New insights on the role of gephyrin in regulating both phasic and tonic GABAergic inhibition in rat hippocampal neurons in culture

Gephyrin is a tubulin-binding protein that acts as a scaffold for clustering glycine and GABA_A receptors at postsynaptic sites. In this study, the role of gephyrin on GABA_A receptor function was assessed at the post-translational level, using gephyrin-specific single chain antibody fragments (scFv-gephyrin). When expressed in cultured rat hippocampal neurons as a fusion protein containing a nuclear localization signal, scFv-gephyrin were able to remove endogenous gephyrin from GABA_A receptor clusters. Immunocytochemical experiments revealed a significant reduction in the number of synaptic γ2-subunit containing GABA_A receptors and a significant decrease in the density of the GABAergic presynaptic marker vesicular GABA transporter (VGAT). These effects were associated with a slow down of the onset kinetics, a reduction in the amplitude and in the frequency of miniature inhibitory postsynaptic currents (mIPSCs). The quantitative analysis of current responses to ultrafast application of GABA suggested that changes in onset kinetics resulted from modifications in the microscopic gating of GABA_A receptors and in particular from a reduced entry into the desensitized state. In addition, hampering gephyrin function with scFv-gephyrin induced a significant reduction in GABA_A receptor-mediated tonic conductance. This effect was probably dependent on the decrease in GABAergic innervation and in GABA release from presynaptic nerve terminals. These results indicate that gephyrin is essential not only for maintaining synaptic GABA_A receptor clusters in the right position but also for regulating both phasic and tonic inhibition.     

Effects of GABA and related agents on the electrical activity of hypothalamic ventromedial nucleus neurons in vitro

Summary Functional and neurochemical evidence suggests significant GABA participation in the basomedial hypothalamus. We have investigated electrophysiological effects of GABA using in vitro recording from hypothalamic tissue slices. Exogenous GABA inhibited 94 out of 121 ventromedial hypothalamic (VMN) neurons tested. In sixty-one percent of these GABA-responsive neurons, the inhibitory action of GABA was blocked by GABA_A antagonists, bicuculline methiodide (BMI) and picrotoxin (PTX). Nevertheless, many (27/69) GABA-responsive neurons were not sensitive to GABA_A blockers: BMI and PTX failed to antagonize inhibitory action of GABA. Most, if it not all, of these inhibitions can be accounted for by GABA_B effects, since baclofen powerfully inhibited 42 of 44 neurons tested. In addition to blocking the inhibitory action of exogenous GABA, BMI (55%) and PTX (36%) also caused changes of neuronal activity indicating blockade of intrinsic GABA-ergic action. Altogether, our results showed that, in the VMN, GABA acts through not only GABA_A but also GABA_B receptors to inhibit neuronal activity, and that there is tonic inhibition by intrinsic GABA neurons. These GABA actions may participate in behaviorallyrelevant VMN hypothalamic mechanisms.     

Effects of the GABA-uptake inhibitor tiagabine in rat globus pallidus

To elucidate the cellular action of tiagabine, an inhibitor of GAT-1 GABA transporter, in the globus pallidus, whole-cell patch-clamp recordings were made from rat globus pallidus neurons in the acutely prepared brain slice. Superfusion of tiagabine significantly prolonged the decay kinetics of both action potential-dependent and -independent (tetrodotoxin-resistant) inhibitory postsynaptic currents (IPSCs) that were mediated by GABA_A receptors. Furthermore, it decreased the frequency of these IPSCs. The latter effect was reversed by the GABA_B receptor antagonist CGP55845, which alone had no effect, suggesting the involvement of presynaptic GABA_B receptors. Thus, tiagabine could inhibit or disinhibit globus pallidus neurons by increasing the activation of the GABA_A receptors and presynaptic GABA_B receptors, respectively. In the behaving animal, tiagabine when injected unilaterally into the globus pallidus caused consistent ipsilateral rotation of the rats indicative of increased inhibition of globus pallidus activity. This finding could be explained by the proposition that in the presence of tiagabine, prolonged action of GABA on GABA receptors would dominate over the inhibitory effect of tiagabine on GABA release. Our findings on the electrophysiological and behavioral effects of tiagabine in globus pallidus suggest that this basal ganglia nucleus is one of the sites of action of tiagabine and provides a rationale for investigating its involvement in epilepsy.     

