Postsynaptic GABA Research Articles

Confocal Analysis of Reciprocal Feedback at Rod Bipolar Terminals in the Rabbit Retina

Amacrine cells in the mammalian retina are famously diverse in shape and function. Here, we show that two wide-field GABA amacrine cells, S1 and S2, have stereotyped synaptic contacts with the appropriate morphology and distribution to perform specific functions. S1 and S2 both supply negative feedback to rod bipolar terminals and thus provide a substrate for lateral inhibition in the rod pathway. Synapses are specialized structures, and the presynaptic compartment is normally characterized by a swelling or varicosity. Each S1 amacrine cell has approximately 280 varicosities, whereas an S2 cell has even more, approximately 500 per cell. Confocal analysis shows that essentially all varicosities aggregate around rod bipolar terminals where they are apposed by postsynaptic GABA receptors. Each rod bipolar terminal is contacted by varicosities from approximately 25 different S1 and 50 different S2 amacrine cells. In fact, rod bipolar cells are the only synaptic target for S1 and S2 amacrine cells: all of the output from these two wide-field GABA amacrine cells goes to rod bipolar terminals. It has long been a puzzle why two amacrine cells, apparently with the same connections, are required. However, an analysis of the distribution of varicosities suggests that S1 and S2 amacrine cells provide different signals. S2 amacrine cells dominate within 200 mu from a rod bipolar terminal and can provide an inhibitory input with spatial characteristics that match the size of the surround signal recorded from AII amacrine cells in the rod pathway. In contrast, the larger, better-coupled S1 amacrine cells may provide a more distant network signal.

Dopamine D4Receptors Modulate GABAergic Signaling in Pyramidal Neurons of Prefrontal Cortex

Dopaminergic neurotransmission in the prefrontal cortex (PFC) plays an important role in regulating cognitive processes and emotional status. The dopamine D4 receptor, which is highly enriched in the PFC, is one of the principal targets of antipsychotic drugs. To understand the cellular mechanisms and functional implications of D4 receptors, we examined the impact of D4 receptors in PFC pyramidal neurons on GABAergic inhibition, a key element in the regulation of "working memory." Application of the D4 agonist N-(methyl)-4-(2-cyanophenyl)piperazinyl-3-methylbenzamide maleate caused a reversible decrease in postsynaptic GABA(A) receptor currents; this effect was blocked by the D4 antagonist 3-[(4-[4-chlorophenyl]piperazine-1-yl)methyl]-[1H]-pyrrolo[2,3-b]pyridine but not by the D2 antagonist sulpiride, suggesting mediation by D4 receptors. Application of PD168077 also reduced the GABA(A) receptor-mediated miniature IPSC amplitude in PFC pyramidal neurons recorded from slices. The D4 modulation of GABA(A) receptor currents was blocked by protein kinase A (PKA) activation and occluded by PKA inhibition. Inhibiting the catalytic activity of protein phosphatase 1 (PP1) also eliminated the effect of PD168077 on GABA(A) currents. Furthermore, disrupting the association of the PKA/PP1 complex with its scaffold protein Yotiao significantly attenuated the D4 modulation of GABA(A) currents, suggesting that Yotiao-mediated targeting of PKA/PP1 to the vicinity of GABA(A) receptors is required for the dopaminergic signaling. Together, our results show that activation of D4 receptors in PFC pyramidal neurons inhibits GABA(A) channel functions by regulating the PKA/PP1 signaling complex, which could underlie the D4 modulation of PFC neuronal activity and the actions of antipsychotic drugs.

GABA(B) receptor activation desensitizes postsynaptic GABA(B) and A(1) adenosine responses in rat hippocampal neurones.

