Muscarinic stimulation of synaptic activity by protein kinase C is inhibited by adenosine in cultured hippocampal neurons.
We have studied the effect of the cholinergic agonist carbachol on the spontaneous release of glutamate in cultured rat hippocampal cells. Spontaneous excitatory postsynaptic currents (sEPSCs) through glutamatergic alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type channels were recorded by means of the patch-clamp technique. Carbachol increased the frequency of sEPSCs in a concentration-dependent manner. The kinetic properties of the sEPSCs and the amplitude distribution histograms were not affected by carbachol, arguing for a presynaptic site of action. This was confirmed by measuring the turnover of the synaptic vesicular pool by means of the fluorescent dye FM 1-43. The carbachol-induced increase in sEPSC frequency was not mimicked by nicotine, but could be blocked by atropine or by pirenzepine, a muscarinic cholinergic receptor subtype M1 antagonist. Intracellular Ca2+ signals recorded with the fluorescent probe Fluo-3 indicated that carbachol transiently increased intracellular Ca2+ concentration. Since, however, carbachol still enhanced the sEPSC frequency in bis(2-aminophenoxy)ethane-N,N,N',N'-tetra-acetate-loaded cells, this effect could not be attributed to the rise in intracellular Ca2+ concentration. On the other hand, the protein kinase inhibitor staurosporine as well as a down-regulation of protein kinase C by prolonged treatment of the cells with 4beta-phorbol 12-myristate 13-acetate inhibited the carbachol effect. This argues for an involvement of protein kinase C in presynaptic regulation of spontaneous glutamate release. Adenosine, which inhibits synaptic transmission, suppressed the carbachol-induced stimulation of sEPSCs by a G protein-dependent mechanism activated by presynaptic A1-receptors.
Bouron
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A
Department of Pharmacology, University of Berne, Friedbuehlstrasse 49, CH-3010 Berne, Switzerland.
Reuter
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H
D000241:Adenosine / Q000494:pharmacology*; D000818:Animals; D000831:Animals, Newborn; D001285:Atropine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity; D004789:Enzyme Activation; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects; D006624:Hippocampus / Q000166:cytology / Q000502:physiology*; D018721:Muscarinic Agonists / Q000494:pharmacology; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000502:physiology; D018733:Nicotinic Antagonists / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018091:Receptors, AMPA / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000378:metabolism*; D019311:Staurosporine / Q000494:pharmacology; D013569:Synapses / Q000187:drug effects; D009435:Synaptic Transmission / Q000187:drug effects*
D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D018733:Nicotinic Antagonists; D018091:Receptors, AMPA; D011976:Receptors, Muscarinic; D001285:Atropine; D002217:Carbachol; D011493:Protein Kinase C; D019311:Staurosporine; D000241:Adenosine; D002118:Calcium