GABA(B) receptor activation modulates GABA(A) receptor-mediated inhibition in chicken nucleus magnocellularis neurons

Neurons of nucleus magnocellularis (NM), a division of avian cochlear nucleus that performs precise temporal encoding, receive glutamatergic excitatory input solely from the eighth nerve and GABAergic inhibitory input primarily from the ipsilateral superior olivary nucleus. GABA activates both ligand-gated Cl^– channels [GABA_A receptors (GABA_ARs)] and G protein-coupled receptors (GABA_B receptors). The net effect of GABA_AR-mediated input to NM is inhibitory, although depolarizing. Several studies have shown that this shunting, inhibitory GABAergic input can evoke action potentials in postsynaptic NM neurons, which could interfere with their temporal encoding. While this GABA-mediated firing is limited by a low-voltage-activated K⁺ conductance, we have found evidence for a second mechanism. We investigated modulation of GABA_AR-mediated responses by GABA_BRs using whole cell recording techniques. Bath-applied baclofen, a GABA_BR agonist, produced dose-dependent suppression of evoked inhibitory postsynaptic currents (eIPSCs). This suppression was blocked by CGP52432 a potent and selective GABA_BR antagonist. Baclofen reduced the frequency but not the amplitude of miniature IPSCs (mIPSCs) and did not affect postsynaptic currents elicited by puff application of a specific GABA_AR agonist muscimol, suggesting a presynaptic mechanism for the GABA_BR-mediated modulation. Firing of NM neurons by synaptic stimulation of GABAergic inputs to NM was eliminated by baclofen. However, endogenous GABA_BR activity in the presynaptic inhibitory terminals was not observed. We propose that presynaptic GABA_BRs function as autoreceptors, regulating synaptic strength of GABA_AR-mediated inhibition, and prevent NM neurons from generating firing during activation of the inhibitory inputs.     

Targeting the interaction of GABAB receptors with CaMKII with an interfering peptide restores receptor expression after cerebral ischemia and inhibits progressive neuronal death in mouse brain cells and slices

Cerebral ischemia is the leading cause for long-term disability and mortality in adults due to massive neuronal death. Currently, there is no pharmacological treatment available to limit progressive neuronal death after stroke. A major mechanism causing ischemia-induced neuronal death is the excessive release of glutamate and the associated overexcitation of neurons (excitotoxicity). Normally, GABA_B receptors control neuronal excitability in the brain via prolonged inhibition. However, excitotoxic conditions rapidly downregulate GABA_B receptors via a CaMKII-mediated mechanism and thereby diminish adequate inhibition that could counteract neuronal overexcitation and neuronal death. To prevent the deleterious downregulation of GABA_B receptors, we developed a cell-penetrating synthetic peptide (R1-Pep) that inhibits the interaction of GABA_B receptors with CaMKII. Administration of this peptide to cultured cortical neurons exposed to excitotoxic conditions restored cell surface expression and function of GABA_B receptors. R1-Pep did not affect CaMKII expression or activity but prevented its T286 autophosphorylation that renders it autonomously and persistently active. Moreover, R1-Pep counteracted the aberrant downregulation of G protein-coupled inwardly rectifying K⁺ channels and the upregulation of N-type voltage-gated Ca²⁺ channels, the main effectors of GABA_B receptors. The restoration of GABA_B receptors activated the Akt survival pathway and inhibited excitotoxic neuronal death with a wide time window in cultured neurons. Restoration of GABA_B receptors and neuroprotective activity of R1-Pep was verified by using brain slices prepared from mice after middle cerebral artery occlusion (MCAO). Treatment with R1-Pep restored normal GABA_B receptor expression and GABA receptor-mediated K⁺ channel currents. This reduced MCAO-induced neuronal excitability and inhibited neuronal death. These results support the hypothesis that restoration of GABA_B receptor expression under excitatory conditions provides neuroprotection and might be the basis for the development of a selective intervention to inhibit progressive neuronal death after ischemic stroke.     

