Functional heterogeneity of hippocampal GABAA receptors

gamma-Aminobutyric acid type A (GABAA) receptors were studied in cultured neurons taken from rat hippocampus at early postnatal stages. GABA-induced whole-cell currents showed a broad range of peak amplitudes and time-courses of desensitization. Dose-response curves of rapidly and slowly desensitizing cells revealed EC50 values of 8.5 and 37.3 microM GABA, respectively, with the Hill coefficient being greater than unity. The main-state conductance of GABAA receptor channels was 28-31 pS in all cells. GABA responses of low-affinity cells were more strongly affected by benzodiazepine receptor agonists (e.g. flunitrazepam, clonazepam) and inverse agonists (e.g. methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate), as compared to cells exhibiting high-affinity GABA responses. Currents were also potentiated by zolpidem, but were little affected by Ro 15-4513 and Zn2+. These data suggest the presence of physiologically and pharmacologically distinct GABAA receptor isoforms in neurons of the early postnatal hippocampus, which may subserve different inhibitory control mechanisms in this brain region.     

Cloning of a putative inhibitory amino acid receptor subunit from the parasitic nematode Haemonchus contortus

A cDNA, HGl, encoding an inhibitory amino-acid receptor subunit has been cloned from a mixed egg population of the parasitic nematode Haemonchus contortus. The predicted amino-acid sequence of the subunit shows 24% to 32% homology with other vertebrate and invertebrate GABAA and glycine receptor subunits and has all the expected motifs for a member of the ligand-gated ion channel superfamily. When expressed in Xenopus oocytes HGl gives a small response to 1 mM glycine, but not to 1 mM GABA, glutamate, taurine or L-alanine.     

GABAB receptor-mediated inhibition of GABAA receptor calcium elevations in developing hypothalamic neurons

In the CNS, gamma-aminobutyric acid (GABA) affects neuronal activity through both the ligand-gated GABAA receptor channel and the G protein-coupled GABAB receptor. In the mature nervous system, both receptor subtypes decrease neural excitability, whereas in most neurons during development, the GABAA receptor increases neural excitability and raises cytosolic Ca2+ levels. We used Ca2+ digital imaging to test the hypothesis that GABAA receptor-mediated Ca2+ rises were regulated by GABAB receptor activation. In young, embryonic day 18, hypothalamic neurons cultured for 5 +/- 2 days in vitro, we found that cytosolic Ca2+ rises triggered by synaptically activated GABAA receptors were dramatically depressed (>80%) in a dose-dependent manner by application of the GABAB receptor agonist baclofen (100 nM-100 microM). Coadministration of the GABAB receptor antagonist 2-hydroxy-saclofen or CGP 35348 reduced the inhibitory action of baclofen. Administration of the GABAB antagonist alone elicited a reproducible Ca2+ rise in >25% of all synaptically active neurons, suggesting that synaptic GABA release exerts a tonic inhibitory tone on GABAA receptor-mediated Ca2+ rises via GABAB receptor activation. In the presence of tetrodotoxin the GABAA receptor agonist muscimol elicited robust postsynaptic Ca2+ rises that were depressed by baclofen coadministration. Baclofen-mediated depression of muscimol-evoked Ca2+ rises were observed in both the cell bodies and neurites of hypothalamic neurons taken at embryonic day 15 and cultured for three days, suggesting that GABAB receptors are functionally active at an early stage of neuronal development. Ca2+ rises elicited by electrically induced synaptic release of GABA were largely inhibited (>86%) by baclofen. These results indicate that GABAB receptor activation depresses GABAA receptor-mediated Ca2+ rises by both reducing the synaptic release of GABA and decreasing the postsynaptic Ca2+ responsiveness. Collectively, these data suggest that GABAB receptors play an important inhibitory role regulating Ca2+ rises elicited by GABAA receptor activation. Changes in cytosolic Ca2+ during early neural development would, in turn, profoundly affect a wide array of physiological processes, such as gene expression, neurite outgrowth, transmitter release, and synaptogenesis.     

