bluuebunny/crossref_metadata_2025 huggingface dataset:
High-Affinity Zinc Inhibition of NMDA NR1–NR2A Receptors
Authors: Paoletti Pierre , Ascher Philippe , Neyton Jacques
Published: April 2018

Micromolar concentrations of extracellular Zn2+are known to antagonize native NMDA receptors via a dual mechanism involving both a voltage-independent and a voltage-dependent inhibition. We have tried to evaluate the relative importance of these two effects and their subunit specificity on recombinant NMDA receptors expressed in HEK 293 cells andXenopusoocytes. The comparison of NR1a–NR2A and NR1a–NR2B receptors shows that the voltage-dependent inhibition is similar in both types of receptors but that the voltage-independent inhibition occurs at much lower Zn2+concentrations in NR1a–NR2A receptors (IC50in the nanomolar range) than in NR1a–NR2B receptors (IC50in the micromolar range). The high affinity of the effect observed with NR1a–NR2A receptors was found to be attributable mostly to the slow dissociation of Zn2+from its binding site. By analyzing the effects of Zn2+on varied combinations of NR1 (NR1a or NR1b) and NR2 (NR2A, NR2B, NR2C), we show that both the NR1 and the NR2 subunits contribute to the voltage-independent Zn2+inhibition. We have observed further that under control conditions, i.e., in zero nominal Zn2+solutions, the addition of low concentrations of heavy metal chelators markedly potentiates the responses of NR1a–NR2A receptors, but not of NR1a–NR2B receptors. This result suggests that traces of a heavy metal (probably Zn2+) contaminate standard solutions and tonically inhibit NR1a–NR2A receptors. Chelation of a contaminant metal also could account for the rapid NR2A subunit-specific potentiations produced by reducing compounds like DTT or glutathione.
Requirement for Tyrosine Phosphatase during Serotonergic Neuromodulation by Protein Kinase C
Authors: Catarsi Stefano , Drapeau Pierre
Published: April 2018

Tyrosine kinases and phosphatases are abundant in the nervous system, where they signal cellular differentiation, mediate the responses to growth factors, and direct neurite outgrowth during development. Tyrosine phosphorylation can also alter ion channel activity, but its physiological significance remains unclear. In an identified leech mechanosensory neuron, the ubiquitous neuromodulator serotonin increases the activity of a cation channel by activating protein kinase C (PKC), resulting in membrane depolarization and modulation of the receptive field properties. We observed that the effects on isolated neurons and channels were blocked by inhibiting tyrosine phosphatases. Serotonergic stimulation of PKC thus activates a tyrosine phosphatase activity associated with the channels, which reverses their constitutive inhibition by tyrosine phosphorylation, representing a novel form of neuromodulation.
Direct Recording of Nicotinic Responses in Presynaptic Nerve Terminals
Authors: Coggan Jay S. , Paysan Jacques , Conroy William G. , Berg Darwin K.
Published: April 2018

Nicotinic acetylcholine receptors are widely expressed in the nervous system, but their functions remain poorly understood. One attractive hypothesis is that the receptors act presynaptically to modulate synaptic transmission. We provide a direct demonstration of presynaptic nicotinic receptorsin situby using whole-cell patch-clamp techniques to record currents in large presynaptic calyces that midbrain neurons form on ciliary neurons. Bath application of nicotine induced inward currents in the calyces capable of generating action potentials that overrode the limited space clamp achievable. The inward currents reversed near 0 mV and showed inward rectification common for neuronal nicotinic receptors. Tetrodotoxin (TTX) blocked the action potentials but not the inward currents. α-Bungarotoxin blocked both, consistent with the presynaptic receptors containing α7 subunits. Recording from the postsynaptic ciliary neurons during nicotine exposure revealed EPSCs that TTX blocked, presumably by blocking presynaptic action potentials. The postsynaptic cells also displayed bimodal inward currents caused by their own nicotinic receptors; the bimodal currents were not blocked by TTX but were blocked partially by α-bungarotoxin and completely byd-tubocurarine. Dye-filling with Lucifer yellow from the recording pipette confirmed the identity of patched structures and showed no dye transfer between calyx and ciliary neuron. When calyces or ciliary neurons were labeled en mass with neurobiotin and biocytin through nerve roots, dye transfer was rarely observed. Thus, electrical synapses were infrequent and unlikely to influence calyx responses. Immunochemical analysis of preganglionic nerve extracts identified receptors that bind α-bungarotoxin and contain α7 subunits. The results unambiguously document the existence of functional presynaptic nicotinic receptors.
Microtubule Transport from the Cell Body into the Axons of Growing Neurons
Authors: Slaughter Theresa , Wang Jun , Black Mark M.
Published: April 2018

