BDNF-induced presynaptic facilitation of GABAergic transmission in the hippocampus of young adults is dependent of TrkB and adenosine A<Subscript>2A</Subscript> receptors

Brain-derived neurotrophic factor (BDNF) and adenosine are widely recognized as neuromodulators of glutamatergic transmission in the adult brain. Most BDNF actions upon excitatory plasticity phenomena are under control of adenosine A_2A receptors (A_2ARs). Concerning gamma-aminobutyric acid (GABA)-mediated transmission, the available information refers to the control of GABA transporters. We now focused on the influence of BDNF and the interplay with adenosine on phasic GABAergic transmission. To assess this, we evaluated evoked and spontaneous synaptic currents recorded from CA1 pyramidal cells in acute hippocampal slices from adult rat brains (6 to 10 weeks old). BDNF (10–100 ng/mL) increased miniature inhibitory postsynaptic current (mIPSC) frequency, but not amplitude, as well as increased the amplitude of inhibitory postsynaptic currents (IPSCs) evoked by afferent stimulation. The facilitatory action of BDNF upon GABAergic transmission was lost in the presence of a Trk inhibitor (K252a, 200 nM), but not upon p75^NTR blockade (anti-p75^NTR IgG, 50 μg/mL). Moreover, the facilitatory action of BDNF onto GABAergic transmission was also prevented upon A_2AR antagonism (SCH 58261, 50 nM). We conclude that BDNF facilitates GABAergic signaling at the adult hippocampus via a presynaptic mechanism that depends on TrkB and adenosine A_2AR activation.     

Propofol suppresses synaptic responsiveness of somatosensory relay neurons to excitatory input by potentiating GABAA receptor chloride channels

Propofol is a widely used intravenous general anesthetic. Propofol-induced unconsciousness in humans is associated with inhibition of thalamic activity evoked by somatosensory stimuli. However, the cellular mechanisms underlying the effects of propofol in thalamic circuits are largely unknown. We investigated the influence of propofol on synaptic responsiveness of thalamocortical relay neurons in the ventrobasal complex (VB) to excitatory input in mouse brain slices, using both current- and voltage-clamp recording techniques. Excitatory responses including EPSP temporal summation and action potential firing were evoked in VB neurons by electrical stimulation of corticothalamic fibers or pharmacological activation of glutamate receptors. Propofol (0.6 – 3 μM) suppressed temporal summation and spike firing in a concentration-dependent manner. The thalamocortical suppression was accompanied by a marked decrease in both EPSP amplitude and input resistance, indicating that a shunting mechanism was involved. The propofol-mediated thalamocortical suppression could be blocked by a GABA_A receptor antagonist or chloride channel blocker, suggesting that postsynaptic GABA_A receptors in VB neurons were involved in the shunting inhibition. GABA_A receptor-mediated inhibitory postsynaptic currents (IPSCs) were evoked in VB neurons by electrical stimulation of the reticular thalamic nucleus. Propofol markedly increased amplitude, decay time, and charge transfer of GABA_A IPSCs. The results demonstrated that shunting inhibition of thalamic somatosensory relay neurons by propofol at clinically relevant concentrations is primarily mediated through the potentiation of the GABA_A receptor chloride channel-mediated conductance, and such inhibition may contribute to the impaired thalamic responses to sensory stimuli seen during propofol-induced anesthesia.     

Opposing Actions of Sevoflurane on GABAergic and Glycinergic Synaptic Inhibition in the Spinal Ventral Horn

Background

The ventral horn is a major substrate in mediating the immobilizing properties of the volatile anesthetic sevoflurane in the spinal cord. In this neuronal network, action potential firing is controlled by GABA_A and glycine receptors. Both types of ion channels are sensitive to volatile anesthetics, but their role in mediating anesthetic-induced inhibition of spinal locomotor networks is not fully understood.

Methodology/Principal Findings

To compare the effects of sevoflurane on GABAergic and glycinergic inhibitory postsynaptic currents (IPSCs) whole-cell voltage-clamp recordings from ventral horn interneurons were carried out in organotypic spinal cultures. At concentrations close to MAC (minimum alveolar concentration), decay times of both types of IPSCs were significantly prolonged. However, at 1.5 MAC equivalents, GABAergic IPSCs were decreased in amplitude and reduced in frequency. These effects counteracted the prolongation of the decay time, thereby decreasing the time-averaged GABAergic inhibition. In contrast, amplitudes and frequency of glycinergic IPSCs were not significantly altered by sevoflurane. Furthermore, selective GABA_A and glycine receptor antagonists were tested for their potency to reverse sevoflurane-induced inhibition of spontaneous action potential firing in the ventral horn. These experiments confirmed a weak impact of GABA_A receptors and a prominent role of glycine receptors at a high sevoflurane concentration.

