EFFECT OF ATP, CARBACHOL AND OTHER AGONISTS ON INTRACELLULAR CALCIUM ACTIVITY AND MEMBRANE VOLTAGE OF PANCREATIC DUCTS

The pancreatic duct has been regarded as a typical cAMP-regulated epithelium, and our knowledge about its Ca2+ homeostasis is limited. Hence, we studied the regulation of intracellular calcium, [Ca2+](i), in perfused rat pancreatic ducts using the Ca2+-sensitive probe fura-2. In some experiments we also measured the basolateral membrane voltage, V-bl, of individual cells. The resting basal [Ca2+](i) was relatively high, corresponding to 263 +/- 28 nmol/l, and it decreased rapidly to 106 +/- 28 nmol/l after removal of Ca2+ from the bathing medium (n = 31). Carbachol increased [Ca2+](i) in a concentration-dependent manner. At 10 mu mol/l the fura-2. fluorescence ratio increased by 0.49 +/- 0.06 (n = 24), corresponding to an increase in [Ca2+](i) by 111 +/- 15 nmol/l (it = 17). ATP, added to the basolateral side at 0.1 mmol/l and 1 mmol/l, increased the fluorescence ratio by 0.67 +/- 0.06 and 1.01 +/- 14 (ii = 46; 12), corresponding to a [Ca2+](i) increase of 136 +/- 22 nmol/l and 294 +/- 73 nmol/l respectively (n = 15; 10), Microelectrode measurements showed that ATP (0.1 mmol/l) hyperpolarized V-bl from -62 +/- 3 mV to -70 +/- 3 mV, an effect which was in some cases only transient (n = 7). This effect of ATP was different from that of carbachol, which depolarized V-bl. Applied together with secretin, ATP delayed the secretin-induced depolarization and prolonged the initial hyperpolarization of V-bl (n = 4). Several other putative agonists of pancreatic HCO3- secretion were also tested for their effects on [Ca2+](i). Bombesin (10 nmol/l) increased the fura-2 fluorescence ratio by 0.24 +/- 0.04 (n = 8), neurotensin (10 nmol/l) by 0.25 +/- 0.03 (rt = 6), substance P (0.1 mu mol/l) by 0.22 +/- 0.06 (n = 6), and cholecystokinin (10 nmol/l) by 0.14 +/- 0.03 (n = 7). Taken together, our studies show that Ca2+ homeostasis plays a role in pancreatic ducts. The most important finding is that carbachol and ATP markedly increase [Ca2+](i), but their different electrophysiological responses indicate that intracellular signalling pathways may differ.