Caveolae from canine airway smooth muscle contain the necessary componentsfor a role in Ca2+ handling

To explain that bronchial smooth muscle undergoes sustained agonist-induced contractions in a Ca2+-free medium, we hypothesized that caveolae in the p lasma membrane (PM) contain protected Ca2+. We isolated caveolae from canin e tracheal smooth muscle by detergent treatment of PM-derived microsomes. D etergent-resistant membranes were enriched in caveolin-1, a specific marker for caveolae as well as for L-type Ca2+ channels and Ca2+ binding proteins (calsequestrin and calreticulin) as determined by Western blotting. Also, the PM Ca2+ pump was present but not connexin 43 (a noncaveolae PM protein) , the sarcoplasmic reticulum (SR) Ca2+ pump, or the type 1 inositol 1,4,5-t risphosphate receptor, supporting the idea that SR-derived membranes were n ot present. Antibodies to caveolin coimmunoprecipitated caveolin with calse questrin or calreticulin. Thus some of the cellular calsequestrin and calre ticulin associated with caveolin on the cytoplasmic face of each caveola. I mmunohistochemistry of tracheal smooth muscle crysosections confirmed the l ocalization of caveolin and the PM Ca2+ pump to the cell periphery, whereas the SR Ca2+ pump was located deeper in the cell. The presence of L-type Ca 2+ channels, the PM Ca2+ pump, and the Ca2+ binding proteins calsequestrin and calreticulin in caveolin-enriched membranes supports caveola involvemen t in airway smooth muscle Ca2+ handling.