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ATP-sensitive K+ (KATP) channels are unique metabolic sensors formed by association of Kir6.2, an inwardly rectifying K+ channel, and the sulfonylurea receptor SUR, an ATP binding cassette protein. We identified an ATPase activity in immunoprecipitates of cardiac KATP channels and in purified fusion proteins containing nucleotide binding domains NBD1 and NBD2 of the cardiac SUR2A isoform. NBD2 hydrolyzed ATP with a twofold higher rate compared to NBD1. The ATPase required Mg2+ and was insensitive to ouabain, oligomycin, thapsigargin, or levamisole. K1348A and D1469N mutations in NBD2 reduced ATPase activity and produced channels with increased sensitivity to ATP. KATP channel openers, which bind to SUR, promoted ATPase activity in purified sarcolemma. At higher concentrations, openers reduced ATPase activity, possibly through stabilization of MgADP at the channel site. K1348A and D1469N mutations attenuated the effect of openers on KATP channel activity. Opener-induced channel activation was also inhibited by the creatine kinase/creatine phosphate system that removes ADP from the channel complex. Thus, the KATP channel complex functions not only as a K+ conductance, but also as an enzyme regulating nucleotide-dependent channel gating through an intrinsic ATPase activity of the SUR subunit. Modulation of the channel ATPase activity and/or scavenging the product of the ATPase reaction provide novel means to regulate cellular functions associated with KATP channel opening.

Mutations in genes encoding ion channels have increasingly been identified to cause disease conditions collectively termed channelopathies. Recognizing the molecular basis of an ion channel disease has provided new opportunities for screening, early diagnosis, and therapy of such conditions. This synopsis provides an overview of progress in the identification of molecular defects in inwardly rectifying potassium (Kir) channels. Structurally and functionally distinct from other channel families, Kir channels are ubiquitously expressed and serve functions as diverse as regulation of resting membrane potential, maintenance of K(+) homeostasis, control of heart rate, and hormone secretion. In humans, persistent hyperinsulinemic hypoglycemia of infancy, a disorder affecting the function of pancreatic beta cells, and Bartter's syndrome, characterized by hypokalemic alkalosis, hypercalciuria, increased serum aldosterone, and plasma renin activity, are the two major diseases linked so far to mutations in a Kir channel or associated protein. In addition, the weaver phenotype, a neurological disorder in mice, has also been associated with mutations in a Kir channel subtype. Further genetic linkage analysis and full understanding of the consequence that a defect in a Kir channel would have on disease pathogenesis are among the priorities in this emerging field of molecular medicine.

Bacteria that produce heat-stable enterotoxins (STs), a leading cause of secretory diarrhea, are a major cause of morbidity and mortality worldwide. ST stimulates guanylyl cyclase C (GCC) and accumulation of intracellular cyclic GMP ([cGMP]i), which opens the cystic fibrosis transmembrane conductance regulator (CFTR)-related chloride channel, triggering intestinal secretion. Although the signaling cascade mediating ST-induced diarrhea is well characterized, antisecretory therapy targeting this pathway has not been developed. 2-ChloroATP (2ClATP) and its cell-permeant precursor, 2-chloroadenosine (2ClAdo), disrupt ST-dependent signaling in intestinal cells. However, whether the ability to disrupt guanylyl cyclase signaling translates into effective antisecretory therapy remains untested. In this study, the efficacy of 2ClAdo to prevent ST-induced water secretion by human intestinal cells was examined. In Caco-2 human intestinal cells, ST increased [cGMP]i, induced a chloride current, and stimulated net basolateral-to-apical water secretion. This effect on chloride current and water secretion was mimicked by the cell-permeant analog of cGMP, 8-bromo-cGMP. Treatment of Caco-2 cells with 2ClAdo prevented ST-induced increases in [cGMP]i, chloride current and water secretion. Inhibition of the downstream consequences of ST-GCC interaction reflects proximal disruption of cGMP production because 8-bromo-cGMP stimulated chloride current and water secretion in 2ClAdo-treated cells. Thus, this study demonstrates that disruption of guanylyl cyclase signaling is an effective strategy for antisecretory therapy and provides the basis for developing mechanism-based treatments for enterotoxigenic diarrhea.