Evidence for nitric oxide mediated effects on islet hormone secretory phospholipase C signal transduction mechanisms.
We have investigated the putative role of nitric oxide (NO) as a modular of islet hormone release, when stimulated by the muscarinic receptor agonist phospholipase C activator, carbachol, with special regard to whether the IP3-Ca2+ or the diacylglycerol-protein kinase C messenger systems might be involved. It was observed that the NO synthase (NOS) inhibitor N(G)-nitro-L-arginine methylester (L-NAME) markedly potentiated insulin release and modestly inhibited glucagon release induced by carbachol. Similarly, insulin release induced by the phorbol ester TPA (protein kinase C activator) was markedly potentiated. Glucagon release, however, was unaffected. Dynamic perifusion experiments with 45C2+ -loaded islets revealed that the inhibitory action of L-NAME on carbachol-stimulated NO-production was reflected in a rapid and sustained increase in insulin secretion above carbachol controls, whereas the 45Ca2+ -efflux pattern was similar in both groups with the exception of a slight elevation of 45C2+ in the L-NAME-carbachol group during the latter part of the perifusion. No difference in either insulin release or 45Ca2+ -efflux pattern between the carbachol group and L-NAME-carbachol group was seen in another series of experiments with identical design but performed in the absence of extracellular Ca2+. However, it should be noted that in the absence of extracellular Ca2+ both 45Ca2+ -efflux and, especially, insulin release were greatly reduced in comparison with experiments in normal Ca2+. Further, in the presence of diazoxide, a potent K+ ATP-channel opener, plus a depolarizing concentration of K+ the NOS-inhibitor L-NAME still markedly potentiated carbachol-induced insulin release and inhibited glucagon release. The enantiomer D-NAME, which is devoid of NOS-inhibitory properties, did not affect carbachol-induced hormone release. TPA-induced hormone release in depolarized islets was not affected by either L-NAME or D-NAME. The pharmacological intracellular NO donor hydroxylamine dose-dependently inhibited insulin release stimulated by TPA. Furthermore, a series of perifusion experiments revealed that hydroxylamine greatly inhibited carbachol-induced insulin release without affecting the 45Ca2+ -efflux pattern. In summary, our results suggest that the inhibitory effect of NO on carbachol-induced insulin release is not to any significant extent exerted on the IP3-Ca2+ messenger system but rather through S-nitrosylation of critical thiol-residues in protein kinase C and/or other secretion-regulatory thiol groups. In contrast, the stimulating action of NO on carbachol-induced glucagon release was, at least partially, connected to the IP3-Ca2+ messenger system. The main effects of NO on both insulin and glucagon release induced by carbachol were apparently exerted independently of membrane depolarization events.
Akesson
B
B
Department of Pharmacology, University of Lund, Sweden.
Lundquist
I
I
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism*; D018679:Cholinergic Agonists / Q000494:pharmacology; D003981:Diazoxide / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005934:Glucagon / Q000378:metabolism*; D019811:Hydroxylamine / Q000494:pharmacology; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000378:metabolism*; D051379:Mice; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000494:pharmacology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D015221:Potassium Channels / Q000187:drug effects; D011976:Receptors, Muscarinic / Q000378:metabolism*; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*
D018679:Cholinergic Agonists; D004791:Enzyme Inhibitors; D007328:Insulin; D015221:Potassium Channels; D011976:Receptors, Muscarinic; D019811:Hydroxylamine; D009569:Nitric Oxide; D002217:Carbachol; D005934:Glucagon; D019001:Nitric Oxide Synthase; D010738:Type C Phospholipases; D013755:Tetradecanoylphorbol Acetate; D003981:Diazoxide; D002118:Calcium; D019331:NG-Nitroarginine Methyl Ester