Enhanced activity of GABA receptors inhibits glutamate release induced by focal cerebral ischemia in rat striatum

Cerebral ischemia causes an excess release of glutamate, which can injure neurons. The striatum is one of the important regions vulnerable to hypoxia and ischemia. Using push–pull perfusion technique, we investigated the regulatory role of γ-aminobutyric acid (GABA) and its receptors in modifying the amount of glutamate in rat striatum with ischemia. Perfusion with exogenous GABA (1 mM) inhibited cerebral ischemia-induced glutamate release by as much as 47%. We further characterized relative roles of subtype receptors of GABA on glutamate release by using pharmacological tools. While baclofen (500 μM), a GABA_B receptor agonist, suppressed ischemia-induced glutamate release by 52%, GABA_B receptor antagonist saclofen (500 μM) failed to produce a significant increase of glutamate release. The GABA_A receptor agonist muscimol (500 μM) also reduced by 38% the release of glutamate induced by cerebral ischemia but the GABA_A receptor antagonist bicuculline (500 μM) had very little effect. The present study demonstrates that the excessive release of glutamate or the overly activated glutamate receptor, triggered by cerebral ischemia, can be down-regulated by exogenous GABA or by increased activity of GABA receptors, especially the presynaptic GABA_B receptors, which might be one of the important mechanisms to protect against striatum neuronal damage from over stimulation by excessive glutamate during ischemia.     

Bi-directional effects of GABA(B) receptor agonists on the mesolimbic dopamine system

The rewarding effect of drugs of abuse is mediated by activation of the mesolimbic dopamine system, which is inhibited by putative anti-craving compounds. Interestingly, different GABA(B) receptor agonists can exert similarly opposing effects on the reward pathway, but the cellular mechanisms involved are unknown. Here we found that the coupling efficacy (EC(50)) of G-protein-gated inwardly rectifying potassium (GIRK, Kir3) channels to GABA(B) receptor was much lower in dopamine neurons than in GABA neurons of the ventral tegmental area (VTA), depending on the differential expression of GIRK subunits. Consequently, in rodent VTA slices, a low concentration of the canonical agonist baclofen caused increased activity, whereas higher doses eventually inhibited dopamine neurons. At behaviorally relevant dosages, baclofen activated GIRK channels in both cell types, but the drug of abuse gamma-hydroxy-butyric acid (GHB) activated GIRK channels only in GABAergic neurons. Thus GABA(B) receptor agonists exert parallel cellular and behavioral effects due to the cell-specific expression of GIRK subunits.     

Stimulation of α1-adrenoceptors reduces glutamatergic synaptic input from primary afferents through GABAA receptors and T-type Ca channels

Activation of the descending noradrenergic system inhibits nociceptive transmission in the spinal cord. Although both α₁- and α₂-adrenoceptors in the spinal cord are involved in the modulation of nociceptive transmission, it is not clear how α₁-adrenoceptors regulate excitatory and inhibitory synaptic transmission at the spinal level. In this study, inhibitory and excitatory postsynaptic currents (IPSCs and EPSCs, respectively) were recorded from lamina II neurons in rat spinal cord slices. The specific α₁-adrenoceptor agonist phenylephrine significantly increased the frequency of GABAergic spontaneous IPSCs in a concentration dependent manner, and this effect was abolished by the α₁-adrenoceptor antagonist 2-(2,6-dimethoxyphenoxy)ethylaminomethyl-1,4-benzodioxane (WB4101). Phenylephrine also significantly reduced the amplitude of monosynaptic and polysynaptic EPSCs evoked from primary afferents. The inhibitory effect of phenylephrine on evoked monosynaptic glutamatergic EPSCs was largely blocked by the GABA_A receptor antagonist picrotoxin and, to a lesser extent, by the GABA_B receptor antagonist CGP55845. Furthermore, blocking T-type Ca²⁺ channels with amiloride or mibefradil diminished the inhibitory effect produced by phenylephrine or the GABA_A receptor agonist muscimol on monosynaptic EPSCs evoked from primary afferents. Collectively, these findings suggest that activation of α₁-adrenoceptors in the spinal cord increases synaptic GABA release, which attenuates glutamatergic input from primary afferents mainly through GABA_A receptors and T-type Ca²⁺ channels. This mechanism of presynaptic inhibition in the spinal cord may be involved in the regulation of nociception by the descending noradrenergic system.     

