Adenosine A(1) receptors selectively target protein kinase C isoforms to the caveolin-rich plasma membrane in cardiac myocytes.

摘要

Adenosine is a naturally occurring nucleoside that has been shown to regulate a variety of functions in the cardiovascular system. However, the mechanisms in adenosine receptor signaling are not completely understood. Given that adenosine receptors have been linked to protein kinase C (PKC) in cardioprotection and caveolae is critical for receptor signaling, we sought to determine whether activation of adenosine A1 receptors induces selective translocation of PKC isoforms to the membrane from the cytosol and whether activated PKC is targeted to the caveolin-rich plasma membrane microdomains. The freshly isolated adult rat cardiac myocytes were used to examine PKC isoforms including PKCalpha, PKCbeta, PKCvarepsilon, PKCdelta and PKCzeta. Immunoblot analysis revealed that the immunoreactivity for PKCvarepsilon or PKCdelta but not for PKCalpha, PKCbeta or PKCzeta increased significantly in the membrane fractions from cells pretreated with the selective adenosine A1 receptor agonist 2-chloro-N(6)-cyclopentyladenosine (CCPA, 100 nM) when compared with non-stimulated cells. The effect of CCPA on PKCvarepsilon or PKCdelta translocation was blocked by adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 100 nM). When Western blot was performed from the caveolin-enriched plasma membrane fractions, the immunoreactivity for PKCvarepsilon or PKCdelta but not PKCalpha, PKCbeta or PKCzeta was enhanced significantly by CCPA. Furthermore, PKCvarepsilon and PKCdelta were detected in the anti-caveolin-3 immunoprecipitates but not in the samples without primary antibody. Immunofluorescence staining further indicates increased colocalization of PKCvarepsilon or PKCdelta with caveolin-3 at cell peripheral region and T-tubular-like structures in response to adenosine A1 receptor activation. In conclusion, we demonstrate that activation of adenosine A1 receptors promotes the selective translocation of PKCvarepsilon and PKCdelta to the caveolin-enriched plasma membrane microdomains in cardiac myocytes.