Xenon modulates synaptic transmission to rat hippocampal CA3 neurons at both pre- and postsynaptic sites.

KEY POINTS: Xenon (Xe) non-competitively inhibited whole-cell excitatory glutamatergic current (IGlu ) and whole-cell currents gated by ionotropic glutamate receptors (IAMPA , IKA , INMDA ), but had no effect on inhibitory GABAergic whole-cell current (IGABA ). Xe decreased only the frequency of glutamatergic spontaneous and miniature excitatory postsynaptic currents and GABAergic spontaneous inhibitory postsynaptic currents without changing the amplitude or decay times of these synaptic responses. Xe decreased the amplitude of both the action potential-evoked excitatory and the action potential-evoked inhibitory postsynaptic currents (eEPSCs and eIPSCs, respectively) via a presynaptic inhibition in transmitter release. We conclude that the main site of action of Xe is presynaptic in both excitatory and inhibitory synapses, and that the Xe inhibition is much greater for eEPSCs than for eIPSCs. ABSTRACT: To clarify how xenon (Xe) modulates excitatory and inhibitory whole-cell and synaptic responses, we conducted an electrophysiological experiment using the 'synapse bouton preparation' dissociated mechanically from the rat hippocampal CA3 region. This technique can evaluate pure single- or multi-synapse responses and enabled us to accurately quantify how Xe influences pre- and postsynaptic aspects of synaptic transmission. Xe inhibited whole-cell glutamatergic current (IGlu ) and whole-cell currents gated by the three subtypes of glutamate receptor (IAMPA , IKA and INMDA ). Inhibition of these ionotropic currents occurred in a concentration-dependent, non-competitive and voltage-independent manner. Xe markedly depressed the slow steady current component of IAMPA almost without altering the fast phasic IAMPA component non-desensitized by cyclothiazide. It decreased current frequency without affecting the amplitude and current kinetics of glutamatergic spontaneous excitatory postsynaptic currents and miniature excitatory postsynaptic currents. It decreased the amplitude, increasing the failure rate (Rf) and paired-pulse rate (PPR) without altering the current kinetics of glutamatergic action potential-evoked excitatory postsynaptic currents. Thus, Xe has a clear presynaptic effect on excitatory synaptic transmission. Xe did not alter the GABA-induced whole-cell current (IGABA ). It decreased the frequency of GABAergic spontaneous inhibitory postsynaptic currents without changing the amplitude and current kinetics. It decreased the amplitude and increased the PPR and Rf of the GABAergic action potential-evoked inhibitory postsynaptic currents without altering the current kinetics. Thus, Xe acts exclusively at presynaptic sites at the GABAergic synapse. In conclusion, our data indicate that a presynaptic decrease of excitatory transmission is likely to be the major mechanism by which Xe induces anaesthesia, with little contribution of effects on GABAergic synapses.

Calcium channel subtypes on glutamatergic mossy fiber terminals synapsing onto rat hippocampal CA3 neurons.

The current electrophysiological study investigated the functional roles of high- and low-voltage-activated Ca2+ channel subtypes on glutamatergic small mossy fiber nerve terminals (SMFTs) that synapse onto rat hippocampal CA3 neurons. Experiments combining both the "synapse bouton" preparation and single-pulse focal stimulation technique were performed using the conventional whole cell patch configuration under voltage-clamp conditions. Nifedipine, at a high concentration, and BAY K 8644 inhibited and facilitated the glutamatergic excitatory postsynaptic currents (eEPSCs) that were evoked by 0.2-Hz stimulation, respectively. However, these drugs had no effects on spontaneous EPSCs (sEPSCs). Following the use of a high stimulation frequency of 3 Hz, however, nifedipine markedly inhibited eEPSCs at the low concentration of 0.3 µM. Moreover, ω-conotoxin GVIA and ω-agatoxin IVA significantly inhibited both sEPSCs and eEPSCs. Furthermore, SNX-482 slightly inhibited eEPSCs. R(-)-efonidipine had no effects on either sEPSCs or eEPSCs. It was concluded that glutamate release from SMFTs depends largely on Ca2+ entry through N- and P/Q-type Ca2+ channels and, to a lesser extent, on R-type Ca2+ channels. The contribution of L-type Ca2+ channels to eEPSCs was small at low-firing SMFTs but more significant at high-firing SMFTs. T-type Ca2+ channels did not appear to be involved in neurotransmission at SMFTs. NEW & NOTEWORTHY Action potential-evoked glutamate release from small mossy fiber nerve terminals (SMFTs) that synapse onto rat hippocampal CA3 neurons is regulated by high-threshold but not low-threshold Ca2+ channel subtypes. The functional contribution mainly depends on N- and P/Q-type Ca2+ channels and, to a lesser extent, on R-type Ca2+ channels. However, in SMFTs stimulated at a high 3-Hz frequency, L-type Ca2+ channels contributed significantly to the currents. The present results are consistent with previous findings from fluorometric studies of large mossy fiber boutons.

