TIME-DEPENDENT EFFECTS OF CHOLINERGIC STIMULATION ON BETA-CELL RESPONSIVENESS

The effects of cholinergic stimulation on beta cell insulin secretory and phosphoinositide (PI) responses were determined in freshly isolate d rat islets. Increasing the glucose level perifusing the islet from 5 .6 to 8 mM was accompanied by a modest insulin secretory response. The further addition of 10 mu M carbachol increased peak first- and secon d-phase responses by 2.6- and 6.8-fold, respectively. In the presence of 5.6 mM glucose, this low level (10 mu M) of carbachol increased ins ulin release two- to three-fold, a response that was maintained for at least 60 min. In contrast to these acute stimulatory actions in the p resence of glucose, chronic 3.5-h exposure of islets to 10 mu M carbac hol abolished beta cell insulin secretory responses to stimulation, wi th the combination of 8 mM glucose plus 10 mu M carbachol. However, th e further addition of 200 mu M tolbutamide to these islets increased i nsulin secretory rates significantly. To establish the role of islet c ell PI hydrolysis in these secretory responses, additional studies wer e conducted with islets whose PI pools were labeled with [H-3]inositol . Acute exposure to 10 mu M carbachol alone significantly increased in ositol phosphate accumulation and the efflux of [H-3]inositol, even in the absence of glucose. Including 10 mu M carbachol during the labeli ng period with [H-3]inositol resulted in significant impairments in su bsequently measured inositol phosphate accumulation and [H-3]inositol efflux responses to 8 mM glucose plus carbachol stimulation. Prior lon g-term exposure to 10 mu M carbachol also induced heterologous desensi tization: 20 mM glucose-stimulated insulin release and inositol phosph ate accumulation were impaired in a parallel fashion. Chronic carbacho l exposure had no deleterious effect on the usage of 8 or 20 mM glucos e or on the insulin content of the islet. The acute stimulatory effect s of carbachol on inositol phosphate accumulation as well as its inhib itory effect on 20 mM glucose-stimulated insulin release after prolong ed exposure to the muscarinic agonist were significantly reduced by at ropine. These findings demonstrate that changes in PI hydrolysis paral lel those observed with insulin secretion and suggest that alterations in phospholipase C activation may account, at least in part, for the insulin secretory responses observed.