INVOLVEMENT OF DIFFERENT CA2-FREE MEDIUM - LACK OF EFFECT OF PKC INHIBITION( POOLS DURING THE CANINE BRONCHIAL SUSTAINED CONTRACTION IN CA2+)

We evaluated the role of protein kinase C (PKC) in the sustained bronc hial contraction (SBC) induced by carbachol (Cch) or histamine in a Ca 2+-free medium and the possibility that each agonist uses a different Ca2+ store for this response. We studied third-order bronchi and airwa y smooth muscle (ASM) from first-order bronchi dissected free of carti lage and epithelium. Bronchial and ASM responsiveness to Cch or histam ine were evaluated in Krebs solution (2.5 mM Ca2+) and in Ca2+-free me dium. Cch and histamine induced an SEC in bronchial tissues in Ca2+-fr ee medium. In ASM each agonist produced a transient contraction, but t he response to histamine was much smaller. Cch induced a concentration -dependent accumulation of inositol phosphates (IPs) in both bronchi a nd ASM; however, histamine did not induce significant accumulation of IPs. Repeated exposure to histamine in bronchial rings abolished contr actile responses in Ca2+-free media, but Cch added afterwards still pr oduced a sustained contraction. This response was blocked when bronchi al tissues were preincubated with 10 mu M cyclopiazonic acid (CPA). Br ief incubation of these preparations with a high EGTA concentration (1 mM) abolished the histamine-induced SEC. The SEC induced by Cch or hi stamine in Ca2+-free medium was not affected by the preincubation of t he tissues with calphostin C, chelerythrine or staurosporine. We concl uded that Cch mobilizes Ca2+ from two different sources during the SEC in Ca2+-free medium: from a CPA-sensitive one from sarcoplasmic retic ulum (SR) and from a putative extracellular membrane Ca2+ pool sensiti ve to 1 mM EGTA, and neither process involved PKC activation. Histamin e appeared to utilize the extracellular membrane pool only.