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FOUR DIFFERENT KINDS OF CHEMICALLY-OPERATED ION CHANNELS IN SYMPATHETIC NEURONS. TOSHIO IIJIMA AND GEN MATSUMOTO, Electrotechnical Laboratory, 1-1-4 Umezono, Tsu-~a-C1---f-~,Ib--a~-akl~05, Japan.

We studied the properties of ion channels involved in the musearinic acety- icholine (ACh) response. Membrane current and intracellular calcium concentration ([Ca2+]i) were simultaneously recorded on cultured superior cervical ganglion cells from rabbit. ACh, OAG (a synthetic diacylglycerol) and phorbol ester (a protein kinase C activator) caused inactivation of M channel. The inactivation was not affected by [Ca2+]i, but was by dramatically enhanced and prolonged intracellular injection of protein kinase C. When the membrane was depolarized to 0 mV (from -60 mV), the [Ca2+]i increased due to activation of voltage-dependen~ Ca 2+ channels. Under these conditions, ACh activated calcium-dependent C1 channels. Membrane depolarization to +20 mV caused a larger [Ca2+]i increase. However, this elevation was significantly suppressed in the presence of ACh. Therefore ACh blocks voltage dependent actvation of L-type Ca 2+ channel.

TWO TYPES OF GLYCINE RECEPTORS EXPRESSED IN XENOPUS OOCYTES INJECTED WITH RAT SPINAL CORD mRNA. HIROYUKI AKAGI, AND RICARDO MILEDI*, Department of Psychobioloqy, University of California r Irvine t CA 92717 t USA.

Poly(A)'-messenger RNAs (mRNAs) isolated from adult and neonatal (3 to 4 days old) rat spinal cord were injected into Xenopus oocytes, which were subsequently examined under voltage clamp. Messenger RNAs obtained from both sources induced the oocytes to acquire functional glycine receptors (GIyR). The glycine-induced currents reversed direction at about -20 mV (close to the equilibrium potential of CI- in the oocytes) and the responses were blocked by strychnine (0.5 ~M) or picrotoxin (20 ~M). Thus, the GlyRs encoded by two kinds of mRNAs share some characteristics. Nevertheless, size fractionation of the GIyR-mRNAs by sucrose density sedimentation revealed contrasting profiles; the majority of the adult cord GIyR-mRNA sedimented in heavy density fractions, close to the position of the 28 S RNA, while the neonatal cord GIyR-mRNA was mainly close to that of the 18 S RNA. The properties of GIyR encoded by adult cord mRNA differed from those of the receptor encoded by neonatal cord mRNA with respect to dose-response relationship, time course of desensitization and some pharmacological actions. These results suggest that in rat spinal cord there exist at least two classes of mRNAs which encode distinct GIyRs. The production of the GIyR-mRNAs in spinal cord appears to be developmentally regulated.

INWARD RECTIFICATION OF THE ACETYLCHOLINE RECEPTOR CURRENT IN MAMMALIAN AUTONOMIC GANGLIA. HIROMU YAWO% Department of Physiology 7 Kyoto University Faculty of Medicine~ Kyoto 606~ Japan.

The nicotinic acetyleholine receptor (AChR) currents recorded from mammalian autonomic ganglion cells are characterized by a non-linear current-voltage (I-V) relation, in contrast to those recorded from skeletal muscle. The mechanism underlying this unique behavior was investigated by tight-seal whole-cell recording. Mouse submandibular ganglion (SMG) cells are spherical in shape and lack dendrites, so that these cells provide favorable conditions for voltage-clamping. Adult mice were anesthetized with pentobarbital (250 mg/kg), and SMGs were removed with attached presynaptic nerves.

Both the AChR currents and the excitatory synaptic currents (EPSC) were strikingly reduced in magnitude during depolarization. The outward current was almost zero at about 50 mV, followed by a sharp increase with further depolarization. This behavior was independent of the type of cations inside (Na, K, or Cs). Single channel properties were investigated by analysis of whole-cell current noise, the power spectrum of which was well fitted with 2-Lorentzian components. The I-V relation of single AChR channel currents was asymmetrical, being ohmic in negative membrane potentials (inward currents) with a slope of 42 pS and tending to saturate in positive potentials (outward currents). Furthermore, the voltage-dependence of the noise time constants implies that the apparent mean open time of the channel was significantly shortened by depolarization. Thus, the probability of the AChR channel being open appears to change with membrane potential.