Myosin regulation of NKCC1: effects on cAMP-mediated Cl- secretion in intestinal epithelia

Myosin regulation of NKCC1: effects on cAMP-mediated Cl- secretion in intes tinal epithelia. Am. J. Physiol. 277 (Cell Physiol. 46): C441-C447, 1999.-T he basally located actin cytoskeleton has been demonstrated previously to r egulate Cl- secretion from intestinal epithelia via its effects on the Na+- K+-2Cl cotransporter (NKCC1). In nontransporting epithelia, inhibition of m yosin light chain kinase (MLCK) prevents cell-shrinkage-induced activation of NKCC1. The aim of this study was to investigate the role of myosin in th e regulation of secretagogue-stimulated Cl- secretion in intestinal epithel ia. The human intestinal epithelial cell line T84 was used for these studie s. Prevention of myosin light chain phosphorylation with the MLCK inhibitor ML-9 or ML-7 and inhibition of myosin ATPase with butanedione monoxime (BD M) attenuated cAMP but not Ca2+-mediated Cl- secretion. Both ML-9 and BDM d iminished cAMP activation of NKCC1. Neither apical Cl- channel activity, ba solateral K+ channel activity, nor Na+-K+-ATPase were affected by these age nts. Cytochalasin D prevented such attenuation, cAMP-induced rearrangement of basal actin microfilaments was prevented by both ML-9 and BDM. The phosp horylation of mosin light chain and subsequent contraction of basal actin-m yosin bundles are crucial to the cAMP-driven activation of NKCC1 and subseq uent apical Cl- efflux.