RGS2 modulates coupling between GABAB receptors and GIRK channels in dopamine neurons of the ventral tegmental area

Agonists of GABA(B) receptors exert a bi-directional effect on the activity of dopamine (DA) neurons of the ventral tegmental area, which can be explained by the fact that coupling between GABA(B) receptors and G protein-gated inwardly rectifying potassium (GIRK) channels is significantly weaker in DA neurons than in GABA neurons. Thus, low concentrations of agonists preferentially inhibit GABA neurons and thereby disinhibit DA neurons. This disinhibition might confer reinforcing properties on addictive GABA(B) receptor agonists such as gamma-hydroxybutyrate (GHB) and its derivatives. Here we show that, in DA neurons of mice, the low coupling efficiency reflects the selective expression of heteromeric GIRK2/3 channels and is dynamically modulated by a member of the regulator of G protein signaling (RGS) protein family. Moreover, repetitive exposure to GHB increases the GABA(B) receptor-GIRK channel coupling efficiency through downregulation of RGS2. Finally, oral self-administration of GHB at a concentration that is normally rewarding becomes aversive after chronic exposure. On the basis of these results, we propose a mechanism that might underlie tolerance to GHB.     

GABAC receptors are functionally expressed in the intermediate zone and regulate radial migration in the embryonic mouse neocortex

Radial neuronal migration in the cerebral cortex depends on trophic factors and the activation of different voltage- and ligand-gated channels. To examine the functional role of GABA_C receptors in radial migration we analyzed the effects of specific GABA_A and GABA_C receptor antagonists on the migration of BrdU-labeled neurons in vitro using organotypic neocortical slice cultures. These experiments revealed that the GABA_A specific inhibitor bicuculline methiodide facilitated neuronal migration, while the GABA_C specific inhibitor (1,2,5,6-tetrahydropyridine-4-yl) methylphosphinic-acid (TPMPA) impeded migration. Co-application of TPMPA and bicuculline methiodide or the unspecific ionotropic GABA receptor antagonist picrotoxin both impeded migration, suggesting that the GABA_C receptor mediated effects dominate. Addition of the specific GABA_C receptor agonist cis-4-aminocrotonic acid (CACA) also hampered migration, indicating that a physiological GABAergic stimulation is required for appropriate function. RT-PCR experiments using specific probes for GABA_C receptor mRNA and Western blot assays using an antibody directed against rho subunits revealed the expression of GABA_C receptor mRNA and translated GABA_C receptor protein in the immature cortex. Microfluorimetric Ca²⁺ imaging in neurons of identified cortical layers using Calcium Green revealed the functional expression of GABA_A and GABA_C receptors in the intermediate zone, while only GABA_A receptor mediated responses were observed in the upper cortical plate. In summary, these results demonstrate that activation of GABA_C receptors is a prerequisite for accurate migration and that GABA_C receptors are functionally expressed in the intermediate zone.     

GABAA receptors involved in sleep and anaesthesia: β1- versus β3-containing assemblies

