MORPHOLOGY AND FUNCTION OF ROOSTER EFFERENT DUCTULE EPITHELIAL-CELLS IN CULTURE

Regulation of the excurrent ducts of the testis is not well understood , particularly in avian species. To investigate the role of steroid ho rmones in the male reproductive tract, we developed a primary cell cul ture of epithelia isolated from rooster ductuli efferentes (efferent d uctules). Efferent ductules of the avian testis comprise 77% of the ep ididymal region and form a mass of tubules containing a heavily folded epithelium enmeshed in connective tissue. The epididymal region was s eparated by microdissection and small epithelial plaques isolated by s erial digestion with collagenase, elastase and repeated pipetting, Iso lated cell plaques were cultured in a bicameral chamber on Millicell-C M inserts coated with two layers of basement membrane matrix, consisti ng primarily of laminin and Types I and IV collagen. Active ciliary be at was observed before plating and this activity was maintained for 14 days in culture. Cell plaques attached within 24 h and outgrowths for med a confluent monolayer by 5-6 days. The epithelial nature of cultur ed cells was demonstrated by immunocytochemical staining for cytokerat in. Light and electron microscopy confirmed that morphology and polari ty of the original epithelial cells were maintained in culture. Cultur ed efferent ductal epithelium was cuboidal in shape and maintained man y of the cytoplasmic organelles typical of these cells in vivo. The up take of cationic ferritin indicated the endocytotic activity of these cultured cells was maintained. Estrogen receptor mRNA expression was m aintained in cultured cells. These data demonstrate avian efferent duc tal epithelium can be isolated and grown in defined culture medium for the purpose of determining the role of hormones and other factors in regulating the function of the epididymal region in the bird.