Targeting of tissue plasminogen activator into the regulated secretory pathway of neuroendocrine cells.

摘要: Abstract Plasma levels of tissue plasminogen activator (tPA) increase rapidly in response to specific vasoactive agents, trauma, and neural stimulation. This has been attributed acute release tPA from stored pools within the vascular endothelium catecholamine storage vesicles chromaffin cells. We have tested directly whether can be sorted into regulated secretory pathway using murine pituitary-derived neuroendocrine cell line AtT-20 transfected with cDNA. Clones cells expressing were isolated, targeting was demonstrated by (1) stimulation secretion 8-bromo-cAMP, secretagogue which promotes dense granule contents; (2) colocalization ACTH, an endogenous protein that is core granules; (3) retention newly synthesized for prolonged periods time. Laser scanning confocal microscopy analysis immunostained antibodies ACTH showed at tips neuritic processes under cytoplasmic membrane, a region where granules are known migrate after maturation. Treatment 5 mM 8-bromo-cAMP 30 min resulted 2.41±0.36-fold secretion. Both magnitude stimulatory effect fraction intracellular released same regardless expression level various clones. Pulse-chase experiments portion retained least 4 h culture medium 8-bromo-cAMP. These studies indicate tPA, appropriate conditions, targeted later cellular stimuli.