Multiparameter morphology as a means of specifically identifying hormone stimulated adrenal cortical tumor cells (Y-1) and granulosa cells in coculture.

In order to test for biologically valid, direct communication between mouse adrenal tumor and porcine ovarian granulosa cells a method was needed for distinguishing the two cell types in culture. To this end granulosa cells were cultured for 24-48 hours after which they were challenged with rhodamine conjugated latex beads for 30 minutes to 24 hours in monolayer culture. When phagocytosis was allowed to occur for longer than one hour the phagocytic vesicles containing the conjugated beads appeared to fuse with each other and with digestive vacuoles. The resultant vesicles contained many beads and were located within the cell body close to the nucleus. At shorter incubation periods, beads were observed predominantly in the periphery of the cell. Y-1 adrenal cortical tumor cells failed to ingest beads and could be distinguished from the bead labeled granulosa cell in coculture with scanning or thin section electron microscopic as well as fluorescence microscopic techniques. Gap junctions were observed between heterotypic cell pairs; however, no evidence of cell fusion was found. ACTH stimulated granulosa: Y-1 adrenal cocultures initiated cAMP-dependent kinase dissociation in the Y-1 cells and in contacting granulosa cells identified positively by the presence of the beads. We have used the bead labeling technique and a procedure for following the kinetics of cAMP-dependent protein kinase dissociation in order to follow the specific hormonal stimulation of these heterotypic cells in coculture.(ABSTRACT TRUNCATED AT 250 WORDS)