ISOLATED-PERFUSED RABBIT COLON CRYPTS - STIMULATION OF CL- SECRETION BY FORSKOLIN

The aim of this study was to characterize ion conductances and carrier mechanisms of isolated in vitro perfused rabbit colonic crypts. Crypts were isolated from rabbit colon mucosa and mounted on a pipette system which allowed controlled perfusion of the lumen. In non-stimulated conditions basolateral membrane voltage (V-bl) was -65 +/- 1 mV (n = 240). Bath Ba2+ (1 mmol/l) and verapamil (O.1 mmol/l) depolarized V-bl by 21 +/- 2 mv (n = 7) and 31 +/- 1 (n = 4), respectively. Lowering of bath Cl- concentration hyperpolarized V-bl from -69 +/- 3 to -75 +/- 3 mV (n = 9). Lowering of luminal Cl- concentration did not change V-bl. Basolateral application of loop diuretics (furosemide, piretanide, bumetanide) had no influence on V-bl in non-stimulated crypts. Forskolin (10(-6) mol/l) in the bath depolarized V-bl by 29 +/- 2 mV (n = 54) and decreased luminal membrane resistance. In one-third of the experiments a spontaneous partial repolarization of V-bl was seen in the presence of forskolin. During forskolin-induced depolarization basolateral application of loop diuretics hyperpolarized V-bl significantly and concentration dependently with a potency sequence of bumetanide > piretanide greater than or equal to furosemide. Lowering bath Cl- concentration hyperpolarized V-bl. Lowering of luminal Cl- concentration from 120 to 32 mmol/l during forskolin-induced depolarization led to a further depolarization of V-bl by 7 +/- 2 mV (n = 10). We conclude that V-bl of rabbit colonic crypt cells is dominated by a K+ conductance. Stimulation of the cells by forskolin opens a luminal Cl- conductance. Basolateral uptake of Cl- occurs via a basolateral Na+: 2Cl(-): K+ cotransport system.