Conclusions

At high concentrations, sevoflurane mediates neuronal inhibition in the spinal ventral horn primarily via glycine receptors, and less via GABA_A receptors. Our results support the hypothesis that the impact of GABA_A receptors in mediating the immobilizing properties of volatile anesthetics is less essential in comparison to glycine receptors.     

Regulation of GABAergic inhibition by serotonin signaling in prefrontal cortex: molecular mechanisms and functional implications

Serotonergic neurotransmission in prefrontal cortex (PFC) plays a key role in regulating emotion and cognition under normal and pathological conditios. Increasing evidence suggests that serotonin receptors are involved in the complex regulation of GABAergic inhibitory transmission in PFC. Activation of postsynaptic 5-HT₂ receptors in PFC pyramidal neurons inhibits GABA_A-receptor currents via phosphorylation of GABA_A receptor γ2 subunits by RACK1-anchored PKC. In contrast, activation of postsynaptic 5-HT₄ receptors produces an activity-dependent bi-directional regulation of GABA-evoked currents in PFC pyramidal neurons, which is mediated through phosphorylation of GABA_A-receptor β subunits by anchored PKA. On the presynaptic side, GABAergic inhibition is regulated by 5-HT through the activation of 5-HT₂, 5-HT₁, and 5-HT₃ receptors on GABAergic intereneurons. These data provide a molecular and cellular mechanism for serotonin to dynamically regulate synaptic transmission and neuronal excitability in the PFC network, which may underlie the actions of many antidepressant and antipsychotic drugs.     

5-HT7 receptors modulate GABAergic transmission in rat hippocampal CA1 area

The effects of the activation of serotonin-7 (5-HT(7)) receptors were investigated in the CA1 area pyramidal cells and stratum radiatum fast spiking GABAergic interneurons of rat hippocampal slices. To activate 5-HT(7) receptors, 5-carboxamidotryptamine (5-CT), a nonselective 5-HT(1A)/5-HT(7) agonist, was applied in the presence of N-[2-[4-(2-methoxyphenyl)-1piperazinyl]ethyl]-N-2-pyridinylcyclohexanecarboxamide (WAY 100635), a selective 5-HT(1A) receptor antagonist. The activation of 5-HT(7) receptors resulted in a dose-dependent increase in the mean frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) recorded from pyramidal neurons while the mean amplitude of sIPSCs remained unaltered. A nonselective glutamate receptor antagonist, kynurenic acid, and voltage-gated sodium channel blocker, tetrodotoxin (TTX), attenuated but did not prevent the 5-HT(7) receptor-mediated increase of sIPSCs frequency in pyramidal cells. 5-CT application did not influence the excitability of stratum radiatum interneurons but it dose-dependently increased the mean frequency of spontaneous excitatory postsynaptic currents (sEPSCs) recorded from interneurons while the mean amplitude of sEPSCs remained unaltered. These data suggest that the activation of 5-HT(7) receptors results in an enhancement of the GABAergic transmission in the hippocampal CA1 area via two mechanisms. The first one involves an enhancement of excitatory glutamatergic input to GABAergic interneurons and is likely to be mediated by presynaptic 5-HT(7) receptors. The second effect, most likely related to the activation of 5-HT(7) receptors located on interneurons, results in an enhancement of the release of GABA.     

Quantum Properties of GABA Release in Cultured Neurons from the Rat Cortex

We analyzed the properties of inhibitory synaptic transmission between neurons in low-density cultures of cortical cells. Miniature, spontaneous, and evoked inhibitory postsynaptic currents were studied using a whole-cell path-clamp technique at a holding potential of -80 mV. These postsynaptic currents were identified as GABA release-activated Cl^- currents mediated by GABA_A receptors. Fitting amplitude histograms of these currents with Gaussian curves and an autocorrelation technique revealed the presence of equidistant peaks corresponding to a mean quantum size of 10 pA.     

