Nucleus accumbens dopamine release modulation by mesolimbic GABAA receptors—an in vivo electrochemical study

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 GABA_A 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 GABA_A receptor antagonist bicuculline. In contrast, the GABA_B agonist baclofen decreased, while 2-OH-saclofen (a GABA_B antagonist) increased DA release in the NAcc. However, when VTA GABA_B 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 GABA_A receptors may be co-localized on both DA neurons and non-DA (GABAergic) interneurons in the VTA, with the effects of GABA_A 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 GABA_A receptors appear to play a complex role in mediating drug reinforcement, depending upon the dynamic functional state of GABA_A receptors on both tegmental DA and non-DA neurons.     

Clathrin-coated vesicles from bovine brain contain uncoupled GABAA receptors

Clathrin-coated vesicles are thought to be a vehicle for the sequestration of GABA_A receptors. For coated vesicles from bovine cerebrum, we examined the binding properties of [³H]muscimol, a GABA_A-specific agonist, [³H]flunitrazepam, a benzodiazepine agonist, and [³⁵S]t-butylbiocyclophosphorthionate (TBPS), a ligand for GABA_A receptor channels. Under standard conditions, the binding level of [³H]muscimol, [³H]flunitrazepam, and [³⁵S]TBPS to coated vesicles represented 12.3±1.8%, 7.9±1%, and 10.2±1.8%, respectively, of that in crude synaptic membranes. Coated vesicles showed a single [³H]flunitrazepam binding site with a K_D value (12 nM) which was 9-fold that for synaptic membranes. The allosteric coupling between binding sites was measured by the addition of GABA to [³H]flunitrazepam and [³⁵S]TBPS binding assays. For [³H]flunitrazepam binding to synaptic membranes, GABA gave an EC₅₀=2.0 μM and at saturation (100 μM) an enhancement of 122%. This stimulation was completely blocked by the GABA antagonist SR95531. In contrast, neither GABA nor SR95531 had a significant effect on [³H]flunitrazepam binding to CCVs, indicating that the allosteric interaction between GABA and benzodiazepine binding sites is abolished. Likewise, GABA displaced nearly all of the [³⁵S]TBPS binding to synaptic membranes but had no effect on binding to coated vesicles, indicating that coupling between the GABA binding sites and chloride channel is also impaired. Thus GABA_A receptors appear to be uncoupled during normal intracellular trafficking via coated vesicles. The presence of major GABA_A receptor subunits on these particles was verified by quantitative immunoblotting. Relative to the levels in synaptic membranes, CCVs contained 110±14% and 29.5±3.8%, respectively, of the immunoreactivity for GABA_A receptor β2 and α1 subunits. Thus, in comparison to GABA_A receptors on synaptic membranes, those on CCVs have a reduced α1/β2-subunit ratio. It may be suggested that a selective decline in the content of α1 subunits in coated vesicles could in part account for GABA_A receptor uncoupling.     

Several GABAA receptor subunits are expressed in LHRH neurons of juvenile female rats

Gamma aminobutyric acid (GABA), the dominant inhibitory neurotransmitter in brain, is involved in the developmental regulation of LHRH secretion. Morphological studies in rodents have demonstrated that LHRH neurons are innervated by GABA-containing processes, suggesting that LHRH secretion is under direct transsynaptic GABAergic control. While GABA acts through two different receptors, GABA_A and GABA_B, to exert its effects, it appears that GABA_A receptors are able to mediate both inhibitory and stimulatory effects of GABA on LHRH neurons. GABA_A receptors are heterooligomeric ligand-gated anion channels that exhibit a diverse array of functional and pharmacological properties. This diversity is determined by the structural heterogeneity of the receptors, which are assembled from the combination of different classes of subunits with multiple isoforms. Although several studies have described the effect of GABA_A receptor stimulation on LHRH and/or gonadotropin release in prepubertal animals, nothing is known about the receptor subunits that may be expressed in LHRH neurons at this phase in development. Double immunohistofluorescence followed by confocal laser microscopy revealed that subsets of prepubertal LHRH neurons are endowed with α₁, α₂, β_2/3, and γ₂ GABA_A receptor subunits. Combined immunohistochemistry for LHRH neurons and in situ hybridization for GABA_A subunit mRNAs confirmed that the genes encoding the α₁, α₂, β₃ and γ₂ subunits, but not the γ₁ subunit, are expressed in LHRH neurons. Notwithstanding the relative insensitivity of these methods, both the immunohistochemical and hybridization histochemical approaches employed indicate that only a fraction of LHRH neurons are endowed with GABA_A receptors. This arrangement suggests that those LHRH neurons bearing the appropriate GABA_A receptors are responsible for either the entire secretory response to direct GABAergic inputs or for its initiation.     

