In-Sun Choi

Presynaptic glycine receptors facilitate spontaneous glutamate release onto hilar neurons in the rat hippocampus

Although glycine receptors are found in most areas of the brain, including the hippocampus, their functional significance remains largely unknown. In the present study, we have investigated the role of presynaptic glycine receptors on excitatory nerve terminals in spontaneous glutamatergic transmission. Spontaneous EPSCs (sEPSCs) were recorded in mechanically dissociated rat dentate hilar neurons attached with native presynaptic nerve terminals using a conventional whole‐cell patch recording technique under voltage‐clamp conditions. Exogenously applied glycine or taurine significantly increased the frequency of sEPSCs in a concentration‐dependent manner. This facilitatory effect of glycine was blocked by 1 μM strychnine, a specific glycine receptor antagonist, but was not affected by 30 μM picrotoxin. In addition, Zn2+ (10 μM) potentiated the glycine action on sEPSC frequency. Pharmacological data suggested that the activation of presynaptic glycine receptors directly depolarizes glutamatergic terminals resulting in the facilitation of spontaneous glutamate release. Bumetanide (10 μM), a specific Na‐K‐2C co‐transporter blocker, gradually attenuated the glycine‐induced sEPSC facilitation, suggesting that the depolarizing action of presynaptic glycine receptors was due to a higher intraterminal Cl− concentration. The present results suggest that presynaptic glycine receptors on excitatory nerve terminals might play an important role in the excitability of the dentate gyrus‐hilus‐CA3 network in physiological and/or pathological conditions.

P2X7 receptors enhance glutamate release in hippocampal hilar neurons.

We examined the effect of 2′-3′-O-(4-benzoylbenzoyl)-adenosine-5′-triphosphate (Bz-ATP), a P2X7 receptor agonist, on action potential-independent glutamate release from nerve terminals attached to mechanically isolated immature hilar neurons. Bz-ATP increased spontaneous excitatory postsynaptic current (sEPSC) frequency, and this effect was blocked by Brilliant blue G, a P2X7 receptor antagonist, suggesting that P2X7 receptors mediate the facilitatory action of Bz-ATP on sEPSCs. In most of hilar neurons tested, the Bz-ATP-induced increase in sEPSC frequency was blocked by tetrodotoxin or Cd2+, suggesting that the activation of P2X7 receptors leads to a presynaptic depolarization. The P2X7 receptor-mediated facilitation of glutamate release would modulate the excitability of hilar neurons, and eventually have a broad impact on the pathophysiological functions mediated by the hippocampus.

Adenosine A1 receptors inhibit GABAergic transmission in rat tuberomammillary nucleus neurons

The adenosinergic modulation of GABAergic spontaneous miniature inhibitory postsynaptic currents (mIPSCs) was investigated in mechanically dissociated rat tuberomammillary nucleus (TMN) neurons using a conventional whole‐cell patch clamp technique. Adenosine (100 μM) reversibly decreased mIPSC frequency without affecting the current amplitude, indicating that adenosine acts presynaptically to decrease the probability of spontaneous GABA release. The adenosine action on GABAergic mIPSC frequency was completely blocked by 1 μM DPCPX, a selective A1 receptor antagonist, and mimicked by 1 μM CPA, a selective A1 receptor agonist. This suggests that presynaptic A1 receptors were responsible for the adenosine‐mediated inhibition of GABAergic mIPSC frequency. CPA still decreased GABAergic mIPSC frequency even either in the presence of 200 μM Cd2+, a general voltage‐dependent Ca2+ channel blocker, or in the Ca2+‐free external solution. However, the inhibitory effect of CPA on GABAergic mIPSC frequency was completely occluded by 1 mM Ba2+, a G‐protein coupled inwardly rectifying K+ (GIRK) channel blocker. In addition, the CPA‐induced decrease in mIPSC frequency was completely occluded by either 100 μM SQ22536, an adenylyl cyclase (AC) inhibitor, or 1 μM KT5720, a specific protein kinase A (PKA) inhibitor. The results suggest that the activation of presynaptic A1 receptors decreases spontaneous GABAergic transmission onto TMN neurons via the modulation of GIRK channels as well as the AC/cAMP/PKA signal transduction pathway. This adenosine A1 receptor‐mediated modulation of GABAergic transmission onto TMN neurons may play an important role in the fine modulation of the excitability of TMN histaminergic neurons as well as the regulation of sleep‐wakefulness.