Whole-cell recordings of EPSCs and G-protein-activated inwardly rectifying (GIRK) currents were made from cultured hippocampal neurones to determine the effect of long-term agonist treatment on the presynaptic and postsynaptic responses mediated by GABA(B) receptors (GABA(B)Rs). GABA(B)R-mediated presynaptic inhibition was unaffected by agonist (baclofen) treatment for up to 48 h, and was desensitized by about one-half after 96 h. In contrast, GABA(B)R-mediated GIRK currents were desensitized by a similar amount after only 2 h of agonist treatment. In addition, presynaptic inhibition mediated by A(1) adenosine receptors (A(1)Rs) was unaffected by prolonged GABA(B)R activation, whereas A(1)R-mediated GIRK currents were desensitized. Desensitization of postsynaptic GABA(B)R and A(1)R responses was blocked by the GABA(B)R antagonist (1-(S)-3,4-dichlorophenylethyl)amino-2-(S) hydroxypropyl-p-benzyl-phosphonic acid (CGP 55845A), but not by the A(1)R antagonist cyclopentyldipropylxanthine (DPCPX). GIRK current amplitude could be partially restored after baclofen treatment by either coapplication of baclofen and adenosine, or intracellular infusion of the non-hydrolysable GTP analog 5'-guanylylimidodiphosphate (Gpp(NH)p). Short-term (4-24 h) baclofen treatment also significantly desensitized the inhibition of postsynaptic voltage-gated calcium channels by activation of GABA(B)Rs or A(1)Rs. These results show that responses mediated by GABA(B)Rs and A(1)Rs desensitize differently in presynaptic and postsynaptic compartments, and demonstrate the heterologous desensitization of postsynaptic A1R responses.

Synaptic Anchoring of Glycine Receptors in Developing Collicular Neurons under Control of Metabotropic Glutamate Receptor Activity

The role of GlyRs in supramedullar neurons is still enigmatic: although present in the plasma membrane, they seem to be excluded from synaptic transmission. Two distinct stages of inhibitory synaptogenesis in cultured collicular neurons were defined on the basis of strychnine sensitivity of IPSCs, presence of EPSCs, KCC2 expression, and transmitter-induced Ca 2+ responses. In the absence of KCC2 and glutamatergic transmission, newborn VIAAT-labeled boutons contained GlyT2, released glycine, and accumulated postsynaptic gephyrin and GlyRs. With ongoing maturation, when neurons expressed KCC2 and displayed glutamatergic activity, inhibitory synapses contained GAD65, released GABA, and accumulated postsynaptic gephyrin and GABA ARs, but not GlyRs. This activity- and transmitter-specific regulation of GlyR distribution could be challenged by chronic blockade of group I mGluRs. It is thus concluded that glutamatergic transmission can contribute to the regulation of synaptic GlyR stabilization during inhibitory synapse development.

Relative picrotoxin insensitivity distinguishes ionotropic GABA receptor-mediated IPSCs in hippocampal interneurons

Inhibitory GABAergic signalling in the hippocampus plays an important role in synchronizing principal cells and regulating the excitability of this seizure-prone structure. Distinct mechanisms modulate release from GABAergic terminals in the hippocampus, depending on whether the postsynaptic partner is an interneuron or a principal cell. Here, we report that postsynaptic ionotropic GABA receptors in principal cells and interneurons also show a striking pharmacological difference. The broad-spectrum antagonist picrotoxin (PTX) was less potent at blocking IPSCs evoked in stratum radiatum interneurons than in pyramidal neurons in the CA1 region. GABA-evoked currents in membrane patches from interneurons showed a smaller mean unitary conductance than in patches from pyramidal neurons. Because retinal GABA C receptors show decreased picrotoxin sensitivity and conductance, we examined the effect of the GABA C receptor agonist cis-aminocrotonic acid (CACA). Although this agent evoked picrotoxin-resistant currents in interneurons, these were enhanced by the GABA A allosteric modulator pentobarbital. Moreover, both picrotoxin-resistant IPSCs and CACA-evoked currents were blocked by the GABA A receptor-selective antagonist bicuculline. The presence of relatively picrotoxin-resistant GABA A receptors in interneurons provides a potential target for agents to modulate the activity of sub-populations of hippocampal neurons.