The present studies test the hypothesis that microtubules (MTs) are transported from the cell body into the axons of growing neurons. Dissociated sympathetic neurons were cultured using conditions that allow us to control the initiation of axon outgrowth. Neurons were injected with biotin-labeled tubulin (Bt-tub) and then stimulated to extend axons. The newly formed axons were then examined using immunofluorescence procedures for MTs with or without Bt-tub. Because the Bt-tub is fully assembly competent, all MTs that assemble after injection will contain Bt-tub. However, MTs that exist in the neuron at the time of injection and persist during the subsequent incubation will not contain Bt-tub. Because the neurons were injected before extending axons, MTs without Bt-tub are initially localized to the cell body. We specifically determined whether these MTs appeared in the newly formed axon. Such a result can only be explained by the transport of these MTs from their initial location in the cell body into the axon. The newly formed axons of many neurons contained MTs both with and without Bt-tub. MTs without Bt-tub were detected all along the axon and in some neurons represented a substantial portion of the total polymer in the proximal and middle regions of the axon. These results show that MTs are transported from the cell body into growing axons and that this transport is robust, delivering MTs to all regions of the newly formed axon.
Epidermal Growth Factor and Fibroblast Growth Factor-2 Have Different Effects on Neural Progenitors in the Adult Rat Brain
Authors: Kuhn H. Georg , Winkler Jürgen , Kempermann Gerd , Thal Leon J. , Gage Fred H.
Published: April 2018

Neurons and glia are generated throughout adulthood from proliferating cells in two regions of the rat brain, the subventricular zone (SVZ) and the hippocampus. This study shows that exogenous basic fibroblast growth factor (FGF-2) and epidermal growth factor (EGF) have differential and site-specific effects on progenitor cellsin vivo. Both growth factors expanded the SVZ progenitor population after 2 weeks of intracerebroventricular administration, but only FGF-2 induced an increase in the number of newborn cells, most prominently neurons, in the olfactory bulb, the normal destination for neuronal progenitors migrating from the SVZ. EGF, on the other hand, reduced the total number of newborn neurons reaching the olfactory bulb and substantially enhanced the generation of astrocytes in the olfactory bulb. Moreover, EGF increased the number of newborn cells in the striatum either by migration of SVZ cells or by stimulation of local progenitor cells. No evidence of neuronal differentiation of newborn striatal cells was found by three-dimensional confocal analysis, although many of these newborn cells were associated closely with striatal neurons. The proliferation of hippocampal progenitors was not affected by either growth factor. However, EGF increased the number of newborn glia and reduced the number of newborn neurons, similar to the effects seen in the olfactory bulb. These findings may be useful for elucidating thein vivorole of growth factors in neurogenesis in the adult CNS and may aid development of neuronal replacement strategies after brain damage.
OCAM: A New Member of the Neural Cell Adhesion Molecule Family Related to Zone-to-Zone Projection of Olfactory and Vomeronasal Axons
Authors: Yoshihara Yoshihiro , Kawasaki Miwa , Tamada Atsushi , Fujita Hiroko , Hayashi Hideyuki , Kagamiyama Hiroyuki , Mori Kensaku
Published: April 2018

Zone-to-zone projection of olfactory and vomeronasal sensory axons underlies the topographic and functional mapping of chemoreceptor expression zones of the sensory epithelia onto zonally arranged glomeruli in the main and accessory olfactory bulbs. Here we identified OCAM (R4B12 antigen), an axonal surface glycoprotein expressed by subsets of both olfactory and vomeronasal axons in a zone-specific manner. OCAM is a novel homophilic adhesion molecule belonging to the immunoglobulin superfamily with striking structural homology to neural cell adhesion molecule. In both the main and accessory olfactory systems, OCAM mRNA is expressed by sensory neurons in restricted chemoreceptor expression zones, and OCAM protein-expressing axons project to the glomeruli in the corresponding zones of the main and accessory bulbs. OCAM protein is expressed on subsets of growing sensory axons in explant cultures even in the absence of the target bulb. These results demonstrate a precisely coordinated zonal expression of chemoreceptors and OCAM and suggest that OCAM may play important roles in selective fasciculation and zone-to-zone projection of the primary olfactory axons.
Caenorhabditis elegansLevamisole Resistance Geneslev-1,unc-29, andunc-38Encode Functional Nicotinic Acetylcholine Receptor Subunits
Authors: Fleming John T. , Squire Michael D. , Barnes Thomas M. , Tornoe Camilla , Matsuda Kazuhiko , Ahnn Joohong , Fire Andrew , Sulston John E. , Barnard Eric A. , Sattelle David B. , Lewis James A.
Published: April 2018

We show that three of the eleven genes of the nematodeCaenorhabditis elegansthat mediate resistance to the nematocide levamisole and to other cholinergic agonists encode nicotinic acetylcholine receptor (nAChR) subunits.unc-38encodes an α subunit whilelev-1andunc-29encode non-α subunits. The nematode nAChR subunits show conservation of many mammalian nAChR sequence features, implying an ancient evolutionary origin of nAChR proteins. Expression inXenopusoocytes of combinations of these subunits that include theunc-38α subunit results in levamisole-induced currents that are suppressed by the nAChR antagonists mecamylamine, neosurugatoxin, andd-tubocurarine but not α-bungarotoxin. The mutant phenotypes reveal thatunc-38andunc-29subunits are necessary for nAChR function, whereas thelev-1subunit is not. An UNC-29–GFP fusion shows that UNC-29 is expressed in body and head muscles. Two dominant mutations oflev-1result in a single amino acid substitution or addition in or near transmembrane domain 2, a region important to ion channel conductance and desensitization. The identification of viable nAChR mutants inC. elegansprovides an advantageous system in which receptor expression and synaptic targeting can be manipulated and studiedin vivo.
Mammalian Homolog ofDrosophila retinal degeneration BRescues the Mutant Fly Phenotype
Authors: Chang Jinghua T. , Milligan Scott , Li Yuanyuan , Chew Christina E. , Wiggs Janey , Copeland Neal G. , Jenkins Nancy A. , Campochiaro Peter A. , Hyde David R. , Zack Donald J.
Published: April 2018

