Phosphorylation of extrajunctional Cx43 in ischemic-preconditioned rat hearts.
Ischemic preconditioning (IP) has been found to intensify ischemia-induced spreading of connexin 43 (Cx43) into the extrajunctional plasma membrane of the rat heart. Since ischemia is known to induce connexin dephosphorylation and plasmalemmal permeabilization, we considered whether IP might delay dephosphorylation of also extrajunctional connexin and thus impede myocyte permeabilization. Regional myocardial ischemia of both 15 and 45 min duration with and without IP, respectively, was induced in anesthetized rats. Sections of the hearts were immunostained for phosphorylated and dephosphorylated Cx43. The ratios of immunofluorescence in gap junctions and extrajunctional membranes, respectively, versus myocyte interiors were determined as relative fluorescence units (RFU). IP, however, was not found to reduce gap junctional dephosphorylated Cx43 (normal perfusion: 1.23 +/- 0.06 RFU; 15 min ischemia without and with IP: 1.80 +/- 0.09 and 2.10 +/- 0.24 RFU; 45 min ischemia without and with IP: 3.12 +/- 0.55 and 2.57 +/- 0.33 RFU, respectively). Extrajunctionally, only dephosphorylated Cx43 was found. Its occurrence was not prevented by IP, it was rather intensified by IP (normal perfusion: 1.02 +/- 0.01 RFU, 15 min ischemia without and with IP: 1.06 +/- 0.03 and 1.15 +/- 0.05 RFU (P < 0.02), and 45 min ischemia without and with IP: 1.30 +/- 0.07 and 1.27 +/- 0.07 RFU, respectively). Although under ischemia, junctional and extrajunctional Cx43 predominated in the dephosphorylated state, only a small fraction of myocytes were found permeabilized to propidium iodide. In conclusion, IP did not prevent ischemia-induced Cx43 dephosphorylation in gap junctions and extrajunctional plasma membranes. It is thus more likely that changes in non-channel functions or other localizations of Cx43 are involved in IP-induced myocardial protection.
Mühlfeld
Christian
C
Center of Anatomy, Division of Electron Microscopy, University of Göttingen, Göttingen, Germany. christian.muehlfeld@anatomie.med.uni-giessen.de
Cetegen
Cenk
C
Freese
Sigrid
S
Volkmann
Rolf
R
Hellige
Gerhard
G
Vetterlein
Friedrich
F
D000818:Animals; D004396:Coloring Agents; D018031:Connexin 43 / Q000378:metabolism*; D019157:Ischemic Preconditioning, Myocardial; D017202:Myocardial Ischemia / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D009243:NAD / Q000378:metabolism; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism*; D011419:Propidium; D051381:Rats
D004396:Coloring Agents; D018031:Connexin 43; D009243:NAD; D011419:Propidium; D010770:Phosphotransferases