GABAB receptors in the dorsomedial hypothalamus and heart rate in anesthetized rats

Previous studies have shown that: (1) activation of neurons in the dorsomedial hypothalamus (DMH) of the rat by blockade of local GABA_A receptors with bicuculline methiodide (BMI) elicits cardiovascular changes resembling those seen in experimental stress, including marked sympathetically-mediated tachycardia, and (2) inhibition of neurons in the same region by local microinjection of the GABA_A receptor agonist muscimol can virtually abolish stress-induced tachycardia. This study examined the possibility that GABA_B receptors exist in the neural circuitry of the DMH, and that stimulation of these receptors might suppress the cardiovascular response to local disinhibition with BMI. Microinjection of BMI 10 pmol into the DMH in urethane-anesthetized rats resulted in marked tachycardia with little or no effect on arterial pressure. Simultaneous injection of the GABA_B receptor agonist baclofen at doses of 2.5, 5.0 and 10 pmol produced dose-related suppression of BMI induced tachycardia. Coinjection of the GABA_B receptor antagonist 2-hydroxysaclofen 100 or 200 pmol had no significant effect on the heart rate response to BMI, but reversed the suppression elicited in the presence of baclofen. These findings indicate that (1) functional GABA_B receptors exist in the DMH, and (2) stimulation of these receptors inhibits the tachycardia resulting from blockade of local GABA_A receptors.     

5-HT2 receptor regulation of acetylcholine release induced by dopaminergic stimulation in rat striatal slices

The role of 5-hydroxytryptamine (5-HT) receptor subtypes in acetylcholine (ACh) release induced by dopamine or neurokinin receptor stimulation was studied in rat striatal slices. The dopamine D₁ receptor agonist SKF 38393 potentiated in a tetrodotoxin-sensitive manner the K⁺-evoked [³H]ACh release while SCH 23390, a dopamine D₁ receptor antagonist, had no effect. [³H]ACh release was decreased by the dopamine D₂ receptor agonist LY 171555 (quinpirole) and slightly potentiated by the dopamine D₂ receptor antagonist haloperidol. The selective neurokinin NK₁ receptor agonist [Sar⁹, met(O₂)¹¹]SP also potentiated K⁺-evoked release of [³H]ACh. GR 82334, a NK₁ receptor antagonist, blocked not only the effect of [Sar⁹, met(O₂)¹¹]SP but also the release of ACh induced by the D₁ receptor agonist SKF 38393. Among the 5-HT agents studied, only the 5-HT_2A receptor antagonists ketanserin and ritanserin were able to reduce the ACh release induced by dopamine D₁ receptor stimulation. Mesulergine, a more selective 5-HT_2C antagonist, showed an intrinsic releasing effect but did not affect K⁺-evoked ACh release induced by SKF 38393. Methysergide and methiothepin, mixed 5-HT_1/2 antagonists, as well as ondansetron, a 5-HT₃ receptor antagonist, showed an intrinsic effect on ACh release, their effects being additive to that of SKF 38393. 5-HT₂ receptor agonists were ineffective. However, the 5-HT₂ agonist DOI was able to prevent the antagonism by ketanserin of the increased [³H]ACh efflux elicited by SKF 38393, suggesting a permissive role of 5-HT_2A receptors. None of the above indicated 5-HT agents was able to reduce the ACh release induced by the selective NK₁ agonist. The results suggest that 5-HT₂ receptors, probably of the 5-HT_2A subtype, modulate the release of ACh observed in slices from the rat striatum after stimulation of dopamine D₁ receptors. It seems that this serotonergic control is exerted on the interposed collaterals of substance P-containing neurons which promote ACh efflux through activation of NK₁ receptors located on cholinergic interneurons.     