Transport activity of FhuA, FhuC, FhuD, and FhuB derivatives in a system free of polar effects, and stoichiometry of components involved in ferrichrome uptake

The role of GABA receptors in regulating the mesolimbic dopamine (DA) system and drug reinforced behaviors has not been well characterized. Using fast-cyclic voltammetry, the effects of specific GABA receptor modulation on DA release in the nucleus accumbens (NAcc) and heroin self-administration (SA) behavior was investigated. The GABAA agonist muscimol, administered either intravenously or directly into the ventral tegmental area (VTA), significantly increased DA release in the NAcc in 7 of the 10 rats tested. DA release decreased in the remaining three rats; both effects were blocked by pretreatment with the GABAA receptor antagonist bicuculline. In contrast, the GABAB agonist baclofen decreased, while 2-OH-saclofen (a GABAB antagonist) increased DA release in the NAcc. However, when VTA GABAB receptors were previously activated or inactivated by microinjections of baclofen or 2-OH-saclofen, systemic injections of muscimol caused an inhibition of NAcc DA release. These results suggest that GABAA receptors may be co-localized on both DA neurons and non-DA (GABAergic) interneurons in the VTA, with the effects of GABAA determined by the net effect of both direct inhibition and indirect disinhibition of DA neurons. Finally, although a DA releaser, muscimol was neither self-administered in drug naive rats, nor did it substitute for heroin in rats previously trained to self-administer heroin, suggesting that GABAA receptors appear to play a complex role in mediating drug reinforcement, depending upon the dynamic functional state of GABAA receptors on both tegmental DA and non-DA neurons.     

Molecular composition of GABAC receptors

In the central nervous system inhibitory neurotransmission is primarily achieved through activation of receptors for gamma-aminobutyric acid (GABA). Three types of GABA receptors have been identified on the basis of their pharmacology and electrophysiology. The predominant type, termed GABAA and a recently identified type, GABAC, have integral chloride channels, whereas GABAB receptors couple to separate K+ or Ca2+ channels via G-proteins. By analogy to nicotinic acetylcholine receptors, native GABAA receptors are believed to be heterooligomers of five subunits, drawn from five classes (alpha, beta, gamma, delta, epsilon/chi). An additional class, called rho, is often categorized with GABAA receptor subunits due to a high degree of sequence similarity. However, rho subunits are capable of forming functional homooligomeric and heterooligomeric receptors, whereas GABAA receptors only express efficiently as heterooligomers. Intriguingly, the pharmacological properties of receptors formed from rho subunits are very similar to those exhibited by GABAC receptors and rho subunits and GABAC responses have been colocalized to the same retina cells, indicating that rho subunits are the sole components of GABAC receptors. In contrast, the propensity of GABAA receptor and rho subunits to form multimeric structures and their coexistence in retinal cells suggests that GABAC receptors might be heterooligomers of rho and GABAA receptor subunits. This review will summarize our current understanding of the molecular composition of GABAC receptors based upon studies of rho subunit assembly.     

Unchanged [3H]MK-801 binding and increased [3H]flunitrazepam binding in turtle forebrain during anoxia

In order to determine if functional changes in N-methyl-D-aspartate receptors and GABAA receptors play a role in the remarkable anoxia tolerance of freshwater turtle brain, we used autoradiographic techniques to assay [3H]MK-801 and [3H]flunitrazepam binding in turtle forebrain after turtles had been subjected to anoxia for 2 or 6 h. The effects of glutamate, glycine, competitive N-methyl-D-aspartate antagonists, glycine antagonists, polyamines, magnesium, and zinc on [3H]MK-801 binding were the same in anoxic and control turtle forebrains. These results indicate that NMDA receptor regulation plays no role in the adaptive responses to anoxia in turtle brain. In contrast, [3H]flunitrazepam binding was significantly increased in the anoxic dorsal cortex and striatum. The most parsimonious explanation for elevated benzodiazepine receptor binding is that the rise in extracellular GABA levels known to accompany anoxia enhances benzodiazepine receptor affinity. It is possible, however, that GABAA receptor upregulation during anoxia increases the effectiveness of the inhibitory action of released GABA and contributes to the anoxia tolerance of turtles.     

