Voltage- and time-dependent inhibition of neuronal calcium channels by a GTP-binding protein in a mammalian cell line.
1. Inhibitory modualtion of the three Ca2+ channel current components by neurotransmitters was studied using the whole-cell patch-clamp method in a mammalian cell line, NG108-15. 2. In cells differentiated with dibutyryl cyclic AMP, both the low-voltage-activated current (ILVA) and omega CgTX-sensitive high-voltage-activated current (I omega CgTX) could be inhibited by [D-phen2, D-phen5]enkephalin, acetylcholine and noradrenaline. In contrast, differentiation with prostaglandin E1 and theophylline eliminated the agonist-induced inhibition of ILVA, but enhanced that of I omega CgTX. The DHP-sensitive high-voltage-activated current was unaffected by the transmitters in most of the cells. 3. The inhibition was prevented by pre-treatment of cells with pertussis toxin, suggesting involvement of a G-protein. Long treatment of the cells with phorbol ester did not prevent the inhibition. 4. The inhibition was always partial: the maximal inhibition was 40% for ILVA and 70% for I omega CgTX. 5. The inhibition of ILVA and I omega CgTX was relieved during depolarization. Half-maximal relief of inhibition of I omega CgTX was attained at 0 mV, irrespective of agonist concentration. 6. The kinetics of removal and re-establishment of inhibition were voltage dependent. Both processes were single exponentials and had identical time constants at a given membrane potential. Time constants were 124 ms at -40 mV, 160 ms at 0 mV and 8 ms at 60 mV, at any agonist concentration. 7. Time courses of tail currents were unaltered by the inhibition. 8. The inhibition of the omega CgTX-sensitive Ca2+ channel can be described as a shift in gating modes; with an additional voltage-dependent gating state activated by the agonists. The voltage-dependent properties of this modulation allow inhibition of Ca2+ channel to be overcome by high-frequency trains of action potentials.
Abetilung Membranbiophysik, Max Planck Institut für biophysikalische Chemie, Göttingen, Germany.
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Kasai
D000109:Acetylcholine / Q000494:pharmacology; D000527:Alprostadil / Q000378:metabolism; D000818:Animals; D016718:Arachidonic Acid / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000502:physiology*; D008978:Mollusk Venoms / Q000494:pharmacology; D018377:Neurotransmitter Agents / Q000494:pharmacology*; D009638:Norepinephrine / Q000494:pharmacology; D013806:Theophylline / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured / Q000502:physiology; D020866:omega-Conotoxin GVIA
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004745:Enkephalins; D008978:Mollusk Venoms; D018377:Neurotransmitter Agents; D016718:Arachidonic Acid; D020881:Enkephalin, D-Penicillamine (2,5)-; D020866:omega-Conotoxin GVIA; D013806:Theophylline; D019204:GTP-Binding Proteins; D000527:Alprostadil; D000109:Acetylcholine; D009638:Norepinephrine