Blocking L-type Voltage-gated Ca2+ Channels with Dihydropyridines Reduces ��-Aminobutyric Acid Type A Receptor Expression and Synaptic Inhibition

γ-Aminobutyric acid type A receptors (GABA_ARs) are the major sites of fast inhibitory neurotransmission in the brain, and the numbers of these receptors at the cell surface can determine the strength of GABAergic neurotransmission. Chronic changes in neuronal activity lead to an adaptive modulation in the efficacy of GABAergic synaptic inhibition, brought about in part by changes in the number of synaptic GABA_ARs, a mechanism known as homeostatic synaptic plasticity. Reduction in the number of GABA_ARs in response to prolonged neuronal activity blockade is dependent on the ubiquitin-proteasome system. The underlying biochemical pathways linking chronic activity blockade to proteasome-dependent degradation of GABA_ARs are unknown. Here, we show that chronic blockade of L-type voltage-gated calcium channels (VGCCs) with nifedipine decreases the number of GABA_ARs at synaptic sites but not the overall number of inhibitory synapses. In parallel, blockade of L-type VGCCs decreases the amplitude but not the frequency of miniature inhibitory postsynaptic currents or expression of the glutamic acid decarboxylase GAD65. We further reveal that the activation of L-type VGCCs regulates the turnover of newly translated GABA_AR subunits in a mechanism dependent upon the activity of the proteasome and thus regulates GABA_AR insertion into the plasma membrane. Together, these observations suggest that activation of L-type VGCCs can regulate the abundance of synaptic GABA_ARs and the efficacy of synaptic inhibition, revealing a potential mechanism underlying the homeostatic adaptation of fast GABAergic inhibition to prolonged changes in activity.     

Intracellular blockade of GABAA receptors in the rat hippocampal neurons

The intracellular blockade of GABA_A-receptor-mediated currents is a useful approach to suppress the GABAergic conductance in a single cell and to isolate the glutamatergic component of network-driven activities. Previously an approach has been described allowing intracellular blockade of GABA_A receptors by means of intracellular dialysis of a neuron with the pipette-filling solution, in which fluoride ions that hardly pass through the GABA_A receptor channels substitute for Cl^? and in which Mg²⁺ and ATP are omitted to induce rundown of the GABA_A receptors during whole-cell patch-clamp recordings. However, the kinetics of suppression of GABAergic conductance and the effect on the currents mediated by glutamate receptors remain unknown. Here, using whole-cell recordings with fluoride-based, Mg²⁺- and ATP-free solution on CA3 hippocampal neurons of neonatal rats, we show that after 1 h of such dialysis, both spontaneous and evoked GABA_A-receptor-mediated synaptic currents and responses induced by the GABA_A receptor agonist isoguvacine were completely suppressed. Inward GABAergic postsynaptic currents were suppressed prior to outward currents. Synaptic responses mediated by AMPA receptors were not affected by the dialysis, whereas the NMDA-receptor-mediated postsynaptic currents were reduced by approximately 20%. Dialysis with fluoride-based Mg²⁺, ATP-free solution either fully blocked giant depolarizing potentials (GDPs) in CA3 pyramidal cells (n = 2) or reduced the charge crossing the membrane during GDPs and shifted the GDP reversal potential to more positive values (n = 5). The dialysis-resistant component of GDPs was mediated by glutamate receptors, since: (i) it reversed around 0 mV; (ii) it demonstrated a negative slope conductance at negative membrane voltages, which is characteristic of NMDA receptor-mediated responses; (iii) kinetics of the individual events composing the dialysis-resistant component of GDPs at negative voltages were very similar to those of AMPA receptor-mediated synaptic currents. Thus, this procedure can be useful to isolate the glutamate receptor-mediated component of neuronal network-driven activities.     

Postnatal maturation of gamma-aminobutyric acidA and B-mediated inhibition in the CA3 hippocampal region of the rat

In the adult central nervous system, GABAergic synaptic inhibition is known to play a crucial role in preventing the spread of excitatory glutamatergic activity. This inhibition is achieved by a membrane hyperpolarization through the activation of postsynaptic γ-aminobutyric acid_A (GABA_A) and GABA_B receptors. In addition, GABA also depress transmitter release acting through presynaptic GABA_B receptors. Despite the wealth of data regarding the role of GABA in regulating the degree of synchronous activity in the adult, little is known about GABA transmission during early stages of development. In the following we report that GABA mediates most of the excitatory drive at early stages of development in the hippocampal CA3 region. Activation of GABA_A receptors induces a depolarization and excitation of immature CA3 pyramidal neurons and increases intracellular Ca²⁺ ([Ca²⁺]_i) during the first postnatal week of life. During the same developmental period, the postsynaptic GABA_B-mediated inhibition is poorly developed. In contrast, the presynaptic GABA_B-mediated inhibition is well developed at birth and plays a crucial role in modulating the postsynaptic activity by depressing transmitter release at early postnatal stages. We have also shown that GABA plays a trophic role in the neuritic outgrowth of cultured hippocampal neurons. © 1995 John Wiley & Sons, Inc.     