Presynaptic GABAB autoreceptor modulation of P/Q-type calcium channels and GABA release in rat suprachiasmatic nucleus neurons

GABA is the primary transmitter released by neurons of the suprachiasmatic nucleus (SCN), the circadian clock in the brain. Whereas GABAB receptor agonists exert a significant effect on circadian rhythms, the underlying mechanism by which GABAB receptors act in the SCN has remained a mystery. We found no GABAB receptor-mediated effect on slow potassium conductance, membrane potential, or input resistance in SCN neurons in vitro using whole-cell patch-clamp recording. In contrast, the GABAB receptor agonist baclofen (1-100 microM) exerted a large and dose-dependent inhibition (up to 100%) of evoked IPSCs. Baclofen reduced the frequency of spontaneous IPSCs but showed little effect on the frequency or amplitude of miniature IPSCs in the presence of tetrodotoxin. The activation of GABAB receptors did not modulate postsynaptic GABAA receptor responses. The depression of GABA release by GABAB autoreceptors appeared to be mediated primarily through a modulation of presynaptic calcium channels. The baclofen inhibition of both calcium currents and evoked IPSCs was greatly reduced (up to 100%) by the P/Q-type calcium channel blocker agatoxin IVB, suggesting that P/Q-type calcium channels are the major targets involved in the modulation of GABA release. To a lesser degree, N-type calcium channels were also involved. The inhibition of GABA release by baclofen was abolished by a pretreatment with pertussis toxin (PTX), whereas the inhibition of whole-cell calcium currents by baclofen was only partially depressed by PTX, suggesting that G-protein mechanisms involved in GABAB receptor modulation at the soma and axon terminal may not be identical. We conclude that GABAB receptor activation exerts a strong presynaptic inhibition of GABA release in SCN neurons, primarily by modulating P/Q-type calcium channels at axon terminals.     

Positive modulation of human gamma-aminobutyric acid type A and glycine receptors by the inhalation anesthetic isoflurane

The interactions of the inhalation anesthetic agent isoflurane with ligand-gated chloride channels were studied using transient expression of recombinant human receptors in a mammalian cell line. Isoflurane enhanced gamma-aminobutyric acid (GABA)-activated chloride currents in cells that expressed heteromeric GABAA receptors consisting of combinations of alpha 1 or alpha 2, beta 1, and gamma 2 subunits and in cells that expressed receptors consisting of combinations of only alpha and beta subunits. Receptors consisting of alpha 2 and gamma 2 subunits were poorly expressed but were sensitive to isoflurane. Receptors consisting of beta 1 and gamma 2 subunits were not expressed. Isoflurane also enhanced glycine-activated chloride currents through homomeric alpha glycine receptors but did not enhance GABA currents in cells expressing homomeric rho 1 receptors. These results show that not all ligand-gated chloride channel receptors are sensitive to isoflurane and, therefore, that the anesthetic interacts with specific structural determinants of these ion channel proteins.     

