Stimulation of intestinal Cl- transport by heat-stable enterotoxin: activation of cAMP-dependent protein kinase by cGMP.
Heat-stable enterotoxins activate guanylate cyclase, whereas heat-labile enterotoxins stimulate adenylate cyclase. Both classes of toxins cause secretory diarrhea at least in part by stimulating Cl- secretion in the intestine. The mechanism for regulation of Cl- secretion by guanosine 3',5'-cyclic monophosphate (cGMP) was investigated using cultured T84 intestinal cells as a model for intestinal crypt cells. Escherichia coli heat-stable enterotoxin (ST) markedly stimulated cGMP production in T84 cells. Cl- secretion across T84 cell monolayers cultured on permeable filters was stimulated by E. coli ST, cholera toxin, or 8-BrcAMP, but 8-BrcGMP was ineffective. cGMP analogues that are known to be potent and specific activators of cGMP-dependent protein kinase (cG-kinase) also had little effect on 36Cl- uptake by T84 cells cultured in plastic dishes. E. coli ST, forskolin, cholera toxin, or membrane-permeant cAMP analogues markedly increased 36Cl- uptake into T84 cells. The general protein kinase inhibitor, staurosporine, inhibited the stimulation of Cl- permeability elicited by E. coli ST, vasoactive intestinal peptide (VIP), or 8-BrcAMP. DEAE-Sephacel chromatography revealed a predominant type II isoform of cAMP-dependent protein kinase (cA-kinase) in T84 cells, whereas little or no cytosolic cG-kinase activity was found. Treatment of T84 cells with E. coli ST or VIP resulted in an increase in the cA-kinase activity ratio (-cAMP/+cAMP) if the cytosolic enzyme was assayed at reduced temperature (on ice).(ABSTRACT TRUNCATED AT 250 WORDS)
Forte
L R
LR
Department of Pharmacology, Missouri University School of Medicine, Columbia.
Thorne
P K
PK
Eber
S L
SL
Krause
W J
WJ
Freeman
R H
RH
Francis
S H
SH
Corbin
J D
JD
D001692:Biological Transport / Q000187:drug effects; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology*; D004355:Drug Stability; D004768:Enterotoxins / Q000494:pharmacology*; D004789:Enzyme Activation; D004926:Escherichia coli; D006358:Hot Temperature; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
D002712:Chlorides; D004768:Enterotoxins; D014660:Vasoactive Intestinal Peptide; D011494:Protein Kinases; D006152:Cyclic GMP