Lesion with the neurotoxin AF64A alters hippocampal cholinergic receptor function

The effect of selective lesion of cholinergic inputs to the hippocampus on the function of hippocampal cholinergic receptors was examined. Hippocampal cholinergic neurons were lesioned in the rat by administration of the selective cholinergic neurotoxin AF64A (ethylcholine mustard azirtdinium). Cholinergic receptor function was examined by assessing the ability of cholinergic agonists and antagonists to modulate the evoked release of radiolabelled acetylcholine (ACh) from hippocampal slices. Nicotine enhanced release, with a bell-shaped dose-response curve. The dose-response curve and EC₅₀ for nicotine was shifted 10-fold to the left in lesioned rats, suggesting an increased sensitivity to nicotine. However, there were no differences in either the number or affinity of nicotinic receptors as determined with binding studies. The muscarinic agonist oxotremorine inhibited the evoked release of ACh in control tissues, but had much less effect in AF64A-lesioned tissues. Binding to the M1 receptor subtype was not changed. However, the K_d for binding to the high affinity subtype of the M2 receptor was increased 10-fold, suggesting that the receptor has become less sensitive to stimulation. Loss of M2 function may allow an increase in the effect of stimulating nicotinic receptors that modulate ACh release.     

GABAB receptor-mediated inhibition of spontaneous action potential discharge in rat supraoptic neurons in vitro

To elucidate the role of GABA_B receptors in the regulation of the electrical activity of magnocellular neurons of the supraoptic nucleus (SON), the effects of GABA_B agonist and antagonist on the firing rate of spontaneous action potentials were studied in SON slice preparations of rats by extracellular recordings. In the presence of the γ-amino butyric acid (GABA)-gated chloride channel blocker, picrotoxin, the selective GABA_B agonist, baclofen, reduced the firing rate of action potentials in both phasic and non-phasic neurons in a dose-dependent manner. The reduction in the firing rate induced by baclofen was reversed by the selective GABA_B antagonist, 2-hydroxy saclofen (2OH-saclofen), also in a dose-dependent manner. In non-phasic neurons, 2OH-saclofen significantly increased the firing rate and the effect was additive to the effect of picrotoxin. In phasic neurons, 2OH-saclofen alone did not increase the firing rate, but it reversed suppression of the firing induced by increasing extracellular Ca²⁺ concentration to 2.1 mM. Baclofen also reduced the firing rate of non-phasic neurons of virgin and lactating female rats, indicating that the GABA_B receptor-mediated inhibition is not confined to SON neurons of male rats. The evidence indicates that activation of GABA_B receptors inhibits electrical activity of SON neurons of both male and female rats and that GABA_B receptors may play an important role in the inhibitory regulation of the electrical activity of SON neurons by GABA.     

GABAB receptors and norepinephrine-stimulated cAMP production in rat brain cortex

The ability of GABAergic compounds to influence cAMP accumulation in rat brain cortex was examined. It was found that GABA_B receptor agonist such as GABA, baclofen, and kojic amine potentiate the cAMP response in cerebral cortex during exposure to norepinephrine. Isoguvacine and THIP, selective GABA_A receptor agonists, did not demonstrate this effect. The response to baclofen was stereoselective, with virtually all activity residing in the (−)isomer. Bicuculline methiodide had no effect on the agonist-induced potentiation and the rank order of potency for GABA_B agonists to potentiate the cAMP response is identical to their rank order of potency in the GABA_B binding assay. These data suggest that GABA_B receptors are capable of influencing the brain cyclic nucleotide system.     