Synaptic inhibition in the isolated respiratory network of neonatal rats

Gramicidin-perforated patch-clamp recording revealed phasic Cl(-)-mediated hyperpolarizations in respiratory neurons of the brainstem-spinal cord preparation from newborn rats. The in vitro respiratory rhythm persisted after block of gamma-aminobutyric acid (GABA), i.e. GABAA, receptor-mediated inhibitory postsynaptic potentials (IPSPs) with bicuculline and/or glycinergic IPSPs with strychnine. In one class of expiratory neurons, bicuculline unmasked inspiration-related excitatory postsynaptic potentials (EPSPs), leading to spike discharge. Bicuculline also blocked hyperpolarizations and respiratory arrest due to bath-applied muscimol, whereas strychnine antagonized similar responses to glycine. The reversal potential of respiration-related IPSPs and responses to GABA, muscimol or glycine was not affected by CO2/HCO3(-)-free solutions, but shifted from about -65 mV to values more positive than -20 mV upon dialysis of the cells with 144 instead of 4 mM Cl-. Impairment of GABA uptake with nipecotic acid or glycine uptake with sarcosine evoked a bicuculline- or strychnine-sensitive decrease of respiratory frequency which could lead to respiratory arrest. Also, the GABAB receptor agonist baclofen led to reversible suppression of respiratory rhythm. This in vitro apnoea was accompanied by a K+ channel-mediated hyperpolarization (reversal potential -88 mV) of tonic cells, whereas membrane potential of neighbouring respiratory neurons remained almost unaffected. Both baclofen-induced hyperpolarization and respiratory depression were antagonised by 2-OH-saclofen, which did not affect respiration-related IPSPs per se. The results show that synaptic inhibition is not essential for rhythmogenesis in the isolated neonatal respiratory network, although (endogenous) GABA and glycine have a strong modulatory action. Hyperpolarizing IPSPs mediated by GABAA and glycine receptors provide a characteristic pattern of membrane potential oscillations in respiratory neurons, whereas GABAB receptors rather appear to be a feature of non-respiratory neurons, possibly providing excitatory drive to the network.     

Inhibitory action of CGS 21680 on cerebral cortical neurons is antagonized by bicuculline and picrotoxin-is GABA involved?

The possibility of an involvement of endogenously released GABA in the inhibitory actions of A1 and A2a adenosine receptor agonists on rat cerebral cortical neurons discharges was examined using the GABAA antagonists bicuculline and picrotoxin. The A1 agonist N6-cyclopentyladenosine (CPA), the A2a agonist CGS 21680 and the non-selective receptor agonist, adenosine, depressed neuronal firing. Applications of bicuculline or picrotoxin enhanced the spontaneous firing rate of cortical neurons, indicating the presence of ongoing GABA-ergic inhibition. Antagonism of GABAA receptors blocked the depressant effects of CGS 21680 on neuronal firing; was without effect on CPA-evoked inhibitions and tended to reduce the duration of adenosine-evoked inhibitions. These results suggest that the depressant effects of A2a receptor activation are due to an increase in GABA-ergic inhibition, likely as a consequence of increased GABA release. GABA does not appear to be involved in adenosine A1 receptor-mediated inhibition of neuronal firing.     

Analysis of LMW RNA profiles of Frankia strains by staircase electrophoresis

The modulation of the Cl- current activated by gamma-aminobutyric acid (GABA) by changes in extracellular pH in freshly isolated rat dorsal root ganglia (DRG) neurons was studied using the whole-cell patch-clamp technique. In the pH range of 5.0-9.0, increased extracellular pH enhanced, and decreased extracellular pH suppressed, current activated by 10 microM GABA in a reversible and concentration-dependent manner with an IC50 of pH 7.1 in these neurons. Acidification to pH 6.5 inhibited currents activated by the GABAA-selective agonist muscimol in all neurons tested. The antagonism of GABA-activated current by lowering the pH was equivalent at holding potentials between -80 and +40 mV and did not involve a significant alteration in reversal potential. Acidification shifted the GABA concentration/response curve to the right, significantly increasing the EC50 for GABA without appreciably changing the slope or maximal value of the curve. Inhibition of the GABA-activated current by protons was not significantly different when the patch-pipette solution was buffered at pH 7.4 or pH 6.5. These results suggest that extracellular protons inhibit GABAA receptor channels in primary sensory neurons by decreasing the apparent affinity of the receptor for GABA. This represents a novel mechanism of inhibition by protons of a neurotransmitter-gated ion channel. Proton inhibition of GABAA receptor channels may account in part for the modulation by protons of sensory information transmission under certain pathophysiological conditions.     

