INVOLVEMENT OF PROTEIN-KINASE-C IN AGONIST-STIMULATED GOLDFISH OVULATION

The effects of two protein kinase C (PKC) inhibitors, calphostin C and staurosporine, on the in vitro ovulation of goldfish (Carassius aurat us) oocytes were investigated. Ovulation was stimulated by prostagland in (PG) F2alpha (PGF2alpha, 2.0 mug/ml), by sodium orthovanadate (0.1 mM), by a combination of the phorbol ester phorbol 12-myristate-13-ace tate (PMA, 0.1 mug/ml) and calcium ionophore A23187 (0.05 mug/ml), by thapsigargin (0.2 mug/ml), and by elevated pH (8.1). In addition, the effects Of these inhibitors on the PKC activity of the goldfish follic le wall was determined by use of a specific peptide substrate phosphor ylation assay. At 0.1 muM, staurosporine significantly blocked ovulati on induced by all agents. However, at lower (0.01 muM) levels it block ed only PMA/A23187-induced ovulation. In contrast, calphostin signific antly blocked only PMA/A23187-induced ovulation, although there was a decrease in pH-induced ovulation at lower calphostin concentrations. B oth calphostin and staurosporine blocked follicular PKC activity at le vels that were inhibitory to ovulation. In addition, staurosporine sig nificantly blocked PKC activity at levels even lower than those needed to block ovulation. The combined results suggest that orthovanadate, PGF2alpha, and thapsigargin do not require PKC activation for the indu ction of ovulation, whereas PMA/A23187 does.