A Study of the Role of GABAA- and GABAB-Receptors in Presynaptic Inhibition of Primary Afferents in the Spinal Cord of the Frog Rana ridibunda

The role of GABA_A- and GABA_B-receptors in presynaptic inhibition of primary afferent fibers was studied on an isolated preparation of the spinal cord of the frog Rana ridibunda. It is shown that the inhibitory effect of GABA on synaptic transmission from afferent fiber to motoneuron is caused by activation of both GABA_A- and GABA_B-receptors. A temporal correlation (± 5 min) was shown between the blocking action of bicuculline (a specific antagonist of GABA_A-receptors) on primary afferent fiber depolarization (PAD) and its potentiating effect on the excitatory postsynaptic potential (EPSP) at parallel intracellular recording of EPSP in motoneuron and PAD in axons of the dorsal root. As a basis of this correlation, the single GABA_A-receptor mechanism is discussed, which mediates the effect of bicuculline on PAD and EPSP. When a specific agonist of GABA_B-receptor, baclofen, and an antagonist of GABA_B-receptor, 2(OH)-saclofen, were applied, the obtained data indicated an involvement of GABA_B-receptors in inhibition of synaptic transmission from afferent fibers to the motoneuron. Analysis of parameters of the unitary synaptic responses recorded in the control experiments and of their changes under the effect of (– )-baclofen indicates that the inhibitory action caused by activation of GABA_B-receptors develops at the presynaptic level.     

Postsynaptic target specificity of neurotrophin-induced presynaptic potentiation

The role of the target cell in neurotrophin-induced modifications of glutamatergic synaptic transmission was examined in cultured hippocampal neurons. Brain-derived neurotrophic factor (BDNF) induced rapid and persistent potentiation of evoked glutamate release when the postsynaptic neuron was glutamatergic, or excitatory (E-->E), but not when it was GABAergic, or inhibitory (E-->1). This target-specific action of BDNF was also found at divergent outputs of a single presynaptic neuron innervating both glutamatergic and GABAergic neurons, suggesting that individual terminals can be independently modified. Surprisingly, BDNF increased the frequency of miniature postsynaptic currents at both E-->E and E-->I, although it had no effect on evoked currents at E-->I. Finally, potentiation by neurotrophin-3 (NT-3) was also target specific. The selective effect at E-->E suggests that retrograde signaling by the postsynaptic target cell endows a localized presynaptic action of neurotrophins.     

The Reduction of EPSC Amplitude in CA1 Pyramidal Neurons by the Peroxynitrite Donor SIN-1 Requires Ca2+ Influx Via Postsynaptic Non-L-Type Voltage Gated Calcium Channels

The peroxynitrite free radical (ONOO^?) modulation of miniature excitatory postsynaptic currents (mEPSCs) and spontaneous excitatory postsynaptic currents (sEPSCs) was investigated in rat CA1 pyramidal neurons using the whole-cell patch clamp technique. SIN-1(3-morpholino-sydnonimine), which can lead the simultaneous generation of superoxide anion and nitric oxide, and then form the highly reactive species ONOO^?, induced dose-dependent inhibition in amplitudes of both mEPSCs and sEPSCs. The SIN-1 action on mEPSC amplitude was completely blocked by U0126, a selective MEK inhibitor, suggesting that MEK contributed to the action of ONOO^? on mEPSCs. The effect of SIN-1 was completely occluded either in the presence of the calcium chelator EGTA or the non-selective calcium channel antagonist Cd²⁺. Furthermore, the application of nifedipine (20 μM), the L-type calcium channel blocker, had no effect on the ONOO^?-induced decrease in mEPSC amplitude, excluding a role for L-type voltage-gated Ca²⁺ channels in this process. SIN-1 inhibited the frequency of sEPSCs but had no effect on mEPSC frequency, which suggested a presynaptic action potential-dependent the action of ONOO^? at CA1 pyramidal neuron synapses. The best-known glutamatergic input to CA1 pyramidal neurons is via Schaffer collaterals from CA3 area. However, no changes were observed in slices treated with SIN-1 on the spontaneous firing rates of CA3 pyramidal neurons. These findings suggested that SIN-1 inhibited glutamatergic synaptic transmission of CA1 pyramidal neurons by a postsynaptic non-L-type voltage gated calcium channel-dependent mechanism.     