GABAA and GABAB receptors modulating basal and footshock-induced nitric oxide releases in rat prefrontal cortex

Using an in vivo brain microdialysis technique, we measured extracellular levels of nitric oxide (NO) metabolites (NO_x⁻) in the medial prefrontal cortex (mPFC) upon perfusion of γ-aminobutyric acid (GABA) receptor antagonists as well as agonists, and also examined the effects of GABA receptor agonists on mild intermittent footshock-induced NO releases in the mPFC in conscious rats. Perfusion of either bicuculline methiodide, a GABA_A receptor antagonist, or saclofen, a GABA_B receptor antagonist, through a microdialysis probe resulted in dose-dependent increases in NO_x⁻ levels. Higher-dose perfusion of either muscimol (50 μM), a GABA_A receptor agonist, or baclofen (250 μM), a GABA_B receptor agonist resulted in a significant decrease in NO_x⁻ levels. The elevated levels of NO_x⁻ after mild intermittent footshock were attenuated by perfusion of either muscimol (10 μM) or baclofen (50 μM), either of which alone did not affect basal NO_x⁻ levels. These findings are likely to provide helpful clues to our understanding of the inhibitory modulation of basal and footshock-induced NO metabolites releases by GABA_A and GABA_B receptors in the mPFC.     

Distribution of GABA binding site subtypes in rat pituitary gland

GABA_A and GABA_B binding sites in rat pituitary gland were investigated using equilibrium binding assays in vitro. Specific binding of both [³H]GABA and [³H]muscimol could be detected in both anterior and neurointermediate lobes, with a relative concentration in the anterior lobe. [³H]GABA binding was discriminated into GABA_A and GABA_B receptor type binding using baclofen. GABA_B sites were detectable in the anterior but not in the neurointermediate lobe. Saturation analysis of [³H]muscimol binding to whole pituitary gland membranes demonstrated that the pituitary contains two classes of GABA_A sites differing in affinity, as found in the CNS, although the number of sites is considerably lower than in the CNS.     

Zinc modulates GABAB binding in rat brain

The effects of ZnCl₂ on [³H]GABA binding to GABA_A and GABA_B binding sites were investigated using receptor autoradiography. At concentrations exceeding 100 μM, zinc non-competitively inhibited GABA_B binding in a dose dependent fashion. GABA_A binding was not inhibited significantly by zinc eliminating the possibility of a non-specific effect of zinc. Increased calcium concentrations up to 10 MM enhanced total GABA_B binding but did not prevent zinc induced inhibition of GABA_B binding, indicating a separate site of action for these cations at the GABA_B binding site. In some regions, zinc modulates GABA_B binding in a biphasic manner as concentrations of 10–100 μM zinc significantly enhanced GABA_B binding in the hippocampus and the molecular layer of the cerebellum but not in the thalamus. These results provide further evidence for a neuromodulatory role for zinc in the central nervous system.     

Distribution and depression of the GABAB receptor in the spinal dorsal horn of adult rat