Kinetics of etomidate actions on GABA A receptors in the rat spinal dorsal horn neurons

Electrophysiological properties of etomidate (ET)-induced current ( I ET) at different concentrations and effects of ET at clinically relevant concentrations (1–10 μM) on postsynaptic GABA A receptor function were investigated using whole-cell patch-clamp technique in mechanically dissociated rat spinal dorsal horn neurons. The results showed that ET actions were concentration-dependent: low concentrations (10 μM) of ET potentiated GABA-activated current ( I GABA), slowed activation, desensitization and deactivation of GABA A receptors; moderate concentrations (10–1000 μM) of ET directly activated and desensitized GABA A receptors; high concentrations (>1000 μM) of ET produced an inhibitory effect on I ET. In addition, ET prolonged the duration of GABAergic miniature inhibitory postsynaptic currents (mIPSCs) in the mechanically dissociated rat dorsal horn neurons. These results suggest that general anesthetics-induced changes at spinal level could significantly contribute to analgesia and general anesthesia.

3H]Tiagabine binding to GABA transporter-1 (GAT-1) in suicidal depression

Background: In depressive disorders, alterations of GABA concentrations and number of postsynaptic GABA related receptor binding sites, as well as antidepressant effects exerted by GABA agonists, suggest a pathogenetic involvement of the GABA system. Method: The binding of the presynaptic GABA ligand [ 3H]tiagabine to GABA transporter-1 (GAT-1) was studied in post mortem human frontal cortex and cingulate gyrus from 13 suicide victims and 19 controls without known neurological or psychiatric disorder. Results: No differences were found between the suicide victims and the controls with regard to the number of [ 3H]tiagabine binding sites ( B max) or apparent affinity ( K d). Limitations: The study was limited to two brain regions. Conclusion: Findings in other studies of alterations in the GABA system in depression seem according to the present results not to be associated with significant changes in the GABA uptake binding sites in the regions investigated.

Ethanol-Induced Death of Postnatal Hippocampal Neurons

Fetal alcohol exposure causes severe neuropsychiatric problems, but mechanisms of the ethanol-associated changes in central nervous system development are unclear. In vivo, ethanol's interaction with N-methyl- d-aspartate (NMDA) and γ-aminobutyric acid type A (GABA A) receptors may cause increased apoptosis in the immature forebrain. We examined whether ethanol affects survival of neonatal hippocampal neurons in primary cultures. A 6-day ethanol exposure killed hippocampal neurons with an LD50 of ∼25 mM. Elevated extracellular potassium or insulin-related growth factor 1 inhibited cell loss. Although potentiation of GABA A receptors or complete block of NMDA receptors also kills hippocampal neurons, pharmacological studies suggest that ethanol's interaction with GABA A and NMDA receptors is not sufficient to explain ethanol's effects on neuronal survival. Ca 2+ influx in response to depolarization was depressed >50% by chronic ethanol treatment. We suggest that chronic ethanol may promote neuronal loss through a mechanism affecting Ca 2+ influx in addition to effects on postsynaptic GABA and glutamate receptors.

GABAergic Terminals Are Required for Postsynaptic Clustering of Dystrophin But Not of GABAA Receptors and Gephyrin

In rat hippocampal cultures, we show by multilabeling immunocytochemistry that pyramidal cells, which receive little or no GABAergic input, mistarget alpha2-GABA(A) receptors and gephyrin to glutamatergic terminals. This mismatch does not occur in neurons innervated by numerous GABAergic terminals. A similar phenomenon has been reported for isolated autaptic hippocampal neurons (Rao et al., 2000). GABAergic synapses typically form multiple release sites apposed to GABA(A) receptor and gephyrin clusters. Remarkably, dystrophin, a protein highly abundant in skeletal muscle membranes, is extensively colocalized with alpha2-GABA(A) receptors exclusively opposite GABAergic terminals. In addition, selective apposition of syntrophin and beta-dystroglycan to GABAergic presynaptic terminals suggests that the entire dystrophin-associated protein complex (DPC) clusters at GABAergic synapses. In contrast to gephyrin and GABA(A) receptors, DPC proteins are not mistargeted to glutamatergic synapses, indicating independent clustering mechanisms. This was confirmed in hippocampal neurons cultured from GABA(A) receptor gamma2 subunit-deficient mice. Clustering of GABA(A) receptor and gephyrin in these neurons was strongly impaired, whereas clustering of dystrophin and associated proteins was unaffected by the absence of the gamma2 subunit. Our results indicate that accumulation of dystrophin and DPC proteins at GABAergic synapses occurs independently of postsynaptic GABA(A) receptors and gephyrin. We suggest that selective signaling from GABAergic terminals contributes to postsynaptic clustering of dystrophin.

