Formation of ecdysteroids by Y-organs of the crab, Menippe mercenaria. II. Incorporation of cholesterol into 7-dehydrocholesterol and secretion products in vitro.

The conversion in vitro of cholesterol (Ch) to nonpolar metabolites and ecdysteroids was studied in Y-organs of the xanthid crab, Menippe mercenaria. In one set of experiments, Y-organs were prelabeled in vivo by injecting crabs with 100 microCi of [3H]Ch, and halved glands were then incubated for 24 and 48 hr in the presence of unlabeled Ch. In another set, unlabeled Y-organs were incubated in standard medium containing 10 microCi/ml of [3H]Ch. Both polar and nonpolar metabolites were surveyed by HPLC. The early metabolite, 7-dehydrocholesterol (7-dhCh), was the only labeled derivative of Ch detectable in Y-organ tissue after incubation; preincubation amounts of 7-dhCh were higher in glands from de-eyestalked crabs vs glands from intact crabs, and labeling was an order of magnitude higher in glands incubated with labeled Ch vs those prelabeled in vivo. Specific activity calculations indicate highly efficient conversion of 7-dhCh to ecdysteroid secretions. Analyses of the incubation media revealed two polar secretory products, synthesized from Ch in vitro. These coeluted with the authentic standards, 3-dehydroecdysone and 25-deoxyecdysone. Secretion of 3-dehydroecdysone always exceeded that of 25-deoxyecdysone (ratio range, 1.9 to 14.4), in both de-eyestalked and intact crabs.