DIFFERENT SITES OF ACTION OF ARACHIDONIC-ACID ON STEROIDOGENESIS IN RAT LEYDIG-CELLS

The present study in purified rat Leydig cells shows that arachidonic acid may act as an intratesticular factor regulating LH-mediated testi cular steroidogenesis. Arachidonic acid decreased, in a dose-dependent manner. the LH-stimulated cAMP and testosterone levels, over 2 h incu bation. Incubation of Leydig cells with arachidonic acid did not modif y I-125-hCG binding to the cells as compared to control, showing that the action of arachidonic acid is not related to a decrease of hCG bin ding to the cells. Forskolin-stimulated cAMP and testosterone producti on were inhibited by 51.65 and 70.9%, respectively, in the presence of arachidonic acid (100 mu M), although the ED(50) for the diterpene wa s not changed. When isobutyl-methyl-xanthine was added to the incubati on medium, the same percentage of inhibition was found indicating that arachidonic acid inhibition of cAMP production is not due to stimulat ion of Leydig cell phosphodiesterase activity. Pretreatment of the cel ls with pertussis toxin, to inactivate Gi, was also without effect on arachidonic acid inhibition of LH-stimulated cAMP production, but pert ussis toxin abolished the inhibitory effects of arachidonic acid when adenylate cyclase was stimulated with forskolin. However, arachidonic acid addition resulted in inhibition of LH- and forskolin-stimulaled t estosterone production, even if the cells were pretreated with pertuss is toxin. It can be concluded that: (1) The inhibitory effect of arach idonic acid is neither due to a decrease of hCG binding to Leydig cell s nor to a stimulation of cell phosphodiesterase activity; (2) arachid onic acid modulates cAMP production at two different levels, either by activation of Gi protein and by inhibition of Gs protein ol adenylate cyclase; (3) the effect of arachidonic acid on steroidogenesis is als o beyond cAMP formation.