Synaptogenesis in Co-Culture of Neurons of Dorsal Root Ganglia and Spinal Cord

In co-culture of spinal cord and dorsal root ganglion (DRG) neurons, we studied at different terms of culturing postsynaptic currents in DRG neurons evoked by direct electrical stimulation of single spinal neurons using a voltage-clamp technique in the whole-cell configuration. According to the reversal potential and sensitivity to bicuculline, these currents were classified as inhibitory postsynaptic currents (IPSC) carried by Cl^- ions through GABA_A receptors. During neuronal development in dissociated co-culture, the amplitude of evoked IPSC and their time to peak significantly increased. The time to peak of spontaneous IPSC (sIPSC) in DRG neurons remained unchanged, while the frequency of these currents increased with increasing culturing time. It is concluded that under culturing conditions spinal neurons establish inhibitory synaptic contacts with the somata of DRG neurons, and the number of such functional contacts increases in the course of culturing. Our findings show that in dissociated co-culture the process of formation of inhibitory synapses on the axon terminals of primary afferent neurons is akin to that realized in vivo, but with dissimilar topography of distribution of such synapses.     

Involvement of Ventral Periaqueductal Gray Dopaminergic Neurons in Propofol Anesthesia

It has been reported that central dopaminergic system is implicated in the mechanism underlying general anesthesia. Whether dopamine (DA) neurons in midbrain ventral periaqueductal gray (vPAG) are involved in general anesthesia and how general anesthetics affect these neurons remain sparsely documented. To determine the role of vPAG DA neurons in propofol-induced anesthesia, we performed microinjection of 6-hydroxydopamine (6-OHDA) into vPAG to damage DA neurons and investigated the alteration in somatosensory electroencephalogram (EEG), as well as the induction and recovery time of propofol anesthesia. Subsequently, we examined the effect of propofol on the electrophysiological activity of DA neurons in vPAG using whole-cell patch clamp. Two weeks after 6-OHDA microinfusion, DA neurons in the vPAG were markedly reduced by 63.6% in the 6-OHDA-treated rats compared with vehicle rats. This lesion significantly shortened the induction time (7.15?±?3.97 s vs. 11.18?±?2.83 s, P?<?0.05) and prolonged the recovery time of propofol anesthesia (780.26?±?150.86 s vs. 590.68?±?107.97 s, P?<?0.05). Meanwhile, EEG in somatosensory cortex revealed that delta power (0–4 Hz) was significantly higher in 6-OHDA-treated rats than vehicle rats. In the electrophysiological experiment, propofol decreased the frequency of spontaneous excitatory postsynaptic currents rather than the amplitude and decay time. In addition, propofol preferentially increased the frequency and prolonged the decay time of spontaneous inhibitory postsynaptic currents without affecting the amplitude. Significance: Propofol can promote presynaptic GABA release, inhibit presynaptic glutamate release and increase postsynaptic GABA_A receptor sensitivity, which eventually inhibits the activity of vPAG DA neurons and thereby influences the state of consciousness.     

Differential Modulation of GABAA Receptors Underlies Postsynaptic Depolarization- and Purinoceptor-Mediated Enhancement of Cerebellar Inhibitory Transmission: A Non-Stationary Fluctuation Analysis Study

