THE EFFECT OF CA2-KINASE-C IN BOVINE TRACHEAL SMOOTH-MUSCLE( ON THE TRANSLOCATION OF PROTEIN)

The role of calcium in protein kinase C redistribution was studied in bovine tracheal smooth muscle preparations contracted by methacholine. Previous results have shown that, in the presence of normal extracell ular Ca2+, 10 mu M methacholine produced a sustained contraction and a sustained translocation of protein kinase C from the cytosol to the m embrane. In the present study, when tissues were preincubated in Ca2+- free buffer containing 1 mM EGTA, methacholine produced a rapid but tr ansient elevation in membrane-associated protein kinase C activity whi ch was detected at 30 s and had returned to basal within 20 min. The r edistribution of protein kinase C from the cytosol to the membrane ind uced by 1 mu M methacholine in normal Ca2+ was reversed by removal of the extracellular Ca2+ and addition of 2 mM EGTA during agonist stimul ation. Removal of the Ca2+ caused approximately 50% relaxation after 1 0 min. Verapamil (30 mu M) partially reversed the methacholine-induced protein kinase C redistribution and caused approximately 40% relaxati on after 15 min. Sodium nitroprusside (10 mu M) caused a rapid relaxat ion and complete reversal of the protein kinase C redistribution induc ed by methacholine. High K+ (60 mM) also induced a sustained contracti on and redistribution of protein kinase C from the cytosol to the memb rane. Suitable antagonists were added to the bathing medium to block t he effects of endogenous mediators which could be released by KCl-indu ced depolarization. Thus, translocation of protein kinase C is obtaine d in the absence of receptor activation. These observations indicate t hat alterations in [Ca2+](i) may be responsible for the changes in pro tein kinase C redistribution observed during agonist-induced contracti ons of bovine tracheal smooth muscle.