5‐HT1B receptors inhibit glutamate release from primary afferent terminals in rat medullary dorsal horn neurons

BACKGROUND AND PURPOSE Although 5‐HT1B receptors are expressed in trigeminal sensory neurons, it is still not known whether these receptors can modulate nociceptive transmission from primary afferents onto medullary dorsal horn neurons.

Multiple effects of bisphenol A, an endocrine disrupter, on GABAA receptors in acutely dissociated rat CA3 pyramidal neurons

Bisphenol A (BPA), an endocrine disrupter, is contained in cans, polycarbonate bottles and some dental sealants. While the toxicological effects of BPA on the endocrine system have been extensively studied, its action on the central nervous system is poorly understood. Herein, we report the effects of BPA on GABA-induced currents (I(GABA)), using a conventional whole-cell patch clamp technique from acutely isolated rat CA3 pyramidal neurons. By itself, BPA concentration-dependently elicited the membrane current, which was significantly blocked by bicuculline, a selective GABA(A) receptor antagonist. BPA potentiated the peak I(GABA) induced by lower concentrations of GABA (<10 microM) in a concentration-dependent manner. The extent of BPA-induced potentiation of I(GABA) was significantly reduced by either diazepam or ethanol, allosteric modulators of GABA(A) receptors. BPA, however, inhibited the peak I(GABA) induced by higher concentrations of GABA (>30 microM), and accelerated the desensitization rate of I(GABA). BPA also greatly inhibited the steady state I(GABA) induced by higher concentrations of GABA (>30 microM) in a noncompetitive manner. In addition, BPA affected synaptic GABA(A) receptors as it decreased the amplitude of GABAergic miniature inhibitory postsynaptic currents in a concentration-dependent manner. Considering its complex modulatory effects on GABA(A) receptors, BPA might have potential toxicological effects on the central nervous system.

Pregnenolone sulfate enhances spontaneous glutamate release by inducing presynaptic Ca2+-induced Ca2+ release

Pregnenolone sulfate (PS) acts as an excitatory neuromodulator and has a variety of neuropharmacological actions, such as memory enhancement and convulsant effects. In the present study, we investigated the effect of PS on glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs) in acutely isolated dentate gyrus (DG) hilar neurons by use of a conventional whole-cell patch-clamp technique. PS significantly increased sEPSC frequency in a concentration-dependent manner without affecting the current amplitude, suggesting that PS acts presynaptically to increase the probability of spontaneous glutamate release. However, known molecular targets of PS, such as α7 nicotinic ACh, NMDA, σ1 receptors and voltage-dependent Ca(2+) channels, were not responsible for the PS-induced increase in sEPSC frequency. In contrast, the PS-induced increase in sEPSC frequency was completely occluded in a Ca(2+)-free external solution, and was significantly reduced by either the depletion of presynaptic Ca(2+) stores or the blockade of ryanodine receptors, suggesting that PS elicits Ca(2+)-induced Ca(2+) release (CICR) within glutamatergic nerve terminals. In addition, the PS-induced increase in sEPSC frequency was completely occluded by transient receptor potential (TRP) channel blockers. These data suggest that PS increases spontaneous glutamate release onto acutely isolated hilar neurons via presynaptic CICR, which was triggered by the influx of Ca(2+) through presynaptic TRP channels. The PS-induced modulation of excitatory transmission onto hilar neurons could have a broad impact on the excitability of hilar neurons and affect the pathophysiological functions mediated by the hippocampus.

GABAB receptor-mediated presynaptic inhibition of glycinergic transmission onto substantia gelatinosa neurons in the rat spinal cord

&NA; The GABAB receptor‐mediated presynaptic inhibition of glycinergic transmission was studied from young rat substantia gelatinosa (SG) neurons using a conventional whole‐cell patch clamp technique. Action potential‐dependent glycinergic inhibitory postsynaptic currents (IPSCs) were recorded from SG neurons in the presence of 3 mM kynurenic acid and 10 μM SR95531. In these conditions, baclofen (30 μM), a selective GABAB receptor agonist, greatly reduced the amplitude of glycinergic IPSCs and increased the paired‐pulse ratio. Such effects were completely blocked by 3 μM CGP55845, a selective GABAB receptor antagonist, indicating that the activation of presynaptic GABAB receptors decreases glycinergic synaptic transmission. Glycinergic IPSCs were largely dependent on Ca2+ influxes passing through presynaptic N‐ and P/Q‐type Ca2+ channels, and these channels contributed equally to the baclofen‐induced inhibition of glycinergic IPSCs. However, the baclofen‐induced inhibition of glycinergic IPSCs was not affected by either 100 μM SQ22536, an adenylyl cyclase inhibitor, or 1 mM Ba2+, a G‐protein coupled inwardly rectifying K+ channel blocker. During the train stimulation (10 pulses at 20 Hz), which caused a marked synaptic depression of glycinergic IPSCs, baclofen at a 30 μM concentration completely blocked glycinergic synaptic depression, but at a 3 μM concentration it largely preserved glycinergic synaptic depression. Such GABAB receptor‐mediated dynamic changes in short‐term synaptic plasticity of glycinergic transmission onto SG neurons might contribute to the central processing of sensory signals.

