneration inDrosophila
Authors: Kretzschmar Doris , Hasan Gaiti , Sharma Sugandha , Heisenberg Martin , Benzer Seymour
Published: April 2018

Swiss cheese(sws) mutant flies develop normally during larval life but show age-dependent neurodegeneration in the pupa and adult and have reduced life span. In late pupae, glial processes form abnormal, multilayered wrappings around neurons and axons. Degeneration first becomes evident in young flies as apoptosis in single scattered cells in the CNS, but later it becomes severe and widespread. In the adult, the number of glial wrappings increases with age. Theswsgene is expressed in neurons in the brain cortex. The conceptual 1425 amino acid protein shows two domains with homology to the regulatory subunits of protein kinase A and to conceptual proteins of yet unknown function in yeast, worm, and human. Sequencing of twoswsalleles shows amino acid substitutions in these two conserved domains. It is suggested that the novel SWS protein plays a role in a signaling mechanism between neurons and glia that regulates glial wrapping during development of the adult brain.
Neural Coding Mechanisms in Tactile Pattern Recognition: The Relative Contributions of Slowly and Rapidly Adapting Mechanoreceptors to Perceived Roughness
Authors: Blake David T. , Hsiao Steven S. , Johnson Kenneth O.
Published: April 2018

Tactile pattern recognition depends on form and texture perception. A principal dimension of texture perception is roughness, the neural coding of which was the focus of this study. Previous studies have shown that perceived roughness is not based on neural activity in the Pacinian or cutaneous slowly adapting type II (SAII) neural responses or on mean impulse rate or temporal patterning in the cutaneous slowly adapting type I (SAI) or rapidly adapting (RA) discharge evoked by a textured surface. However, those studies found very high correlations between roughness scaling by humans and measures of spatial variation in SAI and RA firing rates. The present study used textured surfaces composed of dots of varying height (280–620 μm) and diameter (0.25–2.5 mm) in psychophysical and neurophysiological experiments. RA responses were affected least by the range of dot diameters and heights that produced the widest variation in perceived roughness, and these responses could not account for the psychophysical data. In contrast, spatial variation in SAI impulse rate was correlated closely with perceived roughness over the whole stimulus range, and a single measure of SAI spatial variation accounts for the psychophysical data in this (0.974 correlation) and two previous studies. Analyses based on the possibility that perceived roughness depends on both afferent types suggest that if the RA response plays a role in roughness perception, it is one of mild inhibition. These data reinforce the hypothesis that SAI afferents are mainly responsible for information about form and texture whereas RA afferents are mainly responsible for information about flutter, slip, and motion across the skin surface.
Vector Averaging for Smooth Pursuit Eye Movements Initiated by Two Moving Targets in Monkeys
Authors: Lisberger Stephen G. , Ferrera Vincent P.
Published: April 2018

The visual input for pursuit eye movements is represented in the cerebral cortex as the distributed activity of neurons that are tuned for both the direction and speed of target motion. To probe how the motor system uses this distributed code to compute a command for smooth eye movements, we have recorded the initiation of pursuit for 150 msec presentations of two spots moving at different speeds and/or in different directions. With equal probability, one of the two spots continued to move at the same speed and in the same direction and became the tracking target, whereas the other disappeared and served as a distractor. We measured eye acceleration in the interval from 110 to 206 msec after the onset of spot motion, within both the open-loop interval for pursuit and the interval during which eye motion was affected by the two spots. Our results demonstrate that weighted vector averaging is used to combine the responses to two moving spots. We found only a minute number of responses that were consistent with either vector summation or winner-take-all computations. In addition, our data show that it is difficult for the monkey to defeat vector averaging without extended training on the use of an explicit cue about which spot will become the target. We argue that our experiment reveals the computations done by the pursuit system in the absence of attentional bias and that vector averaging is normally used to read the distributed code of image motion when there is only one target.