In the salmonid ovary, luteinizing hormone (LH) is the major gonadotropic hormone stimulating the production of steroids during the periovulatory period and its effects are mediated by the cAMP-dependent protein kinase (PKA) signaling pathway. We have previously shown that the in vitro steroidogenic activity of LH in the salmonid ovary is inhibited by insulin-like growth factor I (IGF-I) which, like insulin, has specific receptors in both theca and granulosa layers. In the present study, we have investigated the modulatory effects of insulin on salmon LH (sLH)-stimulated steroid production in preovulatory theca layers of brown trout (Salmo trutta) and the effects of both insulin and IGF-I on the sLH-stimulated cAMP/PKA signaling pathway. Our results show that insulin, like IGF-I, blocked the stimulatory effects of sLH, dibutyryl cAMP and IBMX on testosterone (T) production but not those of sLH on cAMP production. Furthermore, insulin and IGF-I blocked the activation of PKA induced by sLH and these effects were correlated with changes in the total protein content of the catalytic (C) and type II regulatory (RII) subunits of PKA. Interestingly, insulin and IGF-I had different effects on total PKA subunit content since insulin potentiated the sLH-stimulated increase in RII subunit content whereas IGF-I blocked the sLH-stimulated increase in total C subunit content. The effects of insulin and IGF-I in trout theca layers appeared to be mediated by the mitogen-activated protein kinase (MAPK) signaling pathway because inhibition of extracellular signal-regulated kinase 1/2(ERK1/2) activity completely blocked the inhibitory effects of insulin and IGF-I on the sLH-stimulated production of T and because insulin and IGF-I increased the total protein content of ERK1/2 in trout theca layers. Therefore, our results suggest that insulin and IGF-I, probably through the MAPK pathway, block the action of sLH in trout theca layers by modulating the cAMP/PKA signaling pathway.