Modulation of Cl- secretion in rat distal colon by genistein, a protein tyrosine kinase inhibitor

The protein tyrosine kinase inhibitor, genistein, caused an increase of short-circuit current (Isc) across the rat distal colon in forskolin-pretreated tissues, suggesting a synergistic interaction of the drug with cAMP-dependent secretion. In the absence of forskolin, genistein had a dual effect on Isc, it increased Isc in tissues with a low baseline, but decreased Isc in tissues with a high baseline Isc. The secretory effect of genistein was dependent on the presence of Cl- and was blocked by inhibitors of Cl- secretion like bumetanide, an inhibitor of the Na+-K+-Cl- cotransporter, or 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), a Cl- channel blocker. Unidirectional flux measurements revealed that genistein inhibited Na+ and Cl- absorption and induced net Cl- secretion. The protein tyrosine phosphatase inhibitor vanadate suppressed the secretory effect of genistein. In contrast, genistein caused an inhibition of carbachol-induced, i.e. Ca2+-mediated secretion. Whole-cell patch-clamp experiments confirmed the synergistic effect of genistein on cAMP-induced Cl- currents. In the presence of forskolin, genistein caused a depolarization concomitant with an increase in membrane inward current. In addition, genistein caused an inhibition of a basal K+ conductance and inhibited the Ca2+-dependent K+ conductance stimulated by carbachol. These results suggest a complex role of the protein tyrosine kinase pathway in the control of colonic Cl- secretion, an antagonistic action on the cAMP pathway and a synergistic action on the Ca2+ pathway as revealed by the opposing effects of genistein. The physiological importance of this regulation remains to be clarified.