Signaling cascade regulating long-term potentiation of GABA(A) receptor responsiveness in cerebellar Purkinje neurons.

Synaptic plasticity, a cellular basis of learning and memory, has been studied extensively at excitatory synapses. Although synaptic plasticity has also been reported at inhibitory synapses, the molecular mechanism remains elusive. Here we attempted to clarify the overall signaling cascades regulating the induction of inhibitory synaptic plasticity in the cerebellum. Rebound potentiation (RP), a long-lasting increase in GABA(A) receptor (GABA(A)R) responsiveness, is induced by postsynaptic depolarization of a Purkinje neuron (PN) at synapses formed with inhibitory interneurons (stellate or basket neurons). Previously, we showed that RP is suppressed by homosynaptic activation during depolarization through activation of the postsynaptic GABA(B) receptor (GABA(B)R). Activation of GABA(B)R reduces cAMP-dependent protein kinase (PKA) activity via the G(i)/G(o)-protein. Here we examined the molecular pathway through which PKA activity affects RP induction. We confirmed that inhibition of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) or PKA suppresses RP. We also found that inhibition of protein phosphatase 1 (PP-1) or calcineurin (PP-2B) impaired suppression of RP induction. Inhibition of either PP-1 or calcineurin abolished RP impairment by PKA inhibition, but not that by CaMKII inhibition. Antisense oligonucleotide-mediated knock down of DARPP-32, which is a substrate of PKA and calcineurin and inhibits PP-1 when phosphorylated by PKA, suppressed RP. Furthermore, activation of GABA(B)R inhibited CaMKII activation through PKA inhibition and PP-1 activity. These results suggest that calcineurin activation accompanied by PKA inhibition in a PN causes dephosphorylation of DARPP-32, which releases PP-1 from inhibition. PP-1 in turn inhibits CaMKII activity, which is then directly involved in the RP induction.

Phosphorylation of the recombinant ρ1 GABA receptor

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian brain. While a growing body of literature indicates that postsynaptic GABA receptors are regulated by phosphorylation, there is discrepancy as to the specific effects of phosphorylation on GABA receptor function. Here, we have identified phosphorylation sites on the human ρ1 GABA receptor for six protein kinases widely expressed in the brain: protein kinase C (PKC); cAMP-dependent protein kinase (PKA); calmodulin-dependent kinase (CaMKII); casein kinase (CKII); mitogen-activated protein kinase (MAPK); and cGMP-dependent protein kinase (PKG). We demonstrate that in nearly all cases, the consensus sites and actual phosphorylation sites do not agree supporting the risk of relying on a sequence analysis to identify potential phosphorylation sites. In addition, of the six kinases examined, only CKII phosphorylated the human ρ2 subunit. Site-directed mutagenesis of the phosphorylation sites, or activation/inhibition of select kinase pathways, did not alter the receptor sensitivity or maximal GABA-activated current of the ρ1 GABA receptor expressed in Xenopus laevis oocytes suggesting phosphorylation of ρ1 does not directly alter receptor properties. This study is a first and necessary step towards elucidating the regulation of ρ1 GABA receptors by phosphorylation.

Reduced endogenous GABA-mediated inhibition in the PVN on renal nerve discharge in rats with heart failure.