The histaminergic neurons of the posterior hypothalamus (tuberomamillary nucleus—TMN) control wakefulness, and their silencing through activation of GABA_A receptors (GABA_AR) induces sleep and is thought to mediate sedation under propofol anaesthesia. We have previously shown that the β1 subunit preferring fragrant dioxane derivatives (FDD) are highly potent modulators of GABA_AR in TMN neurons. In recombinant receptors containing the β3N265M subunit, FDD action is abolished and GABA potency is reduced. Using rat, wild-type and β3N265M mice, FDD and propofol, we explored the relative contributions of β1- and β3-containing GABA_AR to synaptic transmission from the GABAergic sleep-on ventrolateral preoptic area neurons to TMN. In β3N265M mice, GABA potency remained unchanged in TMN neurons, but it was decreased in cultured posterior hypothalamic neurons with impaired modulation of GABA_AR by propofol. Spontaneous and evoked GABAergic synaptic currents (IPSC) showed β1-type pharmacology, with the same effects achieved by 3 μM propofol and 10 μM PI24513. Propofol and the FDD PI24513 suppressed neuronal firing in the majority of neurons at 5 and 100 μM, and in all cells at 10 and 250 μM, respectively. FDD given systemically in mice induced sedation but not anaesthesia. Propofol-induced currents were abolished (1–6 μM) or significantly reduced (12 μM) in β3N265M mice, whereas gating and modulation of GABA_AR by PI24513 as well as modulation by propofol were unchanged. In conclusion, β1-containing (FDD-sensitive) GABA_AR represent the major receptor pool in TMN neurons responding to GABA, while β3-containing (FDD-insensitive) receptors are gated by low micromolar doses of propofol. Thus, sleep and anaesthesia depend on different GABA_AR types.     

GABAA receptor inhibition does not affect mGluR-dependent LTD at hippocampal Schaffer collateral-CA1 synapses

Hippocampal synaptic plasticity between Schaffer collaterals and CA1 pyramidal neurons can be induced by activation of N-methyl-d-aspartate receptors (NMDARs) or of metabotropic glutamate receptors (mGluRs). Inhibitory GABAergic interneurons in this region abundantly terminate on pyramidal neurons and may thus influence synaptic plasticity. Although NMDAR-dependent synaptic plasticity is known to be influenced by inhibitory interneurons, little is known about the role of GABA on mGluR-dependent plasticity. Here, we used field potential recordings of the Schaffer collateral-CA1 synapses in rat hippocampal slices in order to study the effect of GABA_A receptor (GABA_AR) inhibition on mGluR-dependent long-term depression (LTD). Without GABA_AR blockade, mGluR-dependent LTD was induced pharmacologically by the group I mGluR agonist (RS)-3,5-dihydroxyphenylglycine (DHPG, 100 μM, 10 min) as well as electrically by paired-pulse low-frequency stimulation (PP-LFS, 900 paired pulses at 1 Hz) resulting in a stable depression of the field response lasting at least 80 min after LTD induction. The GABA_AR antagonist gabazine (5 μM) itself caused an increase of field responses suggesting an endogenous GABA release inhibiting CA1 field potentials. However, when either DHPG or PP-LFS was applied during GABA_AR inhibition, the field responses were significantly reduced. Moreover, normalizing these responses to experiments without GABA_AR blockade, there was no significant effect of gabazine on both DHPG- and PP-LFS-induced LTD. Thus, our results show that mGluR-dependent LTD at Schaffer collateral-CA1 synapses is unaffected by GABA_AR mediated synaptic transmission.     

Human locus coeruleus neurons express the GABAA receptor γ2 subunit gene and produce benzodiazepine binding

Noradrenergic neurons of the locus coeruleus project throughout the cerebral cortex and multiple subcortical structures. Alterations in the locus coeruleus firing are associated with vigilance states and with fear and anxiety disorders. Brain ionotropic type A receptors for γ-aminobutyric acid (GABA) serve as targets for anxiolytic and sedative drugs, and play an essential regulatory role in the locus coeruleus. GABA_A receptors are composed of a variable array of subunits forming heteropentameric chloride channels with different pharmacological properties. The γ2 subunit is essential for the formation of the binding site for benzodiazepines, allosteric modulators of GABA_A receptors that are clinically often used as sedatives/hypnotics and anxiolytics. There are contradictory reports in regard to the γ2 subunit's expression and participation in the functional GABA_A receptors in the mammalian locus coeruleus. We report here that the γ2 subunit is transcribed and participates in the assembly of functional GABA_A receptors in the tyrosine hydroxylase-positive neuromelanin-containing neurons within postmortem human locus coeruleus as demonstrated by in situ hybridization with specific γ2 subunit oligonucleotides and autoradiographic assay for flumazenil-sensitive [³H]Ro 15-4513 binding to benzodiazepine sites. These sites were also sensitive to the α1 subunit-preferring agonist zolpidem. Our data suggest a species difference in the expression profiles of the α1 and γ2 subunits in the locus coeruleus, with the sedation-related benzodiazepine sites being more important in man than rodents. This may explain the repeated failures in the transition of novel drugs with a promising neuropharmacological profile in rodents to human clinical usage, due to intolerable sedative effects.     

