Abstract 845: Cardioprotection From GSK-3β Inhibition Requires Activation of PI-3 Kinase and Src but not Akt, PKC, or ERK
Objective: Inhibition of glycogen synthase kinase-3β (GSK-3β) at onset of reperfusion is a key event in preconditioning’s protection. To determine GSK-3β’s position in the signal cascade we tested whether protection from its inhibition with SB216763 (SB) requires activation of other kinases including phosphatidylinositol-3 kinase (PI3K), Src, Akt, protein kinase C (PKC), and extracellular signal-regulated kinases (ERK).
Methods and results: In isolated rabbit hearts with 30-min regional ischemia/2-h reperfusion SB (1μM) at reperfusion reduced infarction from 38.5±3.7% of the risk zone in control hearts to 9.9±1.6%. Co-infusion of either the PI3K blocker wortmannin or PP2, a selective Src inhibitor, aborted the protective effects (29.5±4.0% and 30.4±1.6% infarction, resp.). In contrast, inhibitors of PKC, ERK 1/2, or Akt or blockade of adenosine receptors were unable to block SB-induced protection indicating upstream locations of these kinases and receptors. SB increased Akt but not ERK1/2 phosphorylation. Thus Akt serves as a reporter for PI3K but does not participate in protection from direct GSK-3β inhibition. Isolated adult ventricular rat myocytes were loaded with tetramethylrhodamine ethyl ester (TMRE) and exposed to H2O2 (100μM) for 40 min. Fluorescence intensity was reduced compared to untreated cells (99.4±20.5 vs. 292.6±22.4 a.u.) suggesting oxidant injury opened mitochondrial permeability transition pores causing mitochondria to lose TMRE. Cells infected with an adenovirus containing dominant-negative (dn) GSK-3β plasmid were protected from H2O2 incubation (237.2±29.7 a.u.). Interestingly dnGSK-transfected cells had increased Akt phosphorylation.
Conclusion: Therefore inhibition of GSK-3β is protective in hearts and isolated myocytes. PI3K and Src are required downstream components in GSK-3β’s protection, while Akt, ERK 1/2, and PKC are not.