PHORBOL ESTER, BUT NOT ISCHEMIC PRECONDITIONING, ACTIVATES PROTEIN-KINASE-D IN THE RAT-HEART

The signal transduction pathways that mediate the cardioprotective eff ects of ischemic preconditioning remain unclear. Here we have determin ed the role of a novel kinase, protein kinase D (PKD), in mediating pr econditioning in the rat heart, Isolated rat hearts (n=6/group) were s ubjected to either: (i) 36 min aerobic perfusion (control); (ii) 20 mi n aerobic perfusion plus 3 min no-flow ischemia, 3 min reperfusion, 5 min no-flow ischemia, 5 min reperfusion (ischemic preconditioning): (i ii) 20 min aerobic perfusion plus 200 nmol/l phorbol 12-myristate 13-a cetate (PMA) given as a substitute for ischemic preconditioning, The l eft ventricle then was excised, homogenized and PKD immunoprecipitated from the homogenate. Activity of the purified kinase was determined f ollowing incubation with [gamma(32)P]-ATP+/-syntide-2, a substrate for PKD. Significant PKD autophosphorylation and syntide-2 phosphorylatio n occurred in PMA-treated hearts, but not in control or preconditioned hearts. Additional studies confirmed that recovery of LVDP was greate r and initiation of ischemic contracture and time-to-peak contracture were less, in ischemic preconditioned hearts compared with controls (P <0.05). Our results suggest that the early events that mediate ischemi c preconditioning in the rat heart occur via a PKD-independent mechani sm. (C) 1997 Academic Press Limited.