Demonstration of selective protein kinase C-dependent activation of Src and Lck tyrosine kinases during ischemic preconditioning in conscious rabbits

Src tyrosine kinases have been shown to mediate cellular responses to stres s in noncardiac cells. However, the effect of myocardial ischemia on Src ty rosine kinases is unknown. Furthermore, the identity of the tyrosine kinase (s) involved in the genesis of ischemic preconditioning (PC) remains obscur e. Here, we present the first evidence that ischemic PC (6 cycles of 4-minu te coronary occlusion and il-minute reperfusion) induces selective activati on of 2 members of the Src family of tyrosine kinases, Src and Lck, in the heart of conscious rabbits. The activation of Src in the particulate fracti on was not evident at 5 minutes after ischemic PC but became apparent at 30 minutes (+119% versus control), whereas the activation of Lck in the parti culate fraction was apparent both at 5 minutes (+103% versus control) and a t 30 minutes (+89%) after ischemic PC. The activity of the other 5 members of the Src tyrosine kinases expressed in the rabbit heart (Fyn, Fgr, Yes, L yn, and Blk) was not affected by ischemic PC. Ischemic PC had no effect on the activity of epidermal growth factor receptor kinases, either at 5 or at 30 minutes. The activation of Src and Lck was completely abrogated by the tyrosine kinase inhibitor lavendustin A, given at doses that have previousl y been shown to block the protective effect of ischemic PC in this same con scious rabbit model, suggesting that Src and Lck kinases are essential for the development of ischemic PC. The activity of the epsilon isoform of prot ein kinase C (PKC) in the particulate fraction increased at 5 minutes (+72% ) and at 30 minutes (+67%) after ischemic PC. Pretreatment with lavendustin A had no effect on the activation of PKC epsilon, whereas pretreatment wit h the PKC inhibitor chelerythrine (given at doses that have previously been shown to block ischemic PC) blocked not only the activation of PKC epsilon but also that of Src and Lck, indicating that Src and Lck are downstream o f PKC epsilon in the signaling cascade of ischemic PC, This study identifie s a new component of the signaling mechanism of ischemic PC. The results su pport the concept that, in conscious rabbits, 2 specific members of the Src family of tyrosine kinases, Src and Lck, play an important role in the gen esis of late PC by serving as downstream elements of PKC-mediated signal tr ansduction.