Serotoninergic modulation of GABAergic synaptic transmission in developing rat CA3 pyramidal neurons

Serotoninergic modulation of GABAergic mIPSCs was investigated in immature (postnatal 12–16‐days old) rat CA3 pyramidal neurons using a conventional whole‐cell patch clamp technique. Serotonin or 5‐hydroxytryptamine (5‐HT) (10 μmol/L) transiently and explosively increased mIPSC frequency with a small increase in the current amplitude. However, 5‐HT did not affect the GABA‐induced postsynaptic currents, indicating that 5‐HT acts presynaptically to facilitate the probability of spontaneous GABA release. The 5‐HT action on GABAergic mIPSC frequency was completely blocked by 100 nmol/L MDL72222, a selective 5‐HT3 receptor antagonist, and mimicked by mCPBG, a selective 5‐HT3 receptor agonist. The 5‐HT action on GABAergic mIPSC frequency was completely occluded either in the presence of 200 μmol/L Cd2+ or in the Na+‐free external solution, suggesting that the 5‐HT3 receptor‐mediated facilitation of mIPSC frequency requires a Ca2+influx passing through voltage‐dependent Ca2+channels from the extracellular space, and that presynaptic 5‐HT3 receptors are less permeable to Ca2+. The 5‐HT action on mIPSC frequency in the absence or presence of extracellular Na+ gradually increased with postnatal development. Such a developmental change in the 5‐HT3 receptor‐mediated facilitation of GABAergic transmission would play important roles in the regulation of excitability as well as development in CA3 pyramidal neurons.

Multiple effects of allopregnanolone on GABAergic responses in single hippocampal CA3 pyramidal neurons.

3α-Hydroxy, 5α-reduced pregnane steroids, such as allopregnanolone, are potent modulators of GABA(A) receptors and have many biological responses including sedative, anxiolytic, anticonvulsant and anesthetic actions. In the present study, we have investigated the effects of allopregnanolone on GABA(A) receptors in acutely isolated single hippocampal CA3 pyramidal neurons using the whole cell patch-clamp technique. Allopregnanolone induced membrane Cl(-) currents in a concentration-dependent manner, and the allopregnanolone-induced currents (I(AlloP)) were blocked by noncompetitive GABA(A) receptor antagonists. The I(AlloP) was not affected by the intracellular loading of γ-cyclodextrin (γ-CD), which efficiently sequesters several kinds of endogenous neurosteroids including allopregnanolone, suggesting that allopregnanolone accesses extracellular but not intracellular sites to activate GABA(A) receptors. Allopregnanolone prolonged the decay time constant of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs), suggesting that allopregnanolone modulates the desensitization kinetics of postsynaptic GABA(A) receptors. The picrotoxin-sensitive tonic currents (I(tonic)), which were mediated by extrasynaptic GABA(A) receptors, were recorded from CA3 pyramidal neurons. The intracellular loading of γ-CD or allopregnanolone significantly decreased or increased the amplitude of picrotoxin-sensitive I(tonic), respectively, suggesting that endogenous neurosteroids might, at least in part, be involved in the generation of picrotoxin-sensitive I(tonic). Allopregnanolone also increased the frequency of GABAergic sIPSCs, in a manner dependent on the integrity of voltage-dependent Na(+) and Ca(2+) channels, suggesting that allopregnanolone activates presynaptic GABA(A) receptors to depolarize GABAergic nerve terminals. The present results suggest that allopregnanolone exerts its pharmacological and pathophysiological actions via the modulation of multiple types of GABA(A) receptor-mediated responses.

Compound K, a metabolite of ginsenosides, facilitates spontaneous GABA release onto CA3 pyramidal neurons.

J. Neurochem. (2010) 114, 1085–1096.