One characteristic of heart failure (HF) is increased sympathetic activation. The paraventricular nucleus (PVN) of the hypothalamus (involved in control of sympathetic outflow) has been shown to have increased neuronal activation during HF. This study examined the influence of endogenous GABA input (inhibitory in nature) into the PVN on renal sympathetic nerve discharge (RSND), arterial blood pressure (BP), and heart rate (HR) in rats with HF induced by coronary artery ligation. In alpha-chloralose- and urethane-anesthetized rats, microinjection of bicuculline (a GABA antagonist) into the PVN produced a dose-dependent increase in RSND, BP, and HR in both sham-operated control and HF rats. Bicuculline attenuated the increase in RSND and BP in HF rats compared with control rats. Alternatively, microinjection of the GABA agonist muscimol produced a dose-dependent decrease in RSND, BP, and HR in both control and HF rats. Muscimol was also less effective in decreasing RSND, BP, and HR in HF rats than in control rats. These results suggest that endogenous GABA-mediated input into the PVN of rats with HF is less effective in suppressing RSND and BP compared with control rats. This is partly due to the post-release actions of GABA, possibly caused by altered function of post-synaptic GABA receptors in the PVN of rats with HF. Reduced GABA-mediated inhibition in the PVN may contribute to increased sympathetic outflow, which is commonly observed during HF.

Activity Deprivation Reduces Miniature IPSC Amplitude by Decreasing the Number of Postsynaptic GABAAReceptors Clustered at Neocortical Synapses

Maintaining the proper balance between excitation and inhibition is necessary to prevent cortical circuits from either falling silent or generating epileptiform activity. One mechanism through which cortical networks maintain this balance is through the activity-dependent regulation of inhibition, but whether this is achieved primarily through changes in synapse number or synaptic strength is not clear. Previously, we found that 2 d of activity deprivation increased the amplitude of miniature EPSCs (mEPSCs) onto cultured visual cortical pyramidal neurons. Here we find that this same manipulation decreases the amplitude of mIPSCs. This occurs with no change in single-channel conductance but is accompanied by a reduction in the average number of channels open during the mIPSC peak and a reduction in the intensity of staining for GABA(A) receptors (GABA(A)Rs) at postsynaptic sites. In addition, the number of synaptic sites that express detectable levels of GABA(A)Rs was decreased by approximately 50% after activity blockade, although there was no reduction in the total number of presynaptic contacts. These data suggest that activity deprivation reduces cortical inhibition by reducing both the number of GABA(A)Rs clustered at synaptic sites and the number of functional inhibitory synapses. Because excitatory and inhibitory synaptic currents are regulated in opposite directions by activity blockade, these data suggest that the balance between excitation and inhibition is dynamically regulated by ongoing activity.

GABAergic Innervation Organizes Synaptic and Extrasynaptic GABAAReceptor Clustering in Cultured Hippocampal Neurons

We have studied the effects of GABAergic innervation on the clustering of GABA(A) receptors (GABA(A)Rs) in cultured hippocampal neurons. In the absence of GABAergic innervation, pyramidal cells form small (0.36 +/- 0.01 micrometer diameter) GABA(A)R clusters at their surface in the dendrites and soma. When receiving GABAergic innervation from glutamic acid decarboxylase-containing interneurons, pyramidal cells form large (1.62 +/- 0.08 micrometer breadth) GABA(A)R clusters at GABAergic synapses. This is accompanied by a disappearance of the small GABA(A)R clusters in the local area surrounding each GABAergic synapse. Although the large synaptic GABA(A)R clusters of any neuron contained all GABA(A)R subunits and isoforms expressed by that neuron, the small clusters not localized at GABAergic synapses showed significant heterogeneity in subunit and isoform composition. Another difference between large GABAergic and small non-GABAergic GABA(A)R clusters was that a significant proportion of the latter was juxtaposed to postsynaptic markers of glutamatergic synapses such as PSD-95 and AMPA receptor GluR1 subunit. The densities of both the glutamate receptor-associated and non-associated small GABA(A)R clusters were decreased in areas surrounding GABAergic synapses. However, no effect on the density or distribution of glutamate receptor clusters was observed. The results suggest that there are local signals generated at GABAergic synapses that induce both assembly of large synaptic GABA(A)R clusters at the synapse and disappearance of the small GABA(A)R clusters in the surrounding area. In the absence of GABAergic innervation, weaker GABA(A)R-clustering signals, generated at glutamatergic synapses, induce the formation of small postsynaptic GABA(A)R clusters that remain juxtaposed to glutamate receptors at glutamatergic synapses.

