Inhibition of cAMP- and Ca-dependent Cl- secretion by phorbol esters: inhibition of basolateral K+ channels.
Pretreating confluent T84 cells with the phorbol ester phorbol 12-myristate 13-acetate (PMA) inhibits adenosine 3',5'-cyclic monophosphate (cAMP)- and carbachol-induced Cl secretion. Both a sustained short-circuit current (Isc), seen after the addition of 50 microM 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate (CPT-cAMP) and 100 microM 3-isobutyl-1-methylxanthine (IBMX), and a transient current, seen after the subsequent addition of 100 microM carbachol, are inhibited by 80% following pretreatment with 100 nM PMA for 2 h. Pretreatment with PMA has no effect on the level of cystic fibrosis transmembrane conductance regulator protein or apical cAMP-dependent Cl conductance. Carbachol does not induce an increase in apical Cl conductance. Basolateral K conductance was measured in monolayers treated with apical nystatin and exposed to a K gradient. Agonist-independent K conductance is 10-fold greater in Cl media than in gluconate media. Pretreatment with PMA inhibits agonist-independent K conductance by 57% in Cl media but stimulates K conductance by 1.9-fold in gluconate media. The addition of carbachol induces a transient increase in basolateral K conductance, and pretreatment with PMA inhibits the agonist-dependent K conductance by 66% in Cl media and by 92% in gluconate media. In Cl media, serosal barium, at 3 mM, inhibits agonist-independent K conductance but has no significant effect on the carbachol-induced conductance. In nonpermeabilized monolayers, serosal barium inhibits the cAMP-dependent Isc by 56% but has no effect on the carbachol-induced Isc. These results demonstrate that the primary effect of PMA on Cl secretion is not inhibition of apical Cl channels but inhibition of basolateral K channels.(ABSTRACT TRUNCATED AT 250 WORDS)
Reenstra
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Children's Hospital Oakland Research Institute, California.
D001464:Barium / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D018118:Chloride Channels; D002712:Chlorides / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000242:Cyclic AMP / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004594:Electrophysiology; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism; D009761:Nystatin / Q000494:pharmacology; D011188:Potassium / Q000037:antagonists & inhibitors / Q000502:physiology; D015221:Potassium Channels / Q000187:drug effects*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D018118:Chloride Channels; D002712:Chlorides; D008565:Membrane Proteins; D015221:Potassium Channels; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D009761:Nystatin; D001464:Barium; D002217:Carbachol; D000242:Cyclic AMP; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium