Effects of phorbol ester on gap junctions of neonatal rat heart cells.
Myocytes were isolated from the ventricles of neonatal rat hearts and cultured for 1-3 days. Newly formed cell pairs were used to examine the conductance of gap junctions, gj. Measurements were performed using a dual voltage-clamp method in conjunction with a whole-cell, tight-seal recording. Exposure to the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA, 100-160 nM) led to a decrease in gj. Single-channel events recorded immediately before complete uncoupling yielded a single-channel conductance, gamma j, of 40.5 pS, implying that TPA affects the channel kinetics rather than gamma j. TPA-induced uncoupling was observed at subphysiological levels of cytosolic Ca2+ (pipette solution = 18 nM), not at physiological levels (pipette solution = 170 nM). The effects of TPA could not be mimicked by 250 microM 1-oleoyl-2-acetyl-glycerol (OAG). Preincubation with TPA (up to 24 h) revealed no changes in gj attributable to down-regulation of protein kinase C, PKC. Pretreatment with PKC inhibitors, staurosporine or PKCI, prevented the TPA-dependent decrease in gj. TPA-dependent uncoupling was not impaired by 4-bromophenacyl bromide, an inhibitor of phospholipase A2, PLA2; conversely, an arachidonic acid-dependent decrease in gj was not prevented by PKCI. This suggests that gj regulation does not involve an interaction between PLA2 and PKC.
Münster
P N
PN
Department of Physiology, University of Bern, Switzerland.
Weingart
R
R
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D016718:Arachidonic Acid / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004075:Diglycerides / Q000494:pharmacology; D004553:Electric Conductivity; D006352:Heart Ventricles / Q000648:ultrastructure; D007365:Intercellular Junctions / Q000187:drug effects / Q000502:physiology*; D007700:Kinetics; D009206:Myocardium / Q000648:ultrastructure*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D000470:Alkaloids; D004075:Diglycerides; D016718:Arachidonic Acid; C038237:1-oleoyl-2-acetylglycerol; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium