Dissociation of avian salt gland: separation procedures and characterization of dissociated cells

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Abstract
A procedure for dissociation of the nasal salt glands of the domestic duck, Anas platyrhynchos, into suspensions of individual cells has been developed. This technique employs enzymatic digestion with collagenase, hyaluronidase, and chymotrypsin; divalent cation chelation with EDTA; and gentle mechanical dispersion. Average cellular yields of 39 and 26% based on DNA recovered were obtained from the glands of freshwater- and saline-adapted ducks, respectively. Epithelial secretory cells comprised 60-80% of the cell suspensions with the remainder of the populations consisting of endothelial cells, fibroblasts, and blood cells. The dissociated cells were viable as judged by trypan blue exclusion (80-100%, maintenance of ultrastructural integrity, and retention of responsiveness to secretagogues and metabolic inhibitors. Methacholine chloride (0.5 mM) stimulated oxygen consumption by suspensions of both freshwater- and saline-adapted cells, whereas ouabain (0.05 mM) abolished the methacholine-stimulated respiratory response. These cell suspensions provide a promising system for the in vitro study of secretory mechanisms in the avian salt gland.