Guanosine 3',5'-cyclic monophosphate-dependent protein kinase II mediates heat-stable enterotoxin-provoked chloride secretion in rat intestine

Background & Aims: Escherichia coli heat-stable enterotoxins (STa) provoke electrogenic Cl- secretion in the intestine through a guanosine 3',5'-cyclic monophosphate (cGMP)-dependent signal transduction pathway, The cGMP receptor involved in the activation of the Cl- channel is not known with certainty but may comprise either adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase (cAK) or cGMP-dependent protein kinase (cGK) type II, The aim of this study was to discriminate between these possibilities using specific kinase inhibitors, Methods: Intestinal electrogenic Cl- secretion was determined by measuring short-circuit current (Isc) in a Ussing chamber, Results: The general protein kinase inhibitors staurosporine and H-8 inhibited rat cGK II activity in vitro with 50% inhibitory concentration values of 4 nmol/L and 3 mu mol/L, respectively, which are lower than those reported for cAK, Both staurosporine and H-8, when added to rat proximal colon at concentrations that did not affect the Isc response to 8-bromo-cAMPS, inhibited the STa- and 8-bromo-cGMP-provoked Isc response for more than 80%, Furthermore, the relative specific cGK inhibitor Rp isomer of 8-(chlorophenylthio)-cGMP, but not the cAK inhibitor RP isomer of (Rp) 8-bromo-cAMPS, inhibited the Isc response to submaximal levels of STa in rat proximal colon, Conclusions: These data provide further evidence for an important role of cGK II in STa-mediated Cl- secretion in native rat intestinal epithelium.