Septal infusions of the hyperpolarization-activated cyclic nucleotide-gated channel (HCN-channel) blocker ZD7288 impair spontaneous alternation but not inhibitory avoidance.

Blockade of septal hyperpolarization-activated cyclic nucleotide-gated channels (HCN) impairs hippocampal theta, an effect that would be expected to impair memory. To test this hypothesis, the present experiments determined whether septal infusions of the selective HCN channel blocker ZD7288 would impair performance on two memory tasks that involve the septo-hippocampal system: spontaneous alternation (SA) and continuous multiple inhibitory avoidance (CMIA). Fifteen minutes prior to assessing SA or CMIA, different groups of male Sprague-Dawley rats were given septal infusions of saline or ZD7288 (0.2, 0.6 or 1.5 μg/0.5 μ1). Septal infusions of ZD7288 impaired SA in a dose-dependent manner; the same infusions did not affect CMIA acquisition or retention. These results appear to be the first demonstration that HCN channels in the medial septum influence memory. Specifically, they suggest that septal HCN channels play a permissive role in spatial working memory, but do not influence emotional long-term memory. Given that these channels are preferentially located on GABA septo-hippocampal projection neurons, the present data provide further evidence that these projection neurons are involved in memory. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Involvement of GABAA receptors in the outgrowth of cultured hippocampal neurons

Whereas GABA is a major inhibitory neurotransmitter in the adult central nervous system, recent experiments performed in our laboratory have shown that the activation of GABAA receptors in the hippocampus leads to excitatory effects during the early post-natal period. The possible consequence of a depolarizing effect of GABA was assessed on the neuritic outgrowth of embryonic hippocampal neurons in culture. No morphological alterations were observed when hippocampal neurons were cultured for three days in the presence of muscimol, a GABAA receptor agonist. In contrast, the neuritic outgrowth of cultured hippocampal neurons was profoundly affected by the presence of bicuculline in the culture medium. In the presence of this GABAA receptor antagonist neurons displayed a reduction in the number of primary neurites and branching points, resulting in a concomitant decrease of the total neuritic length. Thus, this study suggests that GABA, acting on GABAA subtype of receptors, is able to affect the development of the hippocampus.     

Sites of alcohol and volatile anaesthetic action on GABA(A) and glycine receptors

Volatile anaesthetics have historically been considered to act in a nonspecific manner on the central nervous system. More recent studies, however, have revealed that the receptors for inhibitory neurotransmitters such as gamma-aminobutyric acid (GABA) and glycine are sensitive to clinically relevant concentrations of inhaled anaesthetics. The function of GABA(A) and glycine receptors is enhanced by a number of anaesthetics and alcohols, whereas activity of the related GABA rho1 receptor is reduced. We have used this difference in pharmacology to investigate the molecular basis for modulation of these receptors by anaesthetics and alcohols. By using chimaeric receptor constructs, we have identified a region of 45 amino-acid residues that is both necessary and sufficient for the enhancement of receptor function. Within this region, two specific amino-acid residues in transmembrane domains 2 and 3 are critical for allosteric modulation of both GABA(A) and glycine receptors by alcohols and two volatile anaesthetics. These observations support the idea that anaesthetics exert a specific effect on these ion-channel proteins, and allow for the future testing of specific hypotheses of the action of anaesthetics.     

