Secretory Membranes of the Rat Parotid Gland: Preparation and Comparative Characterization

This chapter focuses on the secretory membranes of the rat parotid gland. The release of secretory products by exocytosis involves the specific interaction of secretion granule and apical plasma membranes, the focal rearrangement of bilayer structures in regions of contact, and eventual coalescence of the two membranes with direct extracellular deposition of secretory products. The chapter describes the preparation of a plasma membrane fraction containing the apical domain. The isolation procedure could partially preserve morphologically recognizable regions constituting the apical surface. Two important features of the procedure developed are (1) it selects for large sheets of membrane, and (2) hypoosmotic media are used at the outset, as in the case of plasma membrane purification from rat liver, so that osmotically sensitive organelles are either damaged or disrupted during homogenization. Because morphological appearance constitutes an important initial evaluation for the purity and integrity of isolated membranes containing distinct regions, aliquots of the fraction are fixed in aldehydes, postfixed in 1% osmium tetroxide, and prepared for microscopy according to standard procedures.