Mutations in theDrosophila rdgBgene, which encodes a transmembrane phosphatidylinositol transfer protein (PITP), cause a light-enhanced retinal degeneration. Cloning of mammalianrdgBorthologs (mrdgB) reveal predicted proteins that are 39% identical tordgB, with highest homology in the N-terminal PITP domain (62%) and in a region near the C terminus (65%). The humanmrdgBgene spans ∼12 kb and maps to 11q13.1, a locus where several retinal diseases have also been mapped. MurinemrdgBmaps to a syntenic region on the proximal region of chromosome 19.MrdgBis specifically expressed in the retina and brain. In the retina, MrdgB protein is localized to photoreceptor inner segments and the outer and inner plexiform layers. Expression of murinemrdgBin mutant flies fully rescues both therdgB-dependent retinal degeneration and abnormal electroretinogram. These results suggest the existence of similarities between the invertebrate and mammalian retina that were not previously appreciated and also identifymrdgBas a candidate gene for retinal diseases that map to 11q13.1.
Path Integration and Cognitive Mapping in a Continuous Attractor Neural Network Model
Authors: Samsonovich Alexei , McNaughton Bruce L.
Published: April 2018

A minimal synaptic architecture is proposed for how the brain might perform path integration by computing the next internal representation of self-location from the current representation and from the perceived velocity of motion. In the model, a place-cell assembly called a “chart” contains a two-dimensional attractor set called an “attractor map” that can be used to represent coordinates in any arbitrary environment, once associative binding has occurred between chart locations and sensory inputs. In hippocampus, there are different spatial relations among place fields in different environments and behavioral contexts. Thus, the same units may participate in many charts, and it is shown that the number of uncorrelated charts that can be encoded in the same recurrent network is potentially quite large. According to this theory, the firing of a given place cell is primarily a cooperative effect of the activity of its neighbors on the currently active chart. Therefore, it is not particularly useful to think of place cells as encoding any particular external object or event. Because of its recurrent connections, hippocampal field CA3 is proposed as a possible location for this “multichart” architecture; however, other implementations in anatomy would not invalidate the main concepts. The model is implemented numerically both as a network of integrate-and-fire units and as a “macroscopic” (with respect to the space of states) description of the system, based on a continuous approximation defined by a system of stochastic differential equations. It provides an explanation for a number of hitherto perplexing observations on hippocampal place fields, including doubling, vanishing, reshaping in distorted environments, acquiring directionality in a two-goal shuttling task, rapid formation in a novel environment, and slow rotation after disorientation. The model makes several new predictions about the expected properties of hippocampal place cells and other cells of the proposed network.
Noxious Cutaneous Thermal Stimuli Induce a Graded Release of Endogenous Substance P in the Spinal Cord: Imaging Peptide ActionIn Vivo
Authors: Allen Brian J. , Rogers Scott D. , Ghilardi Joseph R. , Menning Patrick M. , Kuskowski Michael A. , Basbaum Allan I. , Simone Donald A. , Mantyh Patrick W.
Published: April 2018

Dorsal root ganglia (DRG) neurons synthesize and transport substance P (SP) to the spinal cord where it is released in response to intense noxious somatosensory stimuli. We have shown previously that SP releasein vivocauses a rapid and reversible internalization of SP receptors (SPRs) in dorsal horn neurons, which may provide a pharmacologically specific image of neurons activated by SP. Here, we report that noxious heat (43°, 48°, and 55°C) and cold (10°, 0°, −10°, and −20°C) stimuli, but not innocuous warm (38°C) and cold (20°C) stimuli, applied to the hindpaw of anesthetized rats induce SPR internalization in spinal cord neurons that is graded with respect to the intensity of the thermal stimulus. Thus, with increasing stimulus intensities, both the total number of SPR+ lamina I neurons showing SPR internalization and the number of internalized SPR+ endosomes within each SPR immunoreactive neuron showed a significant increase. These data suggest that thermal stimuli induce a graded release of SP from primary afferent terminals and that agonist dependent receptor endocytosis provides evidence of a spatially and pharmacologically unique “neurochemical signature” after specific somatosensory stimuli.
Prolonged and Extrasynaptic Excitatory Action of Dopamine Mediated by D1 Receptors in the Rat StriatumIn Vivo
Authors: Gonon François
Published: April 2018

The spatiotemporal characteristics of the dopaminergic transmission mediated by D1 receptors were investigatedin vivo. For this purpose dopamine (DA) release was evoked in the striatum of anesthetized rats by train electrical stimulations of the medial forebrain bundle (one to four pulses at 15 Hz), which mimicked the spontaneous activity of dopaminergic neurons. The resulting dopamine overflow was electrochemically monitored in real time in the extracellular space. This evoked DA release induced a delayed increase in discharge activity in a subpopulation of single striatal neurons. This excitation was attributable to stimulation of D1 receptors by released DA because it was abolished by acute 6-hydroxydopamine lesion and strongly reduced by the D1 antagonist SCH 23390. Striatal neurons exhibiting this delayed response were also strongly excited by intravenous administration of the D1 agonist SKF 82958. Whereas the DA overflow was closely time-correlated with stimulation, the excitatory response mediated by DA started 200 msec after release and lasted for up to 1 sec. Moreover, functional evidence presented here combined with previous morphological data show that D1 receptors are stimulated by DA diffusing up to 12 μm away from release sites in the extrasynaptic extracellular space. In conclusion, DA released by bursts of action potentials exerts, via D1 receptors, a delayed and prolonged excitatory influence on target neurons. This phasic transmission occurs outside synaptic clefts but still exhibits a high degree of spatial specificity.
Seasonal Changes in Testosterone, Neural Attributes of Song Control Nuclei, and Song Structure in Wild Songbirds
Authors: Smith G. Troy , Brenowitz Eliot A. , Beecher Michael D. , Wingfield John C.
Published: April 2018

