448 Bio¢~imica et Biophvsic~ Acta 886 (1986) 448-456 Elsevier BBA 11723 Na+-H + exchange in the process of glucose-induced insulin release from the pancreatic B-ceil. Effects of amiloride on 86 Rb, 45Ca fluxes and insulin release P. Lebrun, E. van Ganse, M. Juvent, M. Deleers and A. Herchuelz * Laboratory of Pharmacology, Brussels University School of Medicine, 115, Boulevard de Waterloo, B-1000 Brussels (Belgium) (Received August 28th, 1985) (Revised manuscript received January 24th, 1986) Key words: Amiloride; Na+-H + exchange; lnsulin release; (Pancreatic B-cell) The effect of amiloride, an inhibitor of Na+-H + exchange, on intraceilular pH (pHi), S~Rb outflow, 45Ca outflow and insulin release from pancreatic rat islets was examined. In the 0.1-1 mM range, amUoride transiently reduced pH i of glucose-deprived islets and allowed glucose to induce a sustained decrease in pH~ of the islet cells. Amiloride reproduced the effect of glucose to decrease S6Rb and 45Ca Outflow. In the presence of glucose (5.6 mM or more), amiloride (100 pM) acted synergistically with the sugar to reduce K + outflow, and to stimulate 4°Ca inflow and insulin release from perifused islets. These results add strong support to the view that the generation of protons through the metabolism of glucose represents an important step in the process of glucose-induced release. The stimulation by glucose of Na+-H + exchange apparently masks and even overcomes the glucose-induced decrease in pH i otherwise expected from the increase in catabolic fluxes. Introduction Since the demonstration that glucose provokes a dose-related increase in H + net output from pancreatic islets [1], it has been suggested that a decrease in intracellular pH (pHi) of the pan- creatic B-cell may represent an important step in the process of glucose-induced insulin release [2,3]. A current view is that protons generated by the metabolism of glucose may inhibit both Ca + and K + outflow from the B-cell, leading to membrane depolarization, gating of voltage-sensitive Ca 2+ channels, increase in cytosolic free Ca 2+ and hence * To whom correspondence should be addressed. Abbreviations: DIDS, 4,4'-diisothiocyanostilbene-2,2'-di- sulfonic acid; Hepes, 4-(2-hydroxyethyl)-l-piperazineethane- sulfonic acid; SITS, 4-acetamido-4'-isothiocyanostilbene-2,2'- disulfonic acid. stimulation of insulin release [2,3]. Protons may be extruded from the B-cell by Na+-H + exchange and/or by a process of HCO3/CI- exchange [3,4]. Although supported by several lines of indi- rect evidences, the validity of this hypothesis has been obscured by the finding that glucose induces a sustained increase, rather than a decrease in pH~ of pancreatic islet cells [4-6]. In addition, the use of experimental conditions known to decrease pH~ have so far failed to show unequivocally that a lowering in pH i may indeed reproduce the full sequence of events normally evoked by glucose, namely a reduction in both K + and Ca 2+ outflow with subsequent increase in Ca 2 + inflow and even- tual stimulation of insulin release. Recently, it was shown that amiloride, an in- hibitor of Na+-H + exchange [7] transiently de- creased pH i in glucose-deprived islets and allowed glucose to induce a sustained decrease in pH~ of 0167-4889/86/$03.50 © 1986 Elsevier Science Publishers B.V. (Biomedical Division)