DYNAMIC INSULIN-SECRETION FROM PERIFUSED RAT PANCREATIC-ISLETS

Insulin secretion from isolated pancreatic islets of 8- to 12-day-old rats was investigated in a dynamic in vitro (perifusion) system. The a ims of the study were (i) to describe a carefully controlled in vitro method to study the mechanism of insulin secretion and to analyse the effects and dynamic interactions of bioactive compounds on isolated ra t pancreatic islets, (ii) to validate the method by comparing fundamen tal data on the functions of the islets obtained with this method to t hose collected with other techniques; and (iii) to find novel features of the control of insulin secretion. The method was carefully designe d to maintain the functional capacity of the explanted cells. A functi onal standardization system was elaborated consisting of (i) analysis of the changes in the basal hormone secretion of the cells; (ii) evalu ating responses to a standard, specific stimuli (50 mM glucose for 3 m in); (iii) determining the alteration of the momentary size of the hor mone pool with responses to KCl; and (iv) direct determination of the total intracellular hormone content from the extract of the column. Th e technique provides accurate quantitative data on the dynamic respons es to biologically active compounds that act directly on the pancreati c islets. The islets maintained their full responsiveness for up to 7 days, and responses as close as in 1-min intervals could be distinguis hed. A linear dose-response relationship was found on the glucose-indu ced insulin release in case of 3-min stimulation with 4 and 500 mM of glucose (lin-log graph). Utilizing this method, we showed that no dese nsitization to glucose-induced insulin release can be observed if the responsiveness of the cells is properly maintained and the parameters of the stimulation are carefully designed. Exposure of the explanted i slets to 10 mu M acetylcholine or 30 mM arginine (Arg) induced a trans itory elevation of insulin release similar in shape to that experience d after glucose stimulation. Norepinephrine (NE), dopamine (DA) and so matostatin (SS) did not induce any detectable alteration on the basal insulin secretion of the islets. However, 100 nM SS given together wit h 50 mM glucose, 30 mM Arg or 10 mu M acetylcholine significantly redu ced the insulin-releasing effect of these substances (by 75.5, 71.5 an d 72.5%, respectively). At the same time; SS did not alter the insulin response of the islets to 100 mM elevation of K+ concentration. SS al so inhibited glucose-induced insulin release in a dose-dependent way ( ED50 = 22 nM). A similar dose-dependent inhibitory effect on glucose i nduced insulin release was found with NE (ED50 = 89 nM) and DA (ED50 = 2.2 mu M). gamma-Aminobutyric acid (GABA) did not influence insulin r elease under similar circumstances.