Translocation and Down‐Regulation of Protein Kinase C‐α, ‐β, and ‐γ Isoforms During Ischemia‐Reperfusion in Rat Brain

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Abstract
We investigated the distribution of protein kinase C (PKC) isoforms in the subcellular fractions (P1, 1,000-g pellet; P2, 10,000-g pellet; P3, 100,000-g pellet; S, 100,000-g supernatant) of rat forebrain after ischemia or reperfusion by immunoblotting. PKC-δ and -ε isoforms were predominant in the P2 (synaptosome-rich) fraction, whereas PKC-α, -β, -γ, -ε, and -ζ isoforms were rich in the S (cytosolic) fraction. With time of ischemia (5-30 min), PKC-α, -β, and -γ translocated to the P2 and P3 fractions, whereas reperfusion for 60 min after 30 min of ischemia reduced PKC-β activity greatly and PKC-α and -γ activities to a lesser extent. There was no redistribution of PKC-δ, -ε, and -ζ after ischemia or reperfusion. A calpain inhibitor, acetylleucylleucylnorleucinal, inhibited the down-regulation of PKC-β, through intravenous injection. The PKC translocation to the P2 fraction was accompanied by their dephosphorylation, transition of PKC-α from dimer to trimer, and the decrease in activity. These data show that PKC-α, -β, and -γ isoforms translocate chiefly to the synaptosome in ischemic brain in association with the dephosphorylation, multimeric change, and inactivation, followed by the proteolysis of PKC-β by calpain after postischemic reperfusion.