Downregulation of Protein Kinase C Inhibits Activation of Mitochondrial K _ATP Channels by Diazoxide

Abstract

Background-The mitochondrial K ATP (mitoK ATP ) channel has been shown to confer short-and long-term cardioprotection against prolonged ischemia via protein kinase C (PKC) signaling pathways. However, the exact association between PKC or its isoforms and mitoK ATP channels has not yet been clarified. The present study tested the hypothesis that the activity and translocation of PKC to the mitochondria are important for cardiac protection elicited by mitoK ATP channels. Methods and Results-PKC was downregulated by prolonged (24-hour) treatment with phorbol 12-myristate 13-acetate (4 g/kg body weight) before subsequent experiments in rats. Langendorff-perfused rat hearts were subjected to 40 minutes of ischemia followed by 30 minutes of reperfusion. Effects of PKC downregulation on the activation of mitoK ATP channels and other interventions on hemodynamic, biochemical, and pathological changes were assessed. Subcellular localization of PKC isoforms by Western blot analysis and immunocytochemistry demonstrated that PKC-␣ and PKC-␦ were translocated to the sarcolemma and that PKC-␦ was translocated to the mitochondria after diazoxide treatment. In hearts treated with diazoxide (80 mol/L), a significant improvement in cardiac function and an attenuation of cell injury were observed. In PKC-downregulated hearts, protection was abolished because mitoK ATP channels could not be activated by diazoxide. Conclusions-These data suggest that PKC activation is required for the opening of mitoK ATP channels during protection against ischemia and that this effect is linked to isoform-specific translocation of PKC-␦ to the mitochondria.

Effects of bisindolylmaleimide PKC inhibitors on p90RSK activity in vitro and in adult ventricular myocytes

“…These inhibitors have also been utilised by this and other laboratories to explore the involvement of PKC isoforms in the stimulation of the sarcolemmal Na þ /H þ exchanger (NHE1) by diverse stimuli, such as adrenergic (Puce´at et al, 1993;Snabaitis et al, 2000), thrombin (Yasutake et al, 1996), angiotensin (Gunasegaram et al, 1999) and opioid (Bian et al, 2000) receptor agonists, anaesthetic agents (Kanaya et al, 2001) and oxidative stress (Snabaitis et al, 2002). As an alternative to pharmacological PKC inhibition, some investigators have utilised PKC downregulation by prolonged phorbol ester stimulation (Clerk et al, 1998;Wang et al, 2001c). However, this method requires an extended period (atleast 24 h) of such stimulation and is likely to be accompanied by compensatory changes that would not occur with acute pharmacological inhibition of PKC.…”

Possible Role of JAK-2 in Attenuated Cardioprotective Effect of Ischemic Preconditioning in Hyperhomocysteinemic Rat Hearts

“…46) LDH is an enzyme that increases in myocardial infarction after reperfusion, which may be due to sustained ischemic injury. 47) The increase in infarct size and the release of LDH and CK are an index of I/R-induced myocardial injury. 48,49) In the present study, 30-min ischemia followed by 120-min reperfusion produced myocardial injury as shown by increased infarct size in the heart and elevated release of LDH and CK in the coronary eOEuent, which were consistent with earlier reports.…”

KR-31466, a Benzopyranylindol Analog, Attenuates Hypoxic Injury Through Mitochondrial KATP Channel and Protein Kinase C Activation in Heart-Derived H9c2 Cells

“…We previously reported the role of PKC as a mediator of cardioprotection in H9c2 cells (11) and isolated rat heart model (23). Recently, it has been suggested that the protective effect of mtK ATP channel opener is associated with PKC (24), by showing that the protective effect of DZX was not observed in hearts after PKC downregulation (24). Thus, in this study, we wanted to determine whether PKC is involved in the KR466-induced cardioprotective mechanism, by using a pharmacological inhibitor.…”

“…In the immunocytochemical study, PKC-d has been translocated to the mitochondria and sarcolemma after treatment with DZX, producing cardiac protection against ischemia (28). Furthermore, in PKC-d downregulated hearts, cardioprotection by DZX was abolished (24). In spite of these reports about the role of PKC-d (29) as a mediator of mtK ATP channel opening, it is still unclear which isozymes of PKC are coupled to protective mechanism by mtK ATP channel opening.…”