Cerebellar GABAergic inhibitory transmission between interneurons and Purkinje cells (PCs) undergoes a long-lasting enhancement following different stimulations, such as brief depolarization or activation of purinergic receptors of postsynaptic PCs. The underlying mechanisms, however, are not completely understood. Using a peak-scaled non-stationary fluctuation analysis, we therefore aimed at characterizing changes in the electrophysiological properties of GABA_A receptors in PCs of rat cerebellar cortex during depolarization-induced “rebound potentiation (RP)” and purinoceptor-mediated long-term potentiation (PM-LTP), because both RP and PM-LTP likely depend on postsynaptic mechanisms. Stimulation-evoked inhibitory postsynaptic currents (eIPSCs) were recorded from PCs in neonatal rat cerebellar slices. Our analysis showed that postsynaptic membrane depolarization induced RP of eIPSCs in association with significant increase in the number of synaptic GABA_A receptors without changing the channel conductance. By contrast, bath application of ATP induced PM-LTP of eIPSCs with a significant increase of the channel conductance of GABA_A receptors without affecting the receptor number. Pretreatment with protein kinase A (PKA) inhibitors, H-89 and cAMPS-Rp, completely abolished the PM-LTP. The CaMKII inhibitor KN-62 reported to abolish RP did not alter PM-LTP. These results suggest that the signaling mechanism underlying PM-LTP could involve ATP-induced phosphorylation of synaptic GABA_A receptors, thereby resulting in upregulation of the channel conductance by stimulating adenylyl cyclase-PKA signaling cascade, possibly via activation of P2Y₁₁ purinoceptor. Thus, our findings reveal that postsynaptic GABA_A receptors at the interneuron-PC inhibitory synapses are under the control of two distinct forms of long-term potentiation linked with different second messenger cascades.     

Glucagon-like peptide-1 modulates synaptic transmission to identified pancreas-projecting vagal motoneurons

Using a brainstem slice preparation, we aimed to study the pre- and postsynaptic effects of glucagon-like peptide-1 (GLP-1) on synaptic transmission to identified pancreas-projecting vagal motoneurons. Following blockade of GABAergic mediated currents with bicuculline, perfusion with 100 nM GLP-1 increased both amplitude and frequency of excitatory postsynaptic currents (EPSCs) in 21 of 52 neurons. Perfusion with the GLP-1 selective agonist exendin-4 (100 nM), also increased the frequency of spontaneous EPSCs, while pretreatment with the GLP-1 selective antagonist, exendin 9-39, prevented the effects of GLP-1. In the presence of kynurenic acid to block ionotropic glutamatergic currents, perfusion with GLP-1 increased the frequency of inhibitory postsynaptic currents (IPSCs) in 28 of 74 neurons; in 14 of these responsive neurons, GLP-1 also increased IPSC amplitude, indicating actions at both pre- and postsynaptic sites. Perfusion with exendin-4 increased the frequency of spontaneous IPSCs, while pretreatment with exendin 9-39 prevented the effects of GLP-1. These results suggest that GLP-1 modulates both excitatory and inhibitory synaptic inputs to pancreas-projecting vagal motoneurons.     

A metabotropic glutamate receptor regulates transmitter release from cone presynaptic terminals in carp retinal slices.

The role of group III metabotropic glutamate receptors (mGluRs) in photoreceptor-H1 horizontal cell (HC) synaptic transmission was investigated by analyzing the rate of occurrence and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) in H1 HCs uncoupled by dopamine in carp retinal slices. Red light steps or the application of 100 microM cobalt reduced the sEPSC rate without affecting their peak amplitude, which is consistent with hyperpolarization or the suppression of Ca(2+) entry into cone synaptic terminals reducing vesicular transmitter release. Conversely, postsynaptic blockade of H1 HC AMPA receptors by 500 nM CNQX reduced the amplitude of sEPSCs without affecting their rate. This analysis of sEPSCs represents a novel methodology for distinguishing between presynaptic and postsynaptic sites of action. The selective agonist for group III mGluRs, l-2-amino-4-phosphonobutyrate (L-APB or L-AP4; 20 microM), reduced the sEPSC rate with a slight reduction in amplitude, which is consistent with a presynaptic action on cone synaptic terminals to reduce transmitter release. During L-APB application, recovery of sEPSC rate occurred with 500 microM (s)-2-methyl-2-amino-4-phosphonobutyrate (MAP4), a selective antagonist of group III mGluR, and with 200 microM 4-aminopyridine (4-AP), a blocker of voltage-dependent potassium channels. Whole-cell recordings from cones in the retinal slice showed no effect of L-APB on voltage-activated Ca(2+) conductance. These results suggest that the activation of group III mGluRs suppresses transmitter release from cone presynaptic terminals via a 4-AP-sensitive pathway. Negative feedback, operating via mGluR autoreceptors, may limit excessive glutamate release from cone synaptic terminals.     

