In female zebrafish ovulation could be induced by male holding water, testis homogenates, and testis fractions containing steroid glucuronides. Deglucuronidation of these fractions led to a loss of ovulation-inducing potency, indicating steroid giucuronides as ovulation inducers. The chemical substances were perceived by the recipient females by means of olfaction. Incubation experiments showed the capacity of the testes to synthesize various C,, and C,, steroids and seven different steroid glucuronides, i.e., 17ct,20P-dihydroxy-4-pregnen-3-one-, testosterone-, androsterone-, epiandrosterone-, 5u-androstane-3a,17@diol-, and 5a-androstane-3l3,17P-diol glucuronide. GC-MS analysis showed the presence of glucuronides of 5a-androstane-3a,l7@dioland cholesterol in male holding water, the latter probably originating from the liver. These compounds may be among the steroid glucuronides functioning as ovulation-inducing pheromones. 0 1987 Academic Press. Inc.