γ-Aminobutyric acid (GABA) is a principal inhibitory neurotransmitter in vertebrate nervous system. The metabotropic receptor for GABA, GABA_B receptor, is characterized as a G protein-coupled receptor subtype. In the present study, GABA_B receptor-like immunoreactivity (GABA_BR-LI) in the rat spinal cord and dorsal root ganglion (DRG), as well as GABA_B receptor-mediated depression in the spinal dorsal horn were examined by using immunohistochemistry and whole-cell voltage-clamp recording technique, respectively. Under light microscope, GABA_BR-LI was densely found in laminae I and II of the dorsal horn. DRG cells of various diameters also showed GABA_BR-LI. Electron microscopy further revealed that GABA_BR-LI was also localized in terminals of myelinated, unmyelinated fibers as well as the somatodendritic sites of dorsal horn neurons. Bath application of a GABA_B receptor agonist, baclofen (10 μM, 30 s), induced a slow outward (inhibitory) current in dorsal horn neurons. This slow current was depressed when the postsynaptic G protein-coupled receptor was inhibited, indicating the postsynaptic action of baclofen. Under the condition of postsynaptic GABA_B receptor being inhibited, baclofen (10 μM, 60 s) depressed large (Aβ) and fine (C, Aδ) afferent fiber-evoked monosynaptic excitatory postsynaptic currents, indicating presynaptic inhibition of GABA_B receptor on elicited neurotransmitter release. Taken together, the results suggest that baclofen-sensitive GABA_B receptor is expressed pre- and postsynaptically on primary afferent fibers and neurons in the spinal dorsal horn; activation of GABA_B receptor in the dorsal horn postsynaptically hyperpolarizes dorsal horn neurons and presynaptically inhibits primary afferents.     

In vivo neurochemical evidence that stimulation of accumbal GABAA and GABAB receptors each reduce acetylcholine efflux without affecting dopamine efflux in the nucleus accumbens of freely moving rats

Cholinergic neurons in the nucleus accumbens contain GABA_A and GABA_B receptors that are thought to inhibit neural activity. We analyzed the roles of GABA_A and GABA_B receptors in regulating accumbal acetylcholine efflux of freely moving rats using in vivo microdialysis. The effects of GABA receptor ligands on the accumbal dopamine efflux were also analyzed because accumbal cholinergic and dopaminergic neurons could mutually interact. Drugs were applied intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 30–60 min infusions. To monitor basal acetylcholine, a low concentration of physostigmine (50 nM) was added to the perfusate. GABA_A receptor agonist muscimol (3 and 30 pmol) induced a dose‐related decrease in accumbal acetylcholine. GABA_B receptor agonist baclofen (30 and 300 pmol) also produced a dose‐related decrease in acetylcholine. GABA_A receptor antagonist bicuculline (60 pmol) which failed to alter baseline acetylcholine counteracted the muscimol (30 pmol)‐induced decrease in acetylcholine. GABA_B receptor antagonist 2‐hydroxysaclofen (12 nmol) which failed to change baseline acetylcholine, counteracted the baclofen (300 pmol)‐induced decrease in acetylcholine. Neither muscimol (30 pmol) nor baclofen (300 pmol) which reduced accumbal acetylcholine altered baseline accumbal dopamine. Neither bicuculline (60 pmol) nor 2‐hydroxysaclofen (12 nmol) also affected the baseline dopamine. These results show that GABA_A and GABA_B receptors each exert inhibitory roles in the regulation of accumbal cholinergic neural activity. The present results also provides in vivo neurochemical evidence that stimulation of GABA_A and GABA_B receptors each reduce acetylcholine efflux without affecting dopamine efflux in the nucleus accumbens of freely moving rats.     

Involvement of GABA systems in acetylcholine release induced by 5-HT3 receptor blockade in slices from rat entorhinal cortex

