Isolation and characterization of secretory vesicles from bovine neurohypophyses.
A procedure is described for the isolation of secretory vesicles from bovine neurohypophyses by differential centrifugation followed by density gradient centrifugation on iso-osmolal gradients of percoll/sucrose. Only negligible contamination of the secretory vesicle fraction with markers for mitochondria, microsomes and plasma membranes could be detected. The amount of Ca2-ATPase in the isolated neurohypophysial secretory vesicles was of the same low order of magnitude as that of (Na, K)-ATPase. Thin-section electromicrographs confirmed the high purity of the isolated secretory vesicle fractions, In freeze-fracture electronmicrographs, vesicle fusion was demonstrated after incubation with Ca2. As shown in dodecyl sulfate-gel electrophoresis and subsequent autoradiography secretory vesicles exhibited an endogenous phosphorylation activity. The secretory vesicles contained an average of 23.1 microgram vasopressin/mg of protein. On incubation in media differing in ionic strength, pH and Ca2 concentration the vesicles were stable for at least 1 h.
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000032:analysis; D002417:Cattle; D002498:Centrifugation / Q000379:methods; D002499:Centrifugation, Density Gradient / Q000379:methods; D003594:Cytoplasmic Granules / Q000032:analysis / Q000201:enzymology / Q000648:ultrastructure; D008854:Microscopy, Electron; D009481:Neurophysins / Q000032:analysis; D010904:Pituitary Gland, Posterior / Q000032:analysis / Q000648:ultrastructure*; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis
D009481:Neurophysins; D000252:Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase