Protein kinase Cepsilon mediates salutary effects on electrical coupling induced by ischemic preconditioning.
BACKGROUND
Ischemic preconditioning delays the onset of electrical uncoupling and prevents loss of the primary ventricular gap junction protein connexin 43 (Cx43) from gap junctions during subsequent ischemia.


OBJECTIVE
To test the hypothesis that these effects are mediated by protein kinase C epsilon (PKCepsilon), we studied isolated Langendorff-perfused hearts from mice with homozygous germline deletion of PKCepsilon (PKCepsilon-KO).


METHODS
Cx43 phosphorylation and distribution were measured by quantitative immunoblotting and confocal microscopy. Changes in electrical coupling were monitored using the 4-electrode technique to measure whole-tissue resistivity.


RESULTS
The amount of Cx43 located in gap junctions, measured by confocal microscopy under basal conditions, was significantly greater in PKCepsilon-KO hearts compared with wild-type, but total Cx43 content measured by immunoblotting was not different. These unanticipated results indicate that PKCepsilon regulates subcellular distribution of Cx43 under normal conditions. Preconditioning prevented loss of Cx43 from gap junctions during ischemia in wild-type but not PKCepsilon-KO hearts. Specific activation of PKCepsilon, but not PKCdelta, also prevented ischemia-induced loss of Cx43 from gap junctions. Preconditioning delayed the onset of uncoupling in wild-type but hastened uncoupling in PKCepsilon-KO hearts. Cx43 phosphorylation at the PKC site Ser368 increased 5-fold after ischemia in wild-type hearts, and surprisingly, by nearly 10-fold in PKCepsilon-KO hearts. Preconditioning prevented phosphorylation of Cx43 in gap junction plaques at Ser368 in wild-type but not PKCepsilon-KO hearts.


CONCLUSIONS
Taken together, these results indicate that PKCepsilon plays a critical role in preconditioning to preserve Cx43 signal in gap junctions and delay electrical uncoupling during ischemia.
Hund
Thomas J
TJ
Department of Surgery, School of Medicine, Washington University, St. Louis, Missouri, USA.
Lerner
Deborah L
DL
Yamada
Kathryn A
KA
Schuessler
Richard B
RB
Saffitz
Jeffrey E
JE
D000818:Animals; D018031:Connexin 43 / Q000378:metabolism*; D017097:Electric Impedance; D017629:Gap Junctions / Q000378:metabolism; D019157:Ischemic Preconditioning, Myocardial; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D023421:Models, Animal; D017202:Myocardial Ischemia / Q000378:metabolism*; D015425:Myocardial Reperfusion; D010766:Phosphorylation; D051744:Protein Kinase C-epsilon / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D011817:Rabbits
D018031:Connexin 43; C500216:GJA1 protein, human; D051744:Protein Kinase C-epsilon