The aim of the present study was to examine the role of 5-HT₃ receptors in spontaneous and K⁺-evoked acetylcholine (ACh) release from rat entorhinal cortex and striatal slices. The 5-HT3 receptor antagonists ondansetron and granisetron (0.01–10 μM) produced a concentration-dependent increase in both spontaneous and K⁺-evoked [³H]ACh release in the two brain regions studied. The release of ACh was Ca²⁺-dependent and tetrodotoxin-sensitive. 5-HT₃ receptor agonists, such as 2-methyl-5-HT and 1-phenylbiguanide, at concentrations up to 1 μM, did not show any intrinsic effect on [³H]ACh release in both rat brain regions. However, 2-methyl-5-HT, 1 μM, fully blocked the ondansetron-induced enhancement in both basal and K⁺-evoked ACh release, suggesting that 5-HT₃ through 5-HT₃ receptor activation, tonically inhibits ACh release. The possible implication of interposed inhibitory systems in ACh release after 5-HT₃ receptor blockade was subsequently analyzed. While the effect of ondansetron was not modified by haloperidol or naloxone, the GABA_A receptor antagonist bicuculline produced a marked potentiation of ACh release in the entorhinal cortex but not in the striatum. The results suggest that in this cortical area 5-HT activates 5-HT₃ receptors located on GABAergic neurons which in turn inhibit cholinergic function.     

Involvement of GABAB receptors in convulsant‐induced epileptiform activity in rat neocortex in vitro

The role of γ-aminobutyric acid B (GABA_B) receptors in the generation and maintenance of bicuculline-induced epileptiform activity in rat neocortical slices was studied using electrophysiological methods. A block of GABA_B receptors in the presence of functional GABA_A receptor-mediated inhibition was not sufficient to induce epileptiform activity. In the presence of the GABA_A receptor antagonist bicuculline (10 μm ) and at suprathreshold stimulation, the GABA_B receptor antagonist CGP 35348 (10–300 μm ) significantly potentiated epileptiform activity. With stimulation at threshold intensity, low concentrations of CGP 35348 (10–30 μm ) potentiated bicuculline-induced activity, whereas higher concentrations (100–300 μm ) invariably led to a reversible suppression of stimulus-evoked epileptiform discharges. CGP 35348 also enhanced picrotoxin-induced epileptiform activity, but at higher concentrations it was considerably less effective in suppressing such epileptiform discharges. The GABA uptake inhibitor nipecotic acid partially mimicked the actions of CGP 35348: with stimulation at threshold intensity, it reversibly suppressed bicuculline-induced epileptiform field potentials, but it did not influence epileptiform activity induced by picrotoxin. We conclude that a postsynaptic blockade of GABA_B receptors induces an amplification of epileptiform activity in neocortical slices disinhibited by GABA_A receptor antagonists. An additional blockade of presynaptic GABA_B receptors, especially under conditions of weak stimulation of the neurons, reduces the inhibitory auto-feedback control of GABA release, leading to a displacement of competitive antagonists from the postsynaptic GABA_A receptor and hence, to a suppression of epileptiform activity induced by competitive GABA_A receptor antagonists.     

Subpopulations of neurons in rat substantia nigra display GABABR2 receptor immunoreactivity

A functional gamma-aminobutyric acid (GABA) B receptor is the first metabotropic receptor known to be composed of two heteromeric subunits, GABA_BR1 and GABA_BR2. Our previous report [Neuroscience 99 (2000) 65] has demonstrated that subpopulations of neurons in the rat substantia nigra display distinct patterns of distribution of GABA_BR1 receptor immunoreactivity. A robust level of GABA_BR1 receptor is only found in the dopaminergic neurons of the substantia nigra pars compacta (SNc). The objective of the present study was to determine the precise cellular localization of GABA_BR2 subunit in the rat substantia nigra using double immunofluorescence. Neuropilar elements in the SNc and the substantia nigra pars reticulata (SNr) were found to display GABA_BR2 immunoreactivity. In addition, the tyrosine hydroxylase-immunoreactive dopaminergic neurons and the parvalbumin-immunoreactive GABAergic neurons in the SNr were also found to display GABA_BR2 immunoreactivity. The present results thus demonstrate that a functional GABA_B receptor may be expressed by the dopaminergic neurons in the SNc. It is less clear whether neurons in the SNr express a functional GABA_B receptor. The present findings have important functional implications in GABA neurotransmission in the substantia nigra.     