The GABAA receptor is expressed in human neurons derived from a teratocarcinoma cell line

NT2 cells, a human teratocarcinoma cell line, are shown to be differentiated in neuron-like cells (NT2-N cells) by treatment with retinoic acid. The present study identified the neurotransmitter receptors expressed in NT2-N cells using patch-clamp recording. Voltage-sensitive Na+ currents, which are specific for neurons, were observed in NT2-N cells but not in NT2 cells, suggesting that NT2-N cells actually function as neurons. Glutamate receptor agonists, N-methyl-D-aspartate (NMDA) and kainate, evoked whole-cell currents. In addition, gamma-aminobutyric acid (GABA) evoked currents and the currents were inhibited by the selective GABAA receptor antagonist, bicuculline. In outside-out patches, GABA elicited single channel currents with two classes of the slope conductance (26 and 50 pS). No current, however, was induced by ACh, serotonin, or dopamine NT2-N cells, thus, express at least two types of the major excitatory and inhibitory neurotransmitter receptor in the central nervous system, the glutamate and GAGAA receptors, suggesting that these receptors have a crucial role in neurotransmission from the earlier stage of the brain development.     

Ventral tegmental area dopamine neurons mediate the shock sensitization of acoustic startle: A potential site of action for benzodiazepine anxiolytics.

Dopamine (DA)-containing neurons in the ventral tegmental area (VTA) are thought to play an important role in fear motivation. The primary objective of the present study was to determine the connection between DA D?, gamma aminobutyric acid (GABA)A, and benzodiazepine receptors in the VTA and footshock-associated emotionality. Microinfusion of the DA D? receptor agonist quinpirole, the GABAA receptor agonist muscimol, and the benzodiazepine receptor agonist flurazepam into the VTA was observed to suppress the shock enhancement of acoustic startle amplitudes. None of the drugs depressed baseline startle responding or footshock reactivity. The results indicate the involvement of VTA DA neurons in the fear-arousing properties of footshock and implicate the VTA as a possible neural site for the anxiolytic actions of benzodiazepines. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Action of zolpidem on responses to GABA in relation to mRNAs for GABA(A) receptor alpha subunits within single cells: evidence for multiple functional GABA(A) isoreceptors on individual neurons

The relationship between zolpidem sensitivity and GABA(A) receptor alpha subunits was studied in individual dissociated neurons from rat brain. Using whole-cell recording, similar EC50 values were demonstrated for the effect of gamma-aminobutyric acid (GABA) on gated-chloride currents from substantia nigra reticulata (SNR) and lateral septal neurons. Subsequently, many neurons from both the SNR or lateral septum were found to exhibit enhanced GABA-gated chloride currents across concentrations of zolpidem ranging from 10 to 300 nM. Some neurons exhibited a greater than 20% increase in responsiveness to GABA at 30 nM of zolpidem without further increase at higher concentrations of zolpidem. Conversely, zolpidem enhancement of GABA from another group of neurons was not observed at 30 nM zolpidem, but between 100 and 300 nM the response to GABA increased greater than 20%. Finally, a third group of neurons reached both of these criteria for zolpidem enhancement of GABA. This latter spectrum of responses to GABA after varying concentrations of zolpidem was consistent with the presence of either two GABA(A) receptors or a single receptor with differing affinities for zolpidem on an individual neuron. Following determination of the sensitivity of neurons from SNR or lateral septum to zolpidem, cytoplasm was extracted from some individual cells to allow identification of cellular mRNAs for the alpha1, alpha2 and alpha3 GABA(A) receptor subunits with RT-PCR. Those neurons that responded to the 30 nM zolpidem concentration invariably expressed the alpha1-GABA(A) receptor subunit. This result is consistent with the GABA(A) alpha1-receptor subunit being an integral part of a functional high-affinity zolpidem type 1-BZD receptor complex on neurons in brain. Those neurons which showed enhancement of GABA from 100 to 300 nM zolpidem contained mRNAs for the alpha2 and/or the alpha3 receptor subunits, a finding consistent with these alpha subunits forming type 2-BZD receptors. Some individual dissociated SNR neurons were sensitive to both low and high concentrations of zolpidem and contained mRNAs for all three alpha-receptor subunits. These latter individual neurons are proposed to have at least two functional GABA(A) receptor subtypes. Thus, the present investigation emphasizes the importance of characterizing the relationship between endogenous GABA(A) receptor function and the presence of specific structural components forming GABA(A) receptor subtypes on neurons.     