Seasonal changes in the neural attributes of brain nuclei that control song in songbirds are among the most pronounced examples of naturally occurring plasticity in the adult brain of any vertebrate. The behavioral correlates of this seasonal neural plasticity have not been well characterized, particularly in songbird species that lack adult song learning. To address this question, we investigated the relationship between seasonal changes in gonadal steroids, song nuclei, and song behavior in adult male song sparrows (Melospiza melodia). At four times of the year, we measured plasma concentrations of testosterone, neural attributes of song nuclei, and several aspects of song structure in wild song sparrows of a nonmigratory population. We found seasonal changes in the song nuclei that were temporally correlated with changes in testosterone concentrations and with changes in song stereotypy. Male song sparrows sang songs that were more variable in structure in the fall, when testosterone concentrations were low and song nuclei were small, than in the spring, when testosterone concentrations were higher and song nuclei were larger. Despite seasonal changes in the song nuclei, the song sparrows continued to sing the same number of different song types, indicating that changes in the song nuclei were not correlated with changes in song repertoire size. These results suggest that song stereotypy, but not repertoire size, is a potential behavioral correlate of seasonal plasticity in the avian song control system.
Upregulation of L-Type Ca2+Channels in Reactive Astrocytes after Brain Injury, Hypomyelination, and Ischemia
Authors: Westenbroek Ruth E. , Bausch Suzanne B. , Lin Richard C. S. , Franck Joanne E. , Noebels Jeffery L. , Catterall William A.
Published: April 2018

Anti-peptide antibodies that specifically recognize the α1subunit of class A–D voltage-gated Ca2+channels and a monoclonal antibody (MANC-1) to the α2subunit of L-type Ca2+channels were used to investigate the distribution of these Ca2+channel subtypes in neurons and glia in models of brain injury, including kainic acid-induced epilepsy in the hippocampus, mechanical and thermal lesions in the forebrain, hypomyelination in white matter, and ischemia. Immunostaining of the α2subunit of L-type Ca2+channels by the MANC-1 antibody was increased in reactive astrocytes in each of these forms of brain injury. The α1Csubunits of class C L-type Ca2+channels were upregulated in reactive astrocytes located in the affected regions in each of these models of brain injury, although staining for the α1subunits of class D L-type, class A P/Q-type, and class B N-type Ca2+channels did not change from patterns normally observed in control animals. In all of these models of brain injury, there was no apparent redistribution or upregulation of the voltage-gated Ca2+channels in neurons. The upregulation of L-type Ca2+channels in reactive astrocytes may contribute to the maintenance of ionic homeostasis in injured brain regions, enhance the release of neurotrophic agents to promote neuronal survival and differentiation, and/or enhance signaling in astrocytic networks in response to injury.
A Domain Contributing to the Ion Channel of ATP-Gated P2X2Receptors Identified by the Substituted Cysteine Accessibility Method
Authors: Egan Terrance M. , Haines William R. , Voigt Mark M.
Published: April 2018

P2X receptors are a family of ATP-gated ion channels thought to have intracellular N and C termini and two transmembrane segments separating a large extracellular domain. We examined the involvement of the second putative transmembrane domain (TM2) of the P2X2subunit in ion conduction, using the substituted cysteine accessibility method (SCAM). This method tests the ability of hydrophilic reagents such as Ag+or the methanethiosulfonates to modify covalently the sulfhydryl side chains exposed to aqueous environments. ATP-gated current was measured in HEK293 cells transiently expressing either wild-type or functional mutant P2X2receptors containing a cysteine substitution in or around TM2. Application of Ag+to gating channels had no sustained effect on wild-type P2X2(WT) but irreversibly altered whole-cell currents in 15 mutants. By contrast, bath application of (2-aminoethyl)methanethiosulfonate (MTSEA) to closed channels inhibited 8 of the 15 residues affected by Ag+when the channel was gating. Inhibition of the closed channel was prevented in seven of eight mutants when membrane-permeant MTSEA was scavenged by 20 mmintracellular cysteine, indicating that these seven mutants lie on the intracellular side of the channel gate. Further, MTSEA inhibited current through G342C in the absence of intracellular cysteine but augmented the current when cysteine was present, suggesting that this residue may be part of the gate. Taken together, the data help to the identify a functional domain of the channel pore by mapping residues on either side of the channel gate.
Expression of a Cleaved Brain-Specific Extracellular Matrix Protein Mediates Glioma Cell InvasionIn Vivo
Authors: Zhang Hong , Kelly Gail , Zerillo Cynthia , Jaworski Diane M. , Hockfield Susan
Published: April 2018