The physiological role of pre- and postsynaptic GABAB receptors in membrane excitability and synaptic transmission of neurons in the rat's dorsal cortex of the inferior colliculus

In the inferior colliculus (IC), GABAergic inhibition mediated by GABA_A receptors has been shown to play a significant role in regulating physiological responses, but little is known about the physiological role of GABA_B receptors in IC neurons. In the present study, we used whole-cell patch clamp recording in vitro to investigate the effects of activation of GABA_B receptors on membrane excitability and synaptic transmission of neurons in the rat's dorsal cortex of the inferior colliculus (ICD). Repetitive stimulation of GABAergic inputs to ICD neurons at high frequencies could elicit a slow and long-lasting postsynaptic response, which was reversibly abolished by the GABA_B receptor antagonist, CGP 35348. The results suggest that postsynaptic GABA_B receptors can directly mediate inhibitory synaptic transmission in ICD. The role of postsynaptic GABA_B receptors in regulation of membrane excitability was further investigated by application of the GABA_B receptor agonist, baclofen. Baclofen hyperpolarized the cell, reduced the membrane input resistance and firing rate, increased the threshold for generating action potentials (APs), and decreased the amplitude of the AP and its associated after-hyperpolarization. The Ca²⁺-mediated rebound depolarization following hyperpolarization and the depolarization hump at the beginning of membrane depolarization were also suppressed by baclofen. In voltage clamp experiments, baclofen induced inward rectifying K⁺ current and reduced low- and high-threshold Ca²⁺ currents, which may account for the suppression of membrane excitability by postsynaptic GABA_B receptors. Application of baclofen also reduced excitatory synaptic responses mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors, and inhibitory synaptic responses mediated by GABA_A receptors. Baclofen increased the ratios of 2nd/1st excitatory and inhibitory postsynaptic currents to paired-pulse stimulation of the synaptic inputs. These results suggest that fast glutamatergic and GABAergic synaptic transmission in ICD can be modulated by presynaptic GABA_B receptors.     

Tonic GABA inhibition in hippocampal dentate granule cells: its regulation and function in temporal lobe epilepsies

Both human and experimental evidence strongly supports the view of brain region‐ and cell‐specific changes in tonic GABA inhibition in temporal lobe epilepsies (TLE). This ‘tonic’ form of signalling is not time‐locked to presynaptic action potentials, which depends upon detection of ambient GABA by extrasynaptic GABA_A receptors (GABA_ARs). Extrasynaptic GABA_ARs have distinct physiological and pharmacological features, including high GABA‐binding affinity and low desensitization and a variety of the specific subunit combinations (α₄δ‐,α₆δ‐,α₅γ‐,ε‐containing receptors). These features closely contribute to the function of tonic GABA current, which is preserved properly or increased in dentate gyrus in models of TLE, even in the face of a loss of synaptic inhibition and inhibitory interneurones. Markedly reduced tonic GABA inhibition may facilitate an episode of epilepsy, while persistent elevated tonic inhibition may contribute to the onset of spontaneous recurrent seizures. In dentate granule cells, tonic GABA inhibition is positively modulated by endogenous neurosteroids and other factors, which undergo changes related to hormonal status after TLE. Tonic inhibition regulates neuronal excitability through its effects on membrane potential by both offsetting the threshold and reducing the frequency of action potentials and input resistance. Therefore, extrasynaptic GABA_ARs are expected to be the most important pharmacological targets in TLE. It is likely that both elevate the ambient GABA concentration and potentiate the tonic currents, contributing to the antiepileptic effects.     

