A phase-contrast and immunofluorescence study of adrenal medullary chromaffin cells in culture: Neurite formation, actin and chromaffin granule distribution

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Abstract
Immunofluorescence studies of bovine chromaffin cells in culture with specific antibodies against dopamine-gb-hydroxylase gave a distinct punctate pattern of labelling, reflecting the distribution of chromaffin granules. There was strong staining of cell extensions and growth cones. Linear arrays of fluorescent dots were observed, suggesting an association of granules with a filamentous cytoskeleton. Labelling of neuritic processes was periodic, perhaps indicative of a packaging of secretory granules. Chromaffin cells stained strongly with specific anti-actin antisera. Fine filament bundles were observed, and also diffuse staining, some punctate labelling and staining of the plasma membrane or sub-membranous cytoplasm. Growth cones and non-terminal cytoplasmic varicosities contained significant amounts of actin. Colchicine (5×10-5M) caused retraction of neuritic extensions and formation of lateral growth cones. Cytochalasin (10μg/ml) caused ballooning of terminal growth cones and non-terminal cytoplasmic varicosities. Phalloidin (10-4M) stimulated microspike formation. The results are discussed in terms of the role of the cytoskeleton in growth cone formation, cell-substratum contacts and the transport of chromaffin granules.