Flotillin‐1 promotes formation of glutamatergic synapses in hippocampal neurons

Synapse malformation underlies numerous neurodevelopmental illnesses, including autism spectrum disorders. Here we identify the lipid raft protein flotillin‐1 as a promoter of glutamatergic synapse formation. We cultured neurons from the hippocampus, a brain region important for learning and memory, and examined them at two weeks in vitro, a time period rich with synapse formation. Double‐label immunocytochemistry of native flot‐1 with glutamatergic and GABAergic synapse markers showed that flot‐1 was preferentially colocalized with the glutamatergic presynaptic marker vesicular glutamate transporter 1 (VGLUT1), compared to the GABAergic presynaptic marker glutamic acid decarboxylase‐65 (GAD‐65). Triple‐label immunocytochemistry of native flot‐1, VGLUT1, and NR1, the obligatory subunit of NMDA receptors, indicates that Flot‐1 was preferentially localized to synaptic rather than extrasynaptic NR1. Furthermore, electrophysiological results using whole‐cell patch clamp showed that Flot‐1 increased the frequency of miniature excitatory postsynaptic currents (mEPSCs) but not miniature inhibitory postsynaptic currents (mIPSCs), whereas amplitude and decay kinetics of either type of synaptic current was not affected. Corresponding immunocytochemical data confirmed that the number of glutamatergic synapses increased with flot‐1 overexpression. Overall, our anatomical and physiological results show that flot‐1 enhances the formation of glutamatergic synapses but not GABAergic synapses, suggesting that the role of flot‐1 in neurodevelopmental disorders should be explored. © 2010 Wiley Periodicals, Inc. Develop Neurobiol 70: 875–883, 2010     

Actions of a novel water-soluble benzodiazepine-receptor agonist JM-1232(-) on synaptic transmission in adult rat spinal substantia gelatinosa neurons

Although the intrathecal administration of JM-1232(-) reportedly produces antinociception, this action has not yet been examined at the cellular level. We examined the action of JM-1232(-) on synaptic transmission in spinal substantia gelatinosa (SG) neurons which play an important role in regulating nociceptive transmission from the periphery. The whole-cell patch-clamp technique was applied to the SG neurons of adult rat spinal cord slices. Bath-applied JM-1232(-) prolonged the decay phase of GABA(A)-receptor mediated spontaneous inhibitory postsynaptic current (sIPSC) and increased its frequency without a change in amplitude. The former but not latter action was sensitive to a benzodiazepine-receptor antagonist flumazenil. JM-1232(-) also increased glycinergic sIPSC frequency with no change in amplitude and decay phase. On the other hand, glutamatergic spontaneous excitatory transmission was unaffected by JM-1232(-). These results indicate that JM-1232(-) enhances inhibitory transmission by (1) prolonging the decay phase of GABAergic sIPSC through benzodiazepine-receptor activation and by (2) increasing the spontaneous release of GABA and glycine from nerve terminals without its activation. This enhancement could contribute to at least a part of the antinociceptive effect of intrathecally-administered JM-1232(-).     

Rescue of Inhibitory Synapse Strength following Developmental Hearing Loss

Inhibitory synapse dysfunction may contribute to many developmental brain disorders, including the secondary consequences of sensory deprivation. In fact, developmental hearing loss leads to a profound reduction in the strength of inhibitory postsynaptic currents (IPSCs) in the auditory cortex, and this deficit persists into adulthood. This finding is consistent with the general theory that the emergence of mature synaptic properties requires activity during development. Therefore, we tested the prediction that inhibitory strength can be restored following developmental hearing loss by boosting GABAergic transmission in vivo. Conductive or sensorineural hearing loss was induced surgically in gerbils prior to hearing onset and GABA agonists were then administered for one week. IPSCs were subsequently recorded from pyramidal neurons in a thalamocortical brain slice preparation. Administration of either a GABA_A receptor a1 subunit specific agonist (zolpidem), or a selective GABA reuptake inhibitor (SGRI), rescued IPSC amplitude in hearing loss animals. Furthermore, this restoration persisted in adults, long after drug treatment ended. In contrast, a GABA_B receptor agonist baclofen did not restore inhibitory strength. IPSCs could also be restored when SGRI administration began 3 weeks after sensory deprivation. Together, these results demonstrate long-lasting restoration of cortical inhibitory strength in the absence of normal experience. This suggests that in vivo GABA_A receptor activation is sufficient to promote maturation, and this principle may extend to other developmental disorders associated with diminished inhibitory function.