DIFFERENT REGULATORY PATHWAYS INVOLVED IN ATP-STIMULATED CHLORIDE SECRETION IN RAT EPIDIDYMAL EPITHELIUM

The regulatory pathways involved in the ATP-stimulated Cl- secretion across rat epididymal epithelium were investigated by the short-circuit current (I-SC) technique. Biphasic characteristic was observed in the I-SC responded to ATP (0.01-10 mu M). Inhibitor of P-1 receptor, 8-phenyltheophylline (up to 100 mu M), did not have any effect on both phases of the ATP-stimulated I-SC. The order of potency for stimulation of the two phases in I-SC was ATP > ADP >> AMP, adenosine, consistent with the presence of P-2-purinoceptors. Cl- channel blocker, disulfonic acid stilbene (DIDS, 300 mu M), only inhibited the first peak of the ATP-stimulated I-SC while diphenylamine-dicarboxylic acid (DPC, 1 mM) reduced both, indicating the involvement of different conductance pathways. DIDS was found to have an inhibitory effect on Ca2+-activated I-SC (induced by ionomycin, 10 mu M) but not cAMP-activated I-SC (induced by forskolin, 1 mu M) which could only be blocked by DPC. Both peaks of the ATP-activated I-SC could be significantly inhibited by pretreatment with a Ca2+-chelating agent, BAPTA-AM (50 mu M). An increase in cellular cAMP content upon stimulation of ATP was measured by radioimmunoassay. No significant increase in cAMP production was observed in cells stimulated with adenosine. The ATP-induced cAMP increase was prevented by pretreatment with BAPTA-AM (100 mu M) indicating that cAMP production upon ATP stimulation was secondary to an increase in intracellular Ca2+ concentration. These results indicate that the ATP-stimulated Cl- secretion could be mediated by Ca2+ and cAMP-dependent regulatory pathways giving rise to the biphasic nature of the ATP-induced I-SC. (C) 1995 Wiley-Liss, Inc.