Effects of triphenyltin on glycinergic transmission on rat spinal neurons.

Glycine is a fast inhibitory transmitter like γ-aminobutyric acid in the mammalian spinal cord and brainstem, and it is involved in motor reflex, nociception, and neuronal development. Triphenyltin (TPT) is an organometallic compound causing environmental hazard to many wild creatures. Our previous findings show that TPT ultimately induces a drain and/or exhaustion of glutamate in excitatory presynaptic nerve terminals, resulted in blockage of neurotransmission as well as methylmercury. Therefore, we have investigated the neurotoxic mechanism how TPT modulates inhibitory glycinergic transmission in the synaptic bouton preparation of rat isolated spinal neurons using a patch clamp technique. TPT at environmentally relevant concentrations (3-300 nM) significantly increased the number of frequency of glycinergic spontaneous and miniature inhibitory postsynaptic currents (sIPSC and mIPSC) without affecting the current amplitude and decay time. The TPT effects were also observed in external Ca2+-free solution containing tetrodotoxin (TTX) but removed in Ca2+-free solution with both TTX and BAPTA-AM (Ca2+ chelator). On the other hand, the amplitude of glycinergic evoked inhibitory postsynaptic currents (eIPSCs) increased with decreasing failure rate (Rf) and paired pulse ratio (PPR) in the presence of 300 nM TPT. At a high concentration (1 µM), TPT completely blocked eIPSCs after a transient facilitation. Overall, these results suggest that TPT directly acts transmitter-releasing machinery in glycinergic nerve terminals. Effects of TPT on the nerve terminals releasing fast transmitters were greater in the order of glycinergic > glutamatergic > GABAergic ones. Thus, TPT is supposed to cause a strong synaptic modulations on glycinergic neurotransmission in wild creatures.

Xenon modulates the GABA and glutamate responses at genuine synaptic levels in rat spinal neurons.

Effects of xenon (Xe) on whole-cell currents induced by glutamate (Glu), its three ionotropic subtypes, and GABA, as well as on the fast synaptic glutamatergic and GABAergic transmissions, were studied in the mechanically dissociated "synapse bouton preparation" of rat spinal sacral dorsal commissural nucleus (SDCN) neurons. This technique evaluates pure single or multi-synapse responses from native functional nerve endings and enables us to quantify how Xe influences pre- and postsynaptic transmissions accurately. Effects of Xe on glutamate (Glu)-, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-, kainate (KA)- and N-methyl-d-aspartate (NMDA)- and GABAA receptor-mediated whole-cell currents were investigated by the conventional whole-cell patch configuration. Excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) were measured as spontaneous (s) and evoked (e) EPSCs and IPSCs. Evoked synaptic currents were elicited by paired-pulse focal electric stimulation. Xe decreased Glu, AMPA, KA, and NMDA receptor-mediated whole-cell currents but did not change GABAA receptor-mediated whole-cell currents. Xe decreased the frequency and amplitude but did not affect the 1/e decay time of the glutamatergic sEPSCs. Xe decreased the frequency without affecting the amplitude and 1/e decay time of GABAergic sIPSCs. Xe decreased the amplitude and increased the failure rate (Rf) and paired-pulse ratio (PPR) without altering the 1/e decay time of both eEPSC and eIPSC, suggesting that Xe acts on the presynaptic side of the synapse. The presynaptic inhibition was greater in eEPSCs than in eIPSCs. We conclude that Xe decreases glutamatergic and GABAergic spontaneous and evoked transmissions at the presynaptic level. The glutamatergic presynaptic responses are the main target of anesthesia-induced neuronal responses. In contrast, GABAergic responses minimally contribute to Xe anesthesia.

Effects of Z-338, a novel gastroprokinetic agent, on the actions of excitatory and inhibitory neurotransmitters on neurons in area postrema.