Cellular localizations of AMPA glutamate receptors within the basal forebrain magnocellular complex of rat and monkey

The cellular distributions of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors within the rodent and nonhuman primate basal forebrain magnocellular complex (BFMC) were demonstrated immunocytochemically using anti-peptide antibodies that recognize glutamate receptor (GluR) subunit proteins (i.e., GluR1, GluR4, and a conserved region of GluR2, GluR3, and GluR4c). In both species, many large GluR1-positive neuronal perikarya and aspiny dendrites are present within the medial septal nucleus, the nucleus of the diagonal band of Broca, and the nucleus basalis of Meynert. In this population of neurons in rat and monkey, GluR2/3/4c and GluR4 immunoreactivities are less abundant than GluR1 immunoreactivity. In rat, GluR1 does not colocalize with ChAT, but, within many neurons, GluR1 does colocalize with GABA, glutamic acid decarboxylase (GAD), and parvalbumin immunoreactivities. GluR1- and GABA/GAD-positive neurons intermingle extensively with ChAT-positive neurons. In monkey, however, most GluR1-immunoreactive neurons express ChAT and calbindin-D28 immunoreactivities. The results reveal that noncholinergic GABAergic neurons, within the BFMC of rat, express AMPA receptors, whereas cholinergic neurons in the BFMC of monkey express AMPA receptors. Thus, the cellular localizations of the AMPA subtype of GluR are different within the BFMC of rat and monkey, suggesting that excitatory synaptic regulation of distinct subsets of BFMC neurons may differ among species. We conclude that, in the rodent, BFMC GABAergic neurons receive glutamatergic inputs, whereas cholinergic neurons either do not receive glutamatergic synapses or utilize GluR subtypes other than AMPA receptors. In contrast, in primate, basal forebrain cholinergic neurons are innervated directly by glutamatergic afferents and utilize AMPA receptors.     

Currents evoked by GABA and glycine in acutely dissociated neurons from the rat medial preoptic nucleus

The responses of acutely dissociated medial preoptic neurons to application of GABA, and glycine were studied using the perforated-patch whole-cell recording technique under voltage-clamp conditions. GABA, at a concentration of 1 mM, evoked outward currents in all cells (n = 33) when studied at potentials positive to -80 mV. The I-V relation was roughly linear. The currents evoked by GABA were partially blocked by 25-75 microM picrotoxin and were also partially or completely blocked by 100-200 microM bicuculline. Glycine, at a concentration of 1 mM, did also evoke outward currents in all cells (n = 12) when studied at potentials positive to -75 mV. The I-V relation was roughly linear. The currents evoked by glycine were largely blocked by 1 microM strychnine. In conclusion, the present work demonstrates that neurons from the medial preoptic nucleus of rat directly respond to the inhibitory transmitters GABA and glycine with currents that can be attributed to GABAA receptors and glycine receptors respectively.     

Hippocampal GABA(A) channel conductance increased by diazepam

Benzodiazepines, which are widely used clinically for relief of anxiety and for sedation, are thought to enhance synaptic inhibition in the central nervous system by increasing the open probability of chloride channels activated by the inhibitory neurotransmitter gamma-aminobutyric acid (GABA). Here we show that the benzodiazepine diazepam can also increase the conductance of GABAA channels activated by low concentrations of GABA (0.5 or 5 microM) in rat cultured hippocampal neurons. Before exposure to diazepam, chloride channels activated by GABA had conductances of 8 to 53pS. Diazepam caused a concentration-dependent and reversible increase in the conductance of these channels towards a maximum conductance of 70-80 pS and the effect was as great as 7-fold in channels of lowest initial conductance. Increasing the conductance of GABAA channels tonically activated by low ambient concentrations of GABA in the extracellular environment may be an important way in which these drugs depress excitation in the central nervous system. That any drug has such a large effect on single channel conductance has not been reported previously and has implications for models of channel structure and conductance.