Contribution of GABAC receptors to inhibition in the rodent accessory optic system

The medial terminal nucleus (MTN) of the mammalian accessory optic system controls vertical compensatory eye movements. It consists of two neuronal populations which respond best either to upward or to downward visual image shifts. The two cell classes are located spatially separate in the dorsal or in the ventral subdivision of the MTN, respectively. Pronounced GABAergic pathways have been described to exist between neurons in the two MTN subdivisions indicating that inhibitory interactions play a significant role for the generation of MTN cell response properties. Yet, the types of GABA receptors which mediate these inhibitory interactions are unknown. Functionally, it is of particular interest to know whether GABA_C receptors, as in other subcortical visual centers, participate in inhibitory mechanisms in MTN neurons. We therefore performed whole-cell patch clamp recordings from MTN neurons in acute mouse midbrain slices. We monitored excitatory and inhibitory postsynaptic responses to afferent stimulation and applied specific GABA receptor agonists and antagonists to identify the GABA receptor types present in MTN neurons. We found that more than 80% of the neurons in both MTN subdivisions express functional GABA_C receptors that can be activated by specific receptor agonists. A blockade of GABA_C receptors, on the other hand, either reduced or enhanced postsynaptic inhibition, indicating that both postsynaptic and presynaptic functions are served by this receptor type. This, together with earlier results, suggests that GABA_C receptors play a general role for the control of neuronal excitability in subcortical visual pathways.     

Developmental regulation and neuroprotective effects of striatal tonic GABAA currents

Striatal neurons are known to express GABA_A receptor subunits that underlie both phasic and tonic inhibition. Striatal projection neurons, or medium spiny neurons (MSNs), are divided into two classes: MSNs containing the dopamine D1 receptor (D1-MSNs) form the direct pathway to the substantia nigra and facilitate movement while MSNs expressing the dopamine D2 receptor (D2-MSNs) form the pallidal pathway that inhibits movement. Consequently, modulating inhibition in distinct classes of MSNs will differentially impact downstream network activity and motor behavior. Given the powerful role of extrasynaptic inhibition in controlling neuronal excitability, we examined the nature of striatal tonic inhibition and its potential role in preventing excitotoxicity. Consistent with earlier studies in young (P16–P25) mice, tonic GABA currents in D2-MSNs were larger than in D1-MSNs. However, with age (>P30 mice) the tonic GABA currents increased in D1-MSNs but decreased in D2-MSNs. These data demonstrate a developmental switch in the MSN subtype expressing larger tonic GABA currents. Compared to wild-type, MSNs from adult mice lacking the GABA_AR δ subunit (Gabrd^−/− mice) had both decreased tonic GABA currents and reduced survival following an in vitro excitotoxic challenge with quinolinic acid. Furthermore, muscimol-induced tonic GABA currents were accompanied by reduced acute swelling of striatal neurons after exposure to NMDA in WT mice but not in Gabrd^−/− mice. Our data are consistent with a role for tonic inhibition mediated by GABA_AR δ subunits in neuroprotection against excitotoxic insults in the adult striatum.     

Layer- and cell-type-specific tonic GABAergic inhibition of pyramidal neurons in the rat visual cortex

Tonic inhibition mediated by persistent activation of γ-aminobutyric acid_A (GABA_A) receptors by ambient GABA plays a crucial role in the regulation of network excitability and neuronal signal processing. Varying degrees in the strength of tonic inhibition were detected across different cell types throughout the brain. Since sensory information flows through cortical layers in a specific order, the characteristics of tonic inhibition in different cortical layers are of interest. Therefore, we examined the properties of tonic inhibition in pyramidal neurons (PyNs) throughout the rat visual cortex. Layer 2/3 PyNs and burst-spiking PyNs in layers 5 and 6 showed prominent tonic GABA_A currents. Tonic GABA_A currents in layer 4 star PyNs and regular-spiking PyNs in layers 5 and 6 were much weaker. The magnitude of tonic currents correlated well with the inhibition of spike generation. The amplitude of tonic GABA_A currents measured with bicuculline and gabazine, the two different GABA_A receptor blockers, did not differ. The differences in the expression levels of extrasynaptic GABA_A receptors might be the major contributor to the differences in tonic GABA_A currents among cell types. Furthermore, α5 subunits might contribute significantly to tonic currents in infragranular burst-spiking PyNs, especially in layer 5. These results suggest that ambient GABA might exert differential effects on the neuronal integration in a layer- and cell-type-specific manner and thus contribute to the processing of sensory properties by selectively tuning the signals flowing through the visual cortex.     

