NA-K+-2CL- COTRANSPORT AND CL- SECRETION EVOKED BY HEAT-STABLE ENTEROTOXIN IS MICROFILAMENT DEPENDENT IN T84 CELLS()

We previously reported that adenosine 3',5'-cyclic monophosphate-media ted stimulation of Cl- secretion in the human intestinal epithelial ce ll line T84 is accompanied by significant remodeling of F-actin and th at both the secretory and cytoskeletal responses may be inhibited by p halloidin derivatives, agents that polymerize actin and prevent dynami c reorganization of microfilaments. In contrast, the carbachol-elicite d Cl- secretory response (Ca2+ mediated) was not attenuated by phalloi din (J. Clin. Invest. 87: 1903-1909, 1991). In the present study, we e xamine the effect of phalloidin on the Cl- secretory response elicited by the heat-stable enterotoxin of Escherichia coli (ST.), which induc es elevations in intracellular guanosine 3',5'-cyclic monophosphate. W e find that apical adminstration of 1 muM ST. results in a regionally restricted redistribution of F-actin confined to the basal pole of the cells. In monolayers pretreated with phalloidin, the Cl- secretory re sponse to ST. was inhibited by >60%. Sequential treatment of phalloidi n-loaded monolayers with ST(a) followed by carbachol resulted in a syn ergistic secretory response that was not different from control (unloa ded) monolayers. Examination of efflux/uptake through specific membran e transport pathways involved in ST(a)-stimulated Cl- secretion indica ted normal activation of apical Cl- and basolateral K+ channels under phalloidin-loaded conditions. The ability of ST(a)-treated monolayers to pump Na+ in an absorptive direction was also unaffected by phalloid in. Under phalloidin-loaded conditions, ST(a)-stimulated Na+-K+2Cl-cot ransporter activity was reduced by -60%, sufficient to account for the observed inhibition of net Cl- secretory response. We conclude that S T. elicits a cytoskeletal rearrangement that is integral to its abilit y to stimulate electrogenic Cl-secretion. Phalloidin appears to specif ically inhibit the functional activity of the Na+-K+-2Cl- cotransporte r and thereby inhibits the generation of a sustained secretory respons e to ST(a). This study provides further evidence that the short-term r egulation of intestinal salt and water secretion may be influenced by interaction between the actin-based cytoskeleton and ion transport pro teins.