We investigated the effects of the novel gastroprokinetic agent Z-338 on the actions of excitatory and inhibitory neurotransmitters on neurons in area postrema (AP). Iontophoretic applications of acetylcholine (ACh), AMPA and NMDA increased, while GABA suppressed the firing rates of AP neurons recorded by extracellular electrodes. Z-338 (10 microM) suppressed the ACh-induced acceleratory and GABA-induced inhibitory actions without affecting the excitatory actions of AMPA and NMDA. Under voltage-clamp conditions, nicotine, NMDA, kainic acid (KA) and ATP evoked inward currents in dissociated single AP neurons recorded by whole-cell patch clamp technique, and GABA produced outward currents, at holding potentials (V(H)) of -60 or 0 mV. Z-338 (>3 microM) specifically suppressed the nicotine- and GABA-induced currents without affecting the currents induced by NMDA, KA and ATP. In addition, we found that Z-338 (30 microM) suppressed the spontaneous inhibitory postsynaptic currents (sIPSCs) recorded from AP neurons in slice preparations. Experiments with microelectrode and histochemical methods revealed the presence of direct excitatory and di-synaptic inhibitory neural connections from AP to dorsal motor nucleus of the vagus (DMV). In some AP neurons, Z-338 (10 microM) enhanced the spontaneous firing rates recorded by extracellular electrode. The excitatory or inhibitory effects of Z-338 on the firing rates or actions of nicotine and GABA on AP neurons observed in the present study may explain the postmeal relaxation induced by Z-338 in patients with functional dyspepsia.

Effects of scorpion toxin on excitatory and inhibitory presynaptic terminals.

The effects of scorpion toxin (STX) on both spontaneous and evoked glycinergic and glutamatergic postsynaptic currents were studied by using both the mechanically dissociated single SDCN neuron (synaptic bouton preparation) and the 'focal electrical stimulation technique'. In the experimental condition where Na(+) channels on postsynaptic soma membrane were blocked by intracellular perfusion of QX-314, STX increased dose-dependently the frequency of spontaneous glycinergic and glutamatergic postsynaptic currents (sIPSC and sEPSC, respectively) without affecting the amplitude, suggesting STX acts on inhibitory and excitatory presynaptic nerve terminal. Such a facilitatory effect of STX on sIPSC was stronger than that on sEPSC. On the other hand, STX significantly enhanced the averaged current amplitude and decreased the failure rate (Rf) of both evoked inhibitory and excitatory postsynaptic currents (eIPSC and eEPSC, respectively), indicating that STX increases not only the release frequency of glycine and glutamate but also the amount of their release from the both presynaptic nerve endings. These effects of STX were completely removed by adding Na(+) or Ca(2+) channel blockers, indicating that STX increases Ca(2+) influx through Ca(2+) channels triggered by activating voltage-dependent Na(+) channels on the nerve terminals. In addition, the difference of STX actions on the amplitude of spontaneous and evoked currents was discussed.

Histamine responses of large neostriatal interneurons in histamine H1 and H2 receptor knock-out mice.

Histamine (HA) is an important neuro-modulator, contributing to a variety of physiological responses in the mammalian central nervous system (CNS). However there is little information about the cell/signaling mechanism underlying its role. In the present study, we characterized HA responses in single large neostriatal neurons acutely dissociated from wild type (WT) and HA receptor knock-out (KO) mice, with a particular emphasis on identifying the role of HA receptor subtypes. HA (10 microM) and a selective H(2) receptor agonist dimaprit (1 microM) both evoked an inward current in H(1)-KO mice, and HA and a selective H(1) receptor agonist HTMT (10 microM) both evoked an inward current in H(2)-KO mice. In the H(1) and H(2) double (H(1/2)) KO mice, there was no response to either the application of HA or the selective H(1), H(2) receptor agonists. Hence we have confirmed that the targeted genes were indeed absent in these KO mice and that both receptor subtypes contribute to HA's excitatory actions. Furthermore the HA-induced inward currents were mediated by a decrease in current through K(+) channels. In addition, we observed the effects of methamphetamine (METH) on the locomotor activity of WT and HA receptor KO mice, and found that METH-induced behavioral sensitization is evident in H(1/2)-KO mice, but not in H(1)- or H(2)-KO mice. These observations suggest that suppressive roles of HA on methamphetamine-induced behavioral sensitization would be mediated through both H(1) and H(2) receptors in the CNS including neostriatum.