Tsuneo Tosaka

Presynaptic inhibition by noradrenaline of the EPSC evoked in neonatal rat sympathetic preganglionic neurons

Visually identified and electrophysiologically characterized sympathetic preganglionic neurons (SPNs) were recorded using the whole-cell voltage clamp technique in slices of neonatal rat spinal cord. Monosynaptic excitatory postsynaptic currents (EPSCs) were evoked by electrical stimulation of the nucleus intercalatus in the presence of strychnine (5 microM) and bicuculline (10 microM). These EPSCs were abolished by the antagonist of AMPA-type glutamate receptors, 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX; 10 microM). Bath applied noradrenaline (NA; 0.5-50 microM) dose-dependently and reversibly decreased by up to around 60% the amplitude of the EPSC, without affecting the holding current. The EPSC depression by NA was not accompanied by a change in EPSC reversal potential (around +5 mV), nor were inward currents generated by pressure application of glutamate affected by NA application. A comparable degree of EPSC depression was also seen with the alpha2-adrenoceptor agonist clonidine (5 microM), and the alpha2A-agonist oxymetazoline (5 microM), while the alpha1-agonist phenylephrine (100 microM) caused only a 22% depression. The EPSC depression caused by NA (10 microM) was completely antagonized by either the alpha-antagonist phentolamine (10 microM) or the alpha2-antagonist idazoxan (2 microM). Conversely, the beta-adrenoceptor antagonist popranolol (5 microM), and the alpha1-, alpha2B- and alpha2C-antagonist prazosin (2 microM) were without effect. These results indicate that activation of presynaptic alpha2A-adrenoceptors on inputs to SPNs decreases glutamate release.

Voltage-dependent potassium currents of sympathetic preganglionic neurons in neonatal rat spinal cord thin slices.

Voltage-dependent potassium currents were analyzed in the visually identified sympathetic preganglionic neurons (SPNs) of neonatal rat spinal cord thin slices by the whole-cell patch-clamp technique. Some of the SPNs were identified by the presence of retrogradely transported fluorescent dye, DiI, injected into the superior cervical ganglion several days prior to experimentation. In a tetrodotoxin (TTX)-containing solution, a step depolarization from the holding potential of -72 mV generated a slow outward current that was suppressed by tetraethylammonium (TEA) and by Ca(2+)-free/2.5 ImM Co2+ solution. Ca(2+)-dependent current consisted of a transient and a sustained components. In a Ca(2+)-free (substituted with Mg2+) solution with TTX and TEA, a step depolarization from a hyperpolarized potential evoked a transient outward current that was blocked by 4-aminopyridine (4-AP). A step hyperpolarization evoked a voltage-dependent inward current, the conductance of which was dependent not only on the membrane potential, but also on the extracellular K+ concentration. Tail current analyses revealed that all of these currents were carried by K+ ions. These results indicate that SPN possesses at least five types of voltage-dependent K+ current, including the delayed rectifier current (IK), Ca(2+)-dependent transient current (IC), Ca(2+)-dependent sustained current (IAHP), A-current (IA) and inward rectifying current (Iu), which may be targets of putative transmitters released from various descending and segmental inputs impinging upon the SPN.

Different sensitivities of acetylcholine-induced “after-hyperpolarization” compared to dopamine-induced hyperpolarization, to ouabain or to lithium-replacement of sodium, in rabbit sympathetic ganglia

Abstract The ganglionic potentials elicited by a test dose of acetylcholine (ACh) acting nicotinically consist of an initial depolarization followed by a longer “after-hyperpolarization” (after-HP). The after-HP is abolished selectively and relatively rapidly after treatment with ouabain or with Li-replacement of 90% of Na in Ringer. The hyperpolarizing response to a test dose of dopamine (DA), however, is as or more resistant than the ACh-depolarization to either treatment. These findings provide further strong evidence that the slow inhibitory postsynaptic potential (s-IPSP), for which DA is the transmitter in these ganglia, is not mediated by an activation of an electrogenic NaK pump.

Hyperpolarization following activation of K+ channels by excitatory postsynaptic potentials.

We have postulated that an excitatory postsynaptic potential (e.p.s.p.) may open voltage-sensitive K+ (‘M’) channels1, in an appropriate depolarizing range, and that this could alter the e.p.s.p. waveform. Consequently, the fast e.p.s.p. in neurones of sympathetic ganglia, elicited by a nicotinic action of acetylcholine (ACh)2, could be followed by a hyperpolarization, produced by the opening of M channels during the depolarizing e.p.s.p. and their subsequent slow closure (time constant∼150 ms)1. This introduces the concept that transmitter-induced p.s.ps may trigger voltage-sensitive conductances other than those initiating action potentials, and that in the present case this could produce a true post-e.p.s.p. hyperpolarization. (Some hyperpolarizations other than inhibitory postsynaptic potentials (i.p.s.ps) have been reported to follow e.p.s.ps3,4.) We show here that this is so.