Malignant gliomas (primary brain tumors) aggressively invade the surrounding normal brain. This invasive ability is not demonstrated by brain metastases of nonglial cancers. The brain-specific, brain-enriched hyaluronan binding (BEHAB)/brevican gene, which encodes an extracellular hyaluronan-binding protein, is consistently expressed by human glioma and is not expressed by tumors of nonglial origin (Jaworski et al., 1996). BEHAB/brevican can be cleaved into an N-terminal fragment that contains a hyaluronan-binding domain (HABD) and a C-terminal fragment (Yamada et al., 1995). Here, using antisera to peptides in the predicted N-terminal and C-terminal proteolytic fragments, we demonstrate that the BEHAB/brevican protein is cleaved in invasive human and rodent gliomas. A role for this protein in glioma cell invasion was tested by transfecting a noninvasive cell line with the BEHAB/brevican gene. The noninvasive 9L glioma cell was transfected with either full-length BEHAB/brevican or the HABD and tested for invasion inin vitroandin vivoinvasion assays. Although both constructs increased invasionin vitro, only the HABD increased invasion by tumors growingin vivo. Experimental intracranial tumors from full-length transfectants showed no increase in invasion over control tumors, whereas tumors from HABD transfectants showed a marked potentiation of tumor invasion, producing new tumor foci at sites distant from the main tumor mass. This work demonstrates a role for a brain-specific extracellular matrix protein in glioma invasion, opening new therapeutic avenues for a uniformly fatal disease.
Dentate Gyrus Basket Cell GABAAReceptors Are Blocked by Zn2+via Changes of Their Desensitization Kinetics: AnIn SituPatch-Clamp and Single-Cell PCR Study
Authors: Berger Thomas , Schwarz Cordelia , Kraushaar Udo , Monyer Hannah
Published: April 2018

Although GABA type A receptors (GABAARs) in principal cells have been studied in detail, there is only limited information about GABAARs in interneurons. We have used the patch-clamp technique in acute rat hippocampal slices in combination with single-cell PCR to determine kinetic, pharmacological, and structural properties of dentate gyrus basket cell GABAARs. Application of 1 mmGABA (100 msec) to nucleated patches via a piezo-driven fast application device resulted in a current with a fast rise and a marked biexponential decay (time constants 2.4 and 61.8 msec). This decay could be attributed to strong receptor desensitization. Dose–response curves for the peak and the slow component yielded EC50values of 139 and 24 μm, respectively. Zn2+caused a marked blocking effect on both the peak and the slow component via a noncompetitive mechanism (IC50values of 8 and 16 μm). This led to an acceleration of the slow component as well as a prolongation of recovery from desensitization. Zn2+sensitivity was suggested to depend on the absence of γ-subunits in GABAARs. To test this hypothesis we performed single-cell reverse transcription PCR that revealed primarily the presence of α2-, β2-, β3-, γ1-, and γ2-subunit mRNAs. In addition, flunitrazepam increased the receptor affinity for its agonist, indicating the presence of functional benzodiazepine binding sites, i.e., γ-subunits. Thus, additional factors seem to co-determine the Zn2+sensitivity of native GABAARs. The modulatory effects of Zn2+on GABAAR desensitization suggest direct influences on synaptic integration via changes in inhibition and shunting at GABAergic synapses.
Subunit Composition and Quantitative Importance of Hetero-oligomeric Receptors: GABAAReceptors Containing α6Subunits
Authors: Jechlinger Martin , Pelz Robert , Tretter Verena , Klausberger Thomas , Sieghart Werner
Published: April 2018

In cerebellum, GABAAreceptors containing α6subunits are expressed exclusively in granule cells. The number of α6receptor subtypes formed in these cells and their subunit composition presently are not known. Immunoaffinity chromatography on α6subunit-specific antibodies indicated that 45% of GABAAreceptors in cerebellar extracts contained α6subunits. Western blot analysis demonstrated that α1, β1, β2, β3, γ2, and δ subunits co-purified with α6subunits, suggesting the existence of multiple α6receptor subtypes. These subtypes were identified using a new method based on the one-by-one immunochromatographic elimination of receptors containing the co-purifying subunits in parallel or subsequent experiments. By quantification and Western blot analysis of α6receptors remaining in the extract, the proportion of α6receptors containing the eliminated subunit could be calculated and the subunit composition of the remaining receptors could be determined. Results obtained indicated that α6receptors in cerebellum are composed predominantly of α6βxγ2(32%), α1α6βxγ2(37%), α6βxδ (14%), or α1α6βxδ (15%) subunits. Other experiments indicated that 10%, 51%, or 21% of α6receptors contained homogeneous β1, β2, or β3subunits, respectively, whereas two different β subunits were present in 18% of all α6receptors. The method presented can be used to resolve the total number, subunit composition, and abundancy of GABAAreceptor subtypes in the brain and can also be applied to the investigation of other hetero-oligomeric receptors.
Aquaporin-4 Water Channel Protein in the Rat Retina and Optic Nerve: Polarized Expression in Müller Cells and Fibrous Astrocytes
Authors: Nagelhus Erlend A. , Veruki Margaret L. , Torp Reidun , Haug Finn-M. , Laake Jon H. , Nielsen Søren , Agre Peter , Ottersen Ole P.
Published: April 2018

