Characterization of the mechanism of action of Escherichia coli heat-stable enterotoxin.
The mechanism of activation of intestinal guanylate cyclase by Escherichia coli heat-stable enterotoxin (STa) has been studied by using isolated rat intestinal epithelial cells and purified brush border membrane (BBM) preparations. Inhibitors of prostaglandin biosynthesis, quinacrine and 5,8,11,14-eicosatetraynoic acid (ETYA), significantly reduced intracellular levels of cyclic guanosine 3', 5'-monophosphate in isolated cells treated with STa. Although these data suggested that activation of phospholipase A2 and metabolism of arachidonic acid are involved in the mechanism of action of STa, other data ruled out such a mechanism. (i) The rate of release of [3H]arachidonic acid by prelabeled intestinal cells incubated with STa was the same as control cells not treated with STa. (ii) Thin-layer chromatography of lipid extracts of intestinal cells treated with STa and untreated cells did not reveal any quantitative or qualitative differences in free fatty acids, neutral lipids, and phospholipids. (iii) Amounts of prostaglandin PGE2, prostaglandin PGF2 alpha, and thromboxane B2 in intestinal cells and BBM incubated with STa did not increase compared with controls not incubated with STa. When purified BBM preparations were incubated with phospholipase A2 inhibitors (p-bromophenacyl bromide and quinacrine) or cyclooxygenase inhibitors (ETYA and indomethacin), basal and STa-induced guanylate cyclase activities were significantly reduced. Inhibitors of calcium-calmodulin-mediated reactions (EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid], trifluoperazine, and chlorpromazine) and calcium channel blockers (verapamil and nifedipine) also nonspecifically inhibited both basal and STa-stimulated guanylate cyclase in BBM preparations. Lanthanum, a competitive inhibitor of membrane-bound calcium, did not affect either basal or STa-stimulated guanylate cyclase of BBM preparations.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D006152:Cyclic GMP / Q000378:metabolism; D004768:Enterotoxins / Q000494:pharmacology*; D004848:Epithelium / Q000201:enzymology / Q000378:metabolism; D029968:Escherichia coli Proteins; D005260:Female; D006162:Guanylate Cyclase / Q000378:metabolism*; D007421:Intestine, Small / Q000201:enzymology* / Q000378:metabolism; D007700:Kinetics; D008055:Lipids / Q000302:isolation & purification; D008871:Microvilli / Q000201:enzymology; D011453:Prostaglandins / Q000378:metabolism; D011796:Quinacrine / Q000494:pharmacology; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains
D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D008055:Lipids; D011453:Prostaglandins; C034151:heat stable toxin (E coli); D006162:Guanylate Cyclase; D011796:Quinacrine; D006152:Cyclic GMP