LOCALIZATION OF ATRIAL-NATRIURETIC-PEPTIDE IN CAVEOLAE OF IN-SITU ATRIAL MYOCYTES

The plasma membrane-associated non-clathrin-coated vesicles called cav eolae are multifunctional organelles thought to be implicated in the s equestration and transport of small molecules (potocytosis) as well as in the binding of Ca2+ ions, signal transduction, and processing of h ormonal and mechanosensitive signals. We have previously suggested tha t the apparent contiguity of caveolar and atrial granule membranes obs erved in electron micrographs of in situ mouse atrial myocytes might r eflect externalization of atrial natriuretic peptide through caveolae. Using Tokuyasu's classic technique, we now show by immunoelectron mic roscopy of glutaraldehyde-fixed and cryosectioned mouse and rat atria that antibody against atrial natriuretic peptide prohormone is present within caveolae of in situ atrial myocytes. We confirm this intracave olar localization by stereoimaging colloidal gold-labeled antibody to the prohormone in electron micrographs of glutaraldehyde/osmium tetrox ide-fixed positively stained atrial thin sections. Because profiles of caveolae were rarely immunolabeled with antibody against atrial pepti de unless there was a profile of an immunolabeled atrial granule nearb y in the subjacent cytoplasm, we concluded that the intracaveolar horm one was derived predominantly from a direct interaction of atrial gran ules with caveolae. Perturbations that markedly increase the rate of n atriuretic peptide secretion via the regulated pathway, including atri al stretch, contractions, and increased external Ca2+ concentration, f ailed to alter caveolar immunostaining. These results suggest that atr ial peptide can pass from atrial granules into caveolae by transiently open pathways between the interiors of granules and caveolae. The res ults are interpreted as suggesting the presence of a second pathway fo r externalization of atrial natriuretic peptide through caveolae in ad dition to the classic pathway for regulated atrial peptide secretion a t noncaveolar plasmalemma.