The water permeability of cell membranes differs by orders of magnitude, and most of this variability reflects the differential expression of aquaporin water channels. We have recently found that the CNS contains a member of the aquaporin family, aquaporin-4 (AQP4). As a prerequisite for understanding the cellular handling of water during neuronal activity, we have investigated the cellular and subcellular expression of AQP4 in the retina and optic nerve where activity-dependent ion fluxes have been studied in detail.In situhybridization with digoxigenin-labeled riboprobes and immunogold labeling by a sensitive postembedding procedure demonstrated that AQP4 and AQP4 mRNA were restricted to glial cells, including Müller cells in the retina and fibrous astrocytes in the optic nerve. A quantitative immunogold analysis of the Müller cells showed that these cells exhibited three distinct membrane compartments with regard to AQP4 expression. End feet membranes (facing the vitreous body or blood vessels) were 10–15 times more intensely labeled than non-end feet membranes, whereas microvilli were devoid of AQP4. These data suggest that Müller cells play a prominent role in the water handling in the retina and that they direct osmotically driven water flux to the vitreous body and vessels rather than to the subretinal space. Fibrous astrocytes in the optic nerve similarly displayed a differential compartmentation of AQP4. The highest expression of AQP4 occurred in end feet membranes, whereas the membrane domain facing the nodal axolemma was associated with a lower level of immunoreactivity than the rest of the membrane. This arrangement may allow transcellular water redistribution to occur without inducing inappropriate volume changes in the perinodal extracellular space.
Developmental Expression of the μ, κ, and δ Opioid Receptor mRNAs in Mouse
Authors: Zhu Yanxin , Hsu Ming-Sing , Pintar John E.
Published: April 2018

To characterize further the establishment of the opioid system during prenatal mouse development, we have examined the spatial and temporal expression patterns of μ, κ, and δ opioid receptor mRNAs and find that the expression patterns of these mRNAs are distinct at all ages. Within the embryo, κ is the first opioid receptor expressed, with transcripts detected in the gut epithelium as early as embryonic day 9.5 (E9.5). By E10.5, μ receptor expression is first detected in the facial–vestibulocochlear preganglion complex, whereas δ receptor mRNA is first detected at E12.5 in several peripheral tissues, including the olfactory epithelium, heart, limb bud, and tooth. In the brain, both μ and κ mRNAs are first detected at E11.5 in the basal ganglia and midbrain, respectively. During mid-gestation and late gestation, the expression of both μ and κ receptors extends to other brain regions that exhibit high expression in the adult, including the medial habenula, hypothalamus, pons, and medulla for μ and the basal ganglia, thalamus, hypothalamus, raphe, and ventral tegmental area for κ. Thus by E17.5, many aspects of the adult expression patterns of μ and κ receptors already have been established. Compared with μ and κ, δ receptor mRNA expression in the brain begins relatively late, and the expression levels remain very low even at E19.5. In contrast to its late appearance in the brain, however, δ is the first opioid receptor expressed in the dorsal root ganglion, at E12.5, before its expression in the spinal cord begins at E15.5. μ receptor is the first opioid receptor expressed in the spinal cord, at E11.5. These results extend previous ligand-binding data to significantly earlier ages and suggest that early developmental events in both neural and non-neural tissues may be modulated by opioid receptors. Several examples of possible autocrine and paracrine loops of opioid peptide and receptor expression have been identified, suggesting a role for these local circuits in developmental processes.
Neuronal Signals in the Monkey Ventral Striatum Related to Progress through a Predictable Series of Trials
Authors: Shidara Munetaka , Aigner Thomas G. , Richmond Barry J.
Published: April 2018

Single neurons in the ventral striatum of primates carry signals that are related to reward and motivation. When monkeys performed a task requiring one to three bar release trials to be completed successfully before a reward was given, they seemed more motivated as the rewarded trials approached; they responded more quickly and accurately. When the monkeys were cued as to the progress of the schedule, 89 out of 150 ventral striatal neurons responded in at least one part of the task: (1) at the onset of the visual cue, (2) near the time of bar release, and/or (3) near the time of reward delivery. When the cue signaled progress through the schedule, the neuronal activity was related to the progress through the schedule. For example, one large group of these neurons responded in the first trial of every schedule, another large group responded in trials other than the first of a schedule, and a third large group responded in the first trial of schedules longer than one. Thus, these neurons coded the state of the cue, i.e., the neurons carried the information about how the monkey was progressing through the task. The differential activity disappeared on the first trial after randomizing the relation of the cue to the schedule. Considering the anatomical loop structure that includes ventral striatum and prefrontal cortex, we suggest that the ventral striatum might be part of a circuit that supports keeping track of progress through learned behavioral sequences that, when successfully completed, lead to reward.
Differential Modulation of High-Frequency γ-Electroencephalogram Activity and Sleep–Wake State by Noradrenaline and Serotonin Microinjections into the Region of Cholinergic Basalis Neurons
Authors: Cape Edmund G. , Jones Barbara E.
Published: April 2018

Several lines of evidence indicate that cholinergic basalis neurons play an important role in cortical activation. The present study was undertaken to determine the effect of noradrenergic and serotonergic modulation of the cholinergic neurons on cortical EEG activity and sleep–wake states. The neurotransmitters were injected into the region of the basalis neurons by remote control in freely moving, naturally sleeping–waking rats during the day when the rats are normally asleep the majority of the time. Effects were observed on behavior and EEG activity, including high-frequency γ activity (30–60 Hz), which has been demonstrated to reflect behavioral and cortical arousal in the rat. Noradrenaline, which has been shown in previousin vitrostudies to depolarize and excite the cholinergic cells, produced a dose-dependent increase in γ-EEG activity, a decrease in δ activity, and an increase in waking. Serotonin, which has been found in previousin vitrostudies to hyperpolarize the cholinergic neurons, produced a dose-dependent decrease in γ-EEG activity with no significant change in amounts of wake or slow wave sleep. Both chemicals resulted in a dose-dependent decrease in paradoxical sleep.

