Reciprocal regulation between M3 muscarinic acetylcholine receptor and protein kinase C-ε in ventricular myocytes during myocardial ischemia in rats

We have studied the association between M-3 muscarinic acetylcholine receptors (M-3-mAChR) and protein kinase C-epsilon (PKC-epsilon) during ischemic myocardial injury using Western blot analysis and immunoprecipitation technique. Myocardial ischemia (MI) induced PKC-epsilon translocation from cytosolic to membrane fractions. This translocation participated in the phosphorylation of M-3-mAChR in membrane fractions, which could be abolished by the inhibitor of PKC, chelerythrine chloride. On the other hand, M-3-mAChR could also regulate the expression of PKC-epsilon in ischemic myocardium. Choline (choline chloride, an M-3 receptor agonist, administered at 15 min before occlusion) strengthened the association between PKC-epsilon and M-3-mAChR. However, blockade of M-3-mAChR by 4-diphenylacetoxy-N-methylpiperidine methiodide (an M-3 receptor antagonist, administered at 20 min before occlusion) completely inhibited the effect of choline on the expression of PKC-epsilon. We conclude that the translocation of PKC-epsilon is required for the phosphorylation of M-3-mAChR; moreover, increased PKC-epsilon activity is associated with M-3-mAChR during MI. This reciprocal regulation is likely to play a role in heart signal transduction during ischemia between ventricular myocytes.