Involvement of GABAB receptor systems in experimental depression: baclofen but not bicuculline exacerbates helplessness in rats

There are two γ-aminobutyric acid (GABA) hypotheses of the antidepressants action: an increase in GABA_A neurotransmission or a decrease in GABA_B neurotransmission may contribute to action of antidepressants. In this study, involvement of GABA_A and GABA_B, receptor systems was examined in the learned helplessness paradigm in rats. Rats were injected with bicuculline or baclofen for 14 days. On day 14, the rats were subjected to 15 inescapable shocks. On day 15, they underwent the 40-trial escape test. Baclofen exacerbated the escape failures in the rats subjected to the inescapable shocks, although baclofen had no effects in the animals without shock pre-treatment. Bicuculline failed to influence the escape failures in the rats with the 15-shock pre-treatment. These results suggest that the long-term increase in GABA_B, neurotransmission but not the long-term attenuation of GABA_A neurotransmission may be related to helplessness in rats.     

Opioid peptides modulate GABAA receptor responses in neurons of bullfrog dorsal root ganglia

Effects of enkephalin and selective opioid-receptor agonists on GABA-induced current were examined in dissociated neurons of bullfrog dorsal root ganglia (DRG) by using whole-cell patch-clamp method. Leucine-(Leu)-enkephalin and methionine-(Met)-enkephalin depressed GABA_A receptor-mediated currents. DPDPE, DAMGO and dynorphin-A (Dyn-A) also depressed the inward current produced by GABA: the order of agonist potency was DPDPE ≥ DAMGO> Dyn-A. Naloxone blocked the inhibitory effects of ekephalins and other opioid agonists on the GABA current. Naltrindole (NTI), a δ-receptor antagonist, prevented the DPDPE-induced depression of the GABA current. β-Funaltrexamine (β-FNA), a μ-receptor antagonist, reduced the DAMGO-induced depression of GABA currents. Nor-binaltorphimine (nor-BNI), a κ-receptor antagonist, reduced the effects of Dyn-A in depressing the GABA current The results suggest that enkephalin down-regulates GABA_A receptor function through mainly δ- and μ-opioid receptors in bullfrog DRG neurons.     

Effect of embryonic knock-down of GABAA receptors on the levels of monoamines and their metabolites in the CNS of the mouse

In vitro evidence indicates that γ-aminobutyric acid (GABA), acting at GABA_A receptors, exerts a positive trophic effect on monoaminergic neurons during embryogenesis. To determine whether in vivo antagonism of GABA_A receptors during embryogenesis interferes with the development of monoaminergic neurons, we used mice in which the number of GABA_A receptors was decreased by 50% by targeted deletion of the β₃ subunit gene of the GABA_A receptor. Levels of serotonin, dopamine, norepinephrine, and the metabolites 3,4-deoxyphenylacetic acid, homovanillic acid, and 5-hydroxyindoleacetic acid were measured in the brainstem, cortex, striatum and spinal cord of female adult homozygous null (β₃^−/−) and wild-type (β₃^+/+) mice, as well as progenitor C57BL/6J and Strain 129/SvJ mice. The level of norepinephrine in the spinal cord of β₃^−/− mice was 44% less than that of β₃^+/+ mice, and did not differ in the brainstem, cortex or striatum. This finding suggests that β₃ subunit-containing GABA_A receptors mediate the trophic effects of GABA on a subpopulation of spinally-projecting noradrenergic neurons. In contrast, the levels of serotonin, dopamine or their metabolites were unaffected, suggesting that the development of serotonergic and dopaminergic neurons may require activation of only a small fraction of GABA_A receptors or may not be dependent on β₃ subunit-containing GABA_A receptors. Finally, Strain 129/SvJ and C57BL/6J mice differed with respect to the levels of dopamine and its metabolites in the brainstem, spinal cord and cortex. These differences may need to be considered when assessing the phenotype of gene-targeted mice for which these mice serve as progenitor strains.