UP-REGULATION OF MT1 AND MT2 RECEPTORS BY IN VITRO MELATONIN AND MODULATION OF ALPHA-TUBULIN AND AROMATASE P450 EXPRESSION IN HUMAN GRANULOSA-LUTEIN CELLS

Ovarian granulosa-lutein cells (GLCs) prove to be useful model to study receptors and signalling mechanism involved in steroidogenesis and differentiation plasticity linked to follicular growth and oocyte quality. Reports of high levels of melatonin in pre-ovulatory follicular fluids together with demonstration of specific melatonin receptors in follicular cells let us investigate possible targets of the hormone in GLC aspirated from follicular fluid. We demonstrated by confocal immunofluorescence specific expression of both MT1 and MT2 receptors (MR) which was up-regulated by melatonin treatment and confirmed also by Western blotting. By dual immunofluorescence labelling we were able to show colocalization of the signal for alpha-tubulin and MT1 in the perinuclear cytoplasmic area of GLC, while the MT2 signal appeared weaker around the nuclei and turned diffuse cytoplasmic after 48 h of melatonin treatment. Up-regulation of MR was observed along with stimulation of P450 aromatase protein expression which points out at the significance of melatonin in processes regulating follicular development.