Isolation of nerve endings from the posterior pituitary gland. Electron microscopy of fractions obtained by centrifugation

Abstract

Bovine posterior pituitary glands were homogenized in 10 per cent sucrose and fractionated by differential centrifugation. The following centrifugation procedure resulted in the most satisfactory separation: 1000 g for 15 minutes-nuclei, connective tissue, basement membranes with associated endothelium, giant nerve endings, and whole pituicytes; 4200 g for 15 minutes-free nerve endings, including Herring bodies; 17,000 g for 15 minutes-mitochondria; 68,000 g for 15 minutes-neurosecretory granules. Electron microscopic examination was carried out on whole tissue and on the isolated fractions. Isolated nerve endings were examined also by negative staining techniques. Isolated nerve endings retain an apparently normal complement of mitochondria, neurosecretory granules, and microvesicles ("synaptic" vesicles). The free nerve endings closely resemble those observed in sections of intact posterior pituitary tissue. Free microvesicles were not observed in any of the fractions isolated and apparently sediment at centrifugal forces higher than those employed in this study.