Glycine and GABAA receptor subunits on Renshaw cells: Relationship with presynaptic neurotransmitters and postsynaptic gephyrin clusters

Inhibitory synapses with large and gephyrin-rich postsynaptic receptor areas are likely indicative of higher synaptic strength. We investigated the presynaptic inhibitory neurotransmitter content (GABA, glycine, or both) and the presence and subunit composition of GABA(A) and glycine postsynaptic receptors in one example of gephyrin-rich synapses to determine neurochemical characteristics that could also contribute to enhance synaptic strength. Hence, we analyzed subunit receptor expression in gephyrin patches located on Renshaw cells, a type of spinal interneuron that receives powerful excitatory and inhibitory inputs and displays many large gephyrin patches on its surface. GABA(A) and glycine receptors were almost always colocalized inside Renshaw cell gephyrin clusters. According to the subunit-immunoreactivities detected, the composition of GABA(A) receptors was inferred to be either alpha(3)beta((2or3))gamma(2), alpha(5)beta((2or3))gamma(2), alpha(3)alpha(5)beta((2or3))gamma(2) or a combination of these. The types of neurotransmitters contained inside boutons presynaptic to Renshaw cell gephyrin patches were also investigated. The majority (60-75%) of terminals presynaptic to Renshaw cell gephyrin patches contained immunocytochemical markers for GABA as well as glycine, but a proportion contained markers only for glycine. Significantly, 40% of GABA(A) receptor clusters were opposed to presynaptic boutons that contained only glycinergic markers. We postulate that GABA and glycine corelease, and the presence of alpha3-containing GABA(A) receptors can enhance the postsynaptic current and contribute to strengthen inhibitory input on Renshaw cells. In addition, a certain degree of imprecision in the localization of postsynaptic GABA(A) receptors in regard to GABA release sites onto adult Renshaw cells was also found.

Postsynaptic and presynaptic GABA B receptor-mediated inhibition in rat superficial superior colliculus neurons

It has been reported that GABA B receptors are abundantly expressed in the superficial superior colliculus (sSC). To elucidate the action of GABA B receptors in the sSC, we tested the effect of a specific GABA B receptor agonist baclofen on sSC neurons in slices obtained from rats (16–22 days postnatal) using the whole-cell patch clamp technique. Bath application of baclofen (10–30 μM) elicited hyperpolarization associated with an increase in potassium conductance in morphologically different types of neurons including putative excitatory and inhibitory neurons. In addition, baclofen presynaptically suppressed electrically induced excitatory postsynaptic currents and inhibitory postsynaptic currents. These effects of baclofen were antagonized by a specific GABA B receptor antagonist CGP55845A (1 μM). The results suggest that the GABA B receptors have an inhibitory action at both post- and pre-synaptic sites in the sSC.

New concepts in neonatal seizures.

The immature brain is more prone to seizures than the older brain as a result of an imbalance between excitatory and inhibitory input. The depolarizing, rather than hyperpolarizing effect of GABA(A) during the first week of life in the rodent, and the delay in postsynaptic GABA(B) inhibition coupled with the over-expression of glutamatergic synapses contribute to this increased propensity toward seizures. It is now clear that seizures can be injurious to the immature brain, although the pattern of seizure-induced injury is age-related. While the immature brain is resistant to acute seizure-induced cell loss, there are functional abnormalities following seizures with impairment of visual-spatial memory and reduced seizure threshold. Neonatal seizures are also associated with a number of activity-dependent changes in brain development including altered synaptogenesis and reduction in neurogenesis. These results argue that neonatal seizures should no longer be considered as benign events.

Distinct localization of GABAB receptors relative to synaptic sites in the rat cerebellum and ventrobasal thalamus