Simultaneous estimation of global background synaptic inhibition and excitation from membrane potential fluctuations in layer III neurons of the rat entorhinal cortex in vitro

It is becoming clear that the detection and integration of synaptic input and its conversion into an output signal in cortical neurons are strongly influenced by background synaptic activity or “noise.” The majority of this noise results from the spontaneous release of synaptic transmitters, interacting with ligand-gated ion channels in the postsynaptic neuron [Berretta N, Jones RSG (1996); A comparison of spontaneous synaptic EPSCs in layer V and layer II neurones in the rat entorhinal cortex in vitro. J Neurophysiol 76:1089–1110; Jones RSG, Woodhall GL (2005) Background synaptic activity in rat entorhinal cortical neurons: differential control of transmitter release by presynaptic receptors. J Physiol 562:107–120; LoTurco JJ, Mody I, Kriegstein AR (1990) Differential activation of glutamate receptors by spontaneously released transmitter in slices of neocortex. Neurosci Lett 114:265–271; Otis TS, Staley KJ, Mody I (1991) Perpetual inhibitory activity in mammalian brain slices generated by spontaneous GABA release. Brain Res 545:142–150; Ropert N, Miles R, Korn H (1990) Characteristics of miniature inhibitory postsynaptic currents in CA1 pyramidal neurones of rat hippocampus. J Physiol 428:707–722; Salin PA, Prince DA (1996) Spontaneous GABA_A receptor-mediated inhibitory currents in adult rat somatosensory cortex. J Neurophysiol 75:1573–1588; Staley KJ (1999) Quantal GABA release: noise or not? Nat Neurosci 2:494–495; Woodhall GL, Bailey SJ, Thompson SE, Evans DIP, Stacey AE, Jones RSG (2005) Fundamental differences in spontaneous synaptic inhibition between deep and superficial layers of the rat entorhinal cortex. Hippocampus 15:232–245]. The function of synaptic noise has been the subject of debate for some years, but there is increasing evidence that it modifies or controls neuronal excitability and, thus, the integrative properties of cortical neurons. In the present study we have investigated a novel approach [ Rudolph M, Piwkowska Z, Badoual M, Bal T, Destexhe A (2004) A method to estimate synaptic conductances from membrane potential fluctuations. J Neurophysiol 91:2884–2896] to simultaneously quantify synaptic inhibitory and excitatory synaptic noise, together with postsynaptic excitability, in rat entorhinal cortical neurons in vitro. The results suggest that this is a viable and useful approach to the study of the function of synaptic noise in cortical networks.     

Calcium currents and GABAB receptors in the dorsal sensory cells of the lamprey spinal cord

Patch-clamp studies were performed on the isolated dorsal sensory cells of the spinal cords of three species of lamprey,Ichthyomyzon unicuspis, Petromyzon marinus, andLampetra fluviatilis, to measure changes in the amplitudes of calcium current induced by GABA and its specific antagonists and agonists. The experiments showed that GABA (4 mM) reduced the peak amplitude of the calcium current by 28.5±4.9%, with subsequent recovery to 96.2±9.2% of control (n=45). The GABA_B agonist baclofen had similar effects. The GABA_A agonists glycine and taurine had no effect on the Ca²⁺ current. The inhibitory effect of GABA was blocked by 2-hydroxysaclofen (a GABA_B antagonist), but persisted in the presence of bicuculline (a GABA_A antagonist). These results are evidence that the membranes of dorsal sensory cells contain GABA_B receptors, which significantly increases our under-standing of the mechanisms of presynaptic inhibition in the spinal cords of the cyclostomata. Translated from Rossiiskii Fiziologischeskii Zhurnal imeni I. M. Sechenova, Vol. 83, No. 11-12, pp. 79–91, November–December, 1997.