Glial uptake system of GABA distinct from that of taurine in the bullfrog sympathetic ganglia

The kinetics and specificity of GABA and taurine uptake were studied in the bullfrog sympathetic ganglia. GABA uptake system consisted of simple saturable component and taurine uptake system consisted of two saturable components exclusive of non-saturable influx. Taurine unaffected GABA uptake while GABA inhibited taurine uptake competitively with theKi/Km ratio of 38. GABA (5.14 μM) uptake was inhibited by δ-aminovaleric acid and slightly by 2,4-diaminobutyric acid (5 mM, each) among ten structural analogs. Taurine uptake under high-affinity conditions was most strongly suppressed by hypotaurine and β-alanine competitively with theKi/Km ratio of 1.0 and 1.9, respectively. Autoradiography showed that glial cells were heavily labeled by both [3H]GABA and [3H]taurine. These results suggest that GABA is transported by a highly specific carrier system distinct from the taurine carrier and that taurine, hypotaurine, and β-alanine may share the same high-affinity carrier system in the glial cells of the bullfrog sympathetic ganglia.

Sodium dependency of GABA uptake into glial cells in bullfrog sympathetic ganglia.

The kinetics of sodium dependency of GABA uptake by satellite glial cells was studied in bullfrog sympathetic ganglia. GABA uptake followed simple Michaelis-Menten kinetics at all sodium concentrations tested. Increasing external sodium concentration increased bothKm andVmax for GABA uptake, with an increase in theVmax/Km ratio. The initial rate of uptake as a function of the sodium concentration exhibited sigmoid shape at 100 μM GABA. Hill number was estimated to be 2.0. Removal of external potassium ion or 10 μM ouabain reduced GABA uptake time-dependently. The effect of ouabain was potentiated by 100 μM veratrine. These results suggest that at least two sodium ions are involved with the transport of one GABA molecule and that sodium concentration gradient across the plasma membrane is the main driving force for the transport of GABA. The essential sodium gradient may be maintained by Na+, K+-ATPase acting as an ion pump.

ATP receptors in the smooth muscle of the guinea-pig vas deferens

554 ABNORMAL NEURAL INPUTS FROM THE MEDIAN PREOPTIC NUCLEUS TO THE HYPOTHALAMIC PARAVENTJUCULAR NUCLEUS IN SPONTANEOUSLY HYPERTENSIVE RATS JUNICHI TANAKA’, YASUSHI HAYASHI 2, KATSUHIDE KARIYA3, MASAHIKO NOMURA4 Depts. of ‘Human Dev. and 2Educ. for Handicapped Child., Naruto Univ. of Educ., Naruto, Tokushima 772-8502, 3Res. Lab., Torii & Co. Ltd., Midori-ku, Chiba 267-0056, 4Dept. of Physiol., Saitama Med. Sch., lruma-gun, Saitama, 350-0495 Extracellular single-unit activity was recorded from phasically firmg neurohypophyseal neurons tn the hypothalamic paraventricularnucleus (PVN) of male Wistar-Kyoto (WKY, 40 units) and spontaneously hypertensive rats (SHR, 38 units) under urethane anesthesia. Electrical stimulation of the median preoptic nucleus (MnPO) J)roduced orthodromic excitatton (48% in WKY rats; 51% in SHR) or inhibition (10% in WKY rats; 13% in SHR) of the activity of PVN units. No signifmant differences in the latency, duration or threshold of the MnPG stimulus-induced responses were observed between WKY and SHR. The magnitude of excitatory response, but not the inhtbitory response, much greater in SHR than m WKY rats. Local administration of angiotensin II (ANG II) into the stimulation sites enhanced the neuronal activity of units (64% in WKY rats; 67% in SHR) that displayed the excitation to electrical stimulation of the MnPO. The duration and frequency of excitatory response caused by the ANG II injection was much greater in SHR than in WKY rats. These results suggest that ANG IIsensitive MnPO efferents to the PVN act to facilitate the excitability of putative vasopressin-secreting neurons in the PVN and imply that there is the alteration between WKY and SHR in the functron of the pathways.

2424 Changes in the smooth muscle of the guinea-pig vas deferens by thermal stress

SAEKO SAKAI’ , ISA0 YOSHIHAMA3. FCSIIHIKO HOKOISHI’. KXZUNORI NAMIKI’. 5IASASHI OGAWA2, MAKOTO MIK12, TSUNEO TOSAKA’ The thermal effects on contraction, binding kinetics of [3H]prazosin and histology were examined in the guineapig vas deferens. The contraction was recorded at 36°C after thermal exposure to 43-47°C for 1 h. The binding kinetics of cri-adrenoceptors was determined at 43-55°C. No significant changes in the contraction and the kinetic parameters were observed at 43°C. The contractions induced by phenylephrine. .ATP. ?L1Ch, nicotine. KC1 and electrical stimulation were abolished at 47°C. whereas the number of binding sites reduced by 54% at 55°C. Serious damage of the muscle fibers was observed at 50°C. Autoradiography of tritiated or. a2 and P-antagonists revealed that the muscle membrane was predominantly labeled with silver grains. The results indicate that the nerve fibers and the contractile elements were intact at 43”C, and that the muscle fibers lost their contractility at 47°C; although al-adrenoceptors tolerated against much higher thermal stress.