These results demonstrate that noradrenaline and serotonin exert differential modulatory effects on EEG activity through the basal forebrain, the one facilitating γ activity and eliciting waking and the other diminishing γ activity and not significantly affecting slow wave sleep. The results also confirm that the cholinergic basalis neurons play an important role in cortical activation and particularly in the high-frequency γ activity that underlies cortical and behavioral arousal of the wake state.
Direction Tuning of Individual Retinal Inputs to the Turtle Accessory Optic System
Authors: Kogo Naoki , Rubio Doris McGartland , Ariel Michael
Published: April 2018

Neurons in turtle accessory optic system [basal optic nucleus (BON)] were recorded to study convergence of retinal afferents, using whole-cell patch electrodes in a reducedin vitrobrainstem preparation with the eyes attached. BON cells primarily exhibit EPSPs from a contralateral retinal ganglion cell input and generate an output of action potentials. Visual responses were evoked by different directions of either full-field or local moving patterns. Direction tuning of action potentials was compared with that of EPSPs detected by passing the membrane voltage through an AC amplifier and window discriminator. This rough measure of retinal input indicated that the direction tuning of the full-field excitatory input from the retina matched that of the spike output for the same BON cell.

Using local patterns within the receptive fields of the BON cells, it was estimated that one to four adjacent retinal inputs were being stimulated. The direction tuning of these inputs had preferred directions that were similar to that of the full-field spike output of the cell, irrespective of where the small window was placed within the receptive field. Because more than one retinal input may have been stimulated by the small stimulus window, subsets of those EPSPs that may represent responses of a single retinal afferent were identified based on their amplitude and rise time. Again, the preferred direction of those putative single retinal afferents matched the direction tuning of the spike output of the BON cell. These findings are discussed in terms of the formation of the retinal slip signal by the BON.
Dopamine Axon Varicosities in the Prelimbic Division of the Rat Prefrontal Cortex Exhibit Sparse Immunoreactivity for the Dopamine Transporter
Authors: Sesack Susan R. , Hawrylak Valerie A. , Matus Claudia , Guido Margaret A. , Levey Allan I.
Published: April 2018

The dopamine transporter (DAT) critically regulates the duration of the cellular actions of dopamine and the extent to which dopamine diffuses in the extracellular space. We sought to determine whether the reportedly greater diffusion of dopamine in the rat prefrontal cortex (PFC) as compared with the striatum is associated with a more restricted axonal distribution of the cortical DAT protein. By light microscopy, avidin–biotin–peroxidase immunostaining for DAT was visualized in fibers that were densely distributed within the dorsolateral striatum and the superficial layers of the dorsal anterior cingulate cortex. In contrast, DAT-labeled axons were distributed only sparsely to the deep layers of the prelimbic cortex. By electron microscopy, DAT-immunoreactive profiles in the striatum and cingulate cortex included both varicose and intervaricose segments of axons. However, DAT-labeled processes in the prelimbic cortex were almost exclusively intervaricose axon segments. Immunolabeling for tyrosine hydroxylase in adjacent sections of the prelimbic cortex was localized to both varicosities and intervaricose segments of axons. These qualitative observations were supported by a quantitative assessment in which the diameter of immunoreactive profiles was used as a relative measure of whether varicose or intervaricose axon segments were labeled. These results suggest that considerable extracellular diffusion of dopamine in the prelimbic PFC may result, at least in part, from a paucity of DAT content in mesocortical dopamine axons, as well as a distribution of the DAT protein at a distance from synaptic release sites. The results further suggest that different populations of dopamine neurons selectively target the DAT to different subcellular locations.
D1- Versus D2-Receptor Modulation of Visuospatial Working Memory in Humans
Authors: Müller Ulrich , von Cramon D. Yves , Pollmann Stefan
Published: April 2018

The effects of pergolide, a mixed D1/D2 receptor agonist, and bromocriptine, a selective D2 receptor agonist, were assessed in a visual delay task to further investigate the “dopamine link” of working memory in humans and to look for differential D1 versus D2 receptor contributions. Two groups of 32 healthy young adults (16 female) received either 0.1 mg of pergolide or 2.5 mg of bromocriptine in a placebo-controlled cross-over design. A pretreatment with domperidone, a peripherally active D2 antagonist, was performed in both groups to reduce side effects. Interindividual differences in pharmacokinetics were controlled by the time course of serum prolactin inhibition. The working memory paradigm was a visuospatial delayed matching task; the location of a randomly generated seven-point pattern had to be memorized and compared after 2, 8, or 16 sec with a second pattern that was either identical or slightly shifted within a reference frame. The task was designed with the intention to present unique stimuli at each trial and to require minimal motor demands. Practice effects between the two pharmacological test days were minimized by training sessions that preceded the tests. The paradigm showed significant error and reaction time increases with longer delays. After comparable doses, only pergolide, but not bromocriptine, facilitated visuospatial working memory performance as demonstrated by a significant drug-by-delay interaction. These findings are in accordance with the monkey literature as well as with neuroanatomical findings, and they confirm a preferential role of prefrontal D1 receptors for working memory modulation in humans.
Importance of the Noradrenaline–Dopamine Coupling in the Locomotor Activating Effects of d-Amphetamine
Authors: Darracq Laurent , Blanc Gérard , Glowinski Jacques , Tassin Jean-Pol
Published: April 2018