Metabotropic gamma-aminobutyric acid receptors (GABA(B)Rs) are involved in modulation of synaptic transmission and activity of cerebellar and thalamic neurons. We used subtype-specific antibodies in pre- and postembedding immunohistochemistry combined with three-dimensional reconstruction of labelled profiles and quantification of immunoparticles to reveal the subcellular distribution of pre- and postsynaptic GABA(B)R1a/b and GABA(B)R2 in the rat cerebellum and ventrobasal thalamus. GABA(B)R1a/b and R2 were extensively colocalized in most brain regions including the cerebellum and thalamus. In the cerebellum, immunoreactivity for both subtypes was prevalent in the molecular layer. The most intense immunoreactivity was found in Purkinje cell spines with a high density of immunoparticles at extrasynaptic sites peaking at around 240 nm from glutamatergic synapses between spines and parallel fibre varicosities. This is in contrast to dendrites at sites around GABAergic synapses where sparse and random distribution was found for both subtypes. In addition, more than one-tenth of the synaptic membrane specialization of spine-parallel fibre synapses were labelled at pre- or postsynaptic sites. Weak immunolabelling for both subtypes was also seen in parallel fibres but only rarely in GABAergic axons. In the ventrobasal thalamus, immunolabelling for both receptor subtypes was intense over the dendritic field of thalamocortical cells. Electron microscopy demonstrated an extrasynaptic localization of GABA(B)R1a/b and R2 exclusively in postsynaptic elements. Quantitative analysis further revealed the density of GABA(B)R1a/b around GABAergic synapses was higher than glutamatergic synapses on thalamocortical cell dendrites. The distinct localization of GABA(B)Rs relative to synaptic sites in the cerebellum and ventrobasal thalamus suggests that GABA(B)Rs differentially regulate activity of different neuronal populations.

Early Development of Neuronal Activity in the Primate HippocampusIn Utero

Morphological studies suggest that the primate hippocampus develops extensively before birth, but little is known about its functional development. Patch-clamp recordings of hippocampal neurons and reconstruction of biocytin-filled pyramidal cells were performed in slices of macaque cynomolgus fetuses delivered by cesarean section. We found that during the second half of gestation, axons and dendrites of pyramidal cells grow intensively by hundreds of micrometers per day to attain a high level of maturity near term. Synaptic currents appear around midgestation and are correlated with the level of morphological differentiation of pyramidal cells: the first synapses are GABAergic, and their emergence correlates with the growth of apical dendrite into stratum radiatum. A later occurrence of glutamatergic synaptic currents correlates with a further differentiation of the axodendritic tree and the appearance of spines. Relying on the number of dendritic spines, we estimated that hundreds of new glutamatergic synapses are established every day on a pyramidal neuron during the last third of gestation. Most of the synaptic activity is synchronized in spontaneous slow ( approximately 0.1 Hz) network oscillations reminiscent of the giant depolarizing potentials in neonatal rodents. Epileptiform discharges can be evoked by the GABA(A) receptor antagonist bicuculline by the last third of gestation, and postsynaptic GABA(B) receptors contribute to the termination of epileptiform discharges. Comparing the results obtained in primates and rodents, we conclude that the template of early hippocampal network development is conserved across the mammalian evolution but that it is shifted toward fetal life in primate.

Gabapentin is not a GABA B receptor agonist

Recent experiments have demonstrated that formation of functional type B gamma-aminobutyric acid (GABA B) receptors requires co-expression of two receptor subunits, GABA B1 and GABA B2. Despite the identification of these subunits and a number of associated splice variants, there has been little convincing evidence of pharmacological diversity between GABA B receptors comprising different subunit combinations. However, Ng et al. [Mol. Pharmacol., 59 (2000) 144] have recently suggested a novel and important pharmacological difference between GABA B receptor heterodimers expressing the GABA B1a and GABA B1b receptor subunits. This study suggested that the antiepileptic GABA analogue gabapentin (Neurontin) is an agonist at GABA B receptors expressing the GABA B1a but not the GABA B1b receptor subunit. The importance of this finding with respect to identifying novel GABA B receptor subunit specific agonists prompted us to repeat these experiments in our own [ 35 S]− GTPγS binding and second messenger assay systems. Here we report that gabapentin was completely inactive at recombinant GABA B heterodimers expressing either GABA B1a or GABA B1b receptor subunits in combination with GABA B2 receptor subunits. In addition, in both CA1 and CA3 pyramidal neurones from rodent hippocampal slices we were unable to demonstrate any agonist-like effects of gabapentin at either pre- or post-synaptic GABA B receptors. In contrast, gabapentin activated a GABA A receptor mediated chloride conductance. Our data suggest that gabapentin is not a GABA B-receptor agonist let alone a GABA B receptor subunit selective agonist.