The aim of our in vitro experiments was to study the effects of EGF on rabbit ovarian cells, as well as the possible mechanisms of these effects. The influence of EGF on steroidogenesis, proliferation, cyclic nucleotides and MAP-kinase in rabbit granulosa cells were studied. Results of RIA showed, that EGF stimulated the release of progesterone (1-100 ng/ml), cAMP (at 100 ng/ml), cGMP (1-100 ng/ml). EGF effect on estradiol output was biphasic: at dose 1 ng/ml it inhibited, whilst at 100 ng/ml it strongly increased estradiol secretion. Immunocytochemical study demonstrated an EGF-induced (10 ng/ml) increase in the proportion of cells revealing proliferating cell nuclear antigen (41% vs 24.7% in control, p < 0.01). EGF (10 ng/ml) increased the proportion of cells with immunoreactivity to ERK-1 (more than two-fold) and ERK-3 (three-fold) members of the MAP-kinase family. Moreover, EGF induced the translocation of ERK-1 to the nucleus, whilst preferentially cytoplasmic localization of ERK-3 was not changed after EGF addition. This can indicate regulation of ERK-1 and -3 by EGF, as well as differential patterns of ERK-1 and ERK-3 expression in response to EGF in cultured granulosa cells. - These results indicate that EGF can be a stimulator of proliferation, steroidogenesis and cyclic nucleotide release by rabbit granulosa cells. Stimulation of cAMP and cGMP release, and activation of ERK-related MAP kinase in granulosa cells after EGF addition indicates the involvement of these intracellular messengers in mediating the EGF action on the ovary.