The locomotor hyperactivity induced by systemic or local (nucleus accumbens)d-amphetamine injections can be blocked by systemic or local (prefrontal cortex) injections of prazosin, an α1-adrenergic antagonist (Blanc et al., 1994). Microdialysis studies performed on freely moving animals indicated that prazosin (0.5 mg/kg, i.p.) does not modify the increase in the extracellular dopamine (DA) levels in the nucleus accumbens that are induced byd-amphetamine (2.0 mg/kg, i.p.), but it inhibits thed-amphetamine-induced locomotor hyperactivity (−63%,p< 0.0001). No behavioral activation occurred after the bilateral local perfusion of 3 μmd-amphetamine in the nucleus accumbens, although it led to a fivefold increase in extracellular DA levels. This increase in extracellular DA levels was not affected by prazosin (0.5 mg/kg, i.p.). When an intraperitoneal injection ofd-amphetamine (0.5 mg/kg) was superimposed to the continuous local perfusion of 3 μmd-amphetamine, it induced a 64% increase in the extracellular DA levels in the nucleus accumbens, and this response was associated with simultaneous behavioral activation. Both the increases in extracellular DA levels and in locomotor activity were completely blocked by a pretreatment with prazosin, injected either systemically (0.5 mg/kg, i.p.) or locally and bilaterally into the prefrontal cortex (500 pmol/side). Complementary experiments indicated that the focal application ofd-amphetamine requires at least a 4.8-fold higher increase in DA output compared with systemicd-amphetamine for the behavioral effects to be elicited. Altogether, these results suggest that locomotor activating effects ofd-amphetamine are caused by the stimulation of cortical α1-adrenergic receptors by noradrenaline, which increases the release of a functional part of subcortical DA.
Time Course and Permeation of Synaptic AMPA Receptors in Cochlear Nuclear Neurons Correlate with Input
Authors: Gardner Stephanie M. , Trussell Laurence O. , Oertel Donata
Published: April 2018

AMPA receptors mediate rapid glutamatergic synaptic transmission. In the mammalian cochlear nuclei, neurons receive excitatory input from either auditory nerve fibers, parallel fibers, or both fiber systems. The functional correlates of differences in the source of input were examined by recording AMPA receptor-mediated, miniature EPSCs (mEPSCs) in whole-cell voltage-clamp mode from identified neurons. Bushy, octopus, and T-stellate cells of the ventral cochlear nucleus (VCN) and tuberculoventral cells of the dorsal cochlear nucleus (DCN) receive most of their excitatory input from the auditory nerve; fusiform cells receive excitatory inputs from both the auditory nerve and parallel fibers; cartwheel cells receive excitatory input from parallel fibers alone. mEPSCs from bushy, octopus, T-stellate, and tuberculoventral cells had significantly faster decay time constants (0.35–0.40 msec) than did those from fusiform and cartwheel cells (1.32–1.79 msec). Some fusiform cells had two populations of mEPSCs with distinct time courses. mEPSCs in cells with auditory nerve input alone were inhibited by philanthotoxin, a blocker of calcium-permeable AMPA receptors, whereas mEPSCs in cells with parallel fiber input were not. Thus AMPA receptors postsynaptic to the auditory nerve differ from those postsynaptic to parallel fibers both in channel-gating kinetics and in their permeability to calcium. These results confirm the conclusion that synaptic AMPA receptors are specialized according to the source of input (Hunter et al., 1993; Rubio and Wenthold, 1997;Wang et al., 1998).
Developmental Expression of anamn+Transgene Rescues the Mutant Memory Defect ofamnesiacAdults
Authors: DeZazzo James , Xia Shouzhen , Christensen Jeff , Velinzon Klara , Tully Tim
Published: April 2018

TheDrosophilamemory geneamnesiac(amn) has been proposed to encode a neuropeptide protein, which includes regions homologous to vertebrate pituitary adenylyl cyclase-activating peptide (PACAP; Feany and Quinn, 1995). Definitive experiments to link this gene to memory formation, however, have not yet been accomplished (Kandel and Abel, 1995). The experiments described here demonstrate that the putativeamntranscript is involved in adult memory formation. With the use of aUAS–amn+transgene, we show complete rescue of memory defects inamn28A, a mutant allele caused by the insertion of a GAL4 enhancer trap transposon (Moore et al., 1998). Study of theamn28Areporter reveals widespread expression in the adult brain but also enriched expression in the embryonic and larval nervous systems. To begin addressing the temporal requirement ofamnin memory, we asked whether the memory defects could be rescued by restricting transgenic expression to the adult stage. A heat-shock regimen shown previously to rescue fully theamnethanol sensitivity defect (Moore et al., 1998) failed to rescue the memory defect. These results, coupled with previous genetic and anatomical studies, suggest that adult memory formation and ethanol sensitivity have different temporal and spatial requirements foramn.