Aromatase activity in the brain of Medaka and its regulation by sex steroids.

摘要

The cytochrome P450aromatase (aromatase) catalyses the final step in the synthesis of all estrogens from androgens and thus is central to estrogen regulated biological functions. Our hypothesis was that the localization of brain aromatase activity is sexually dimorphic and sex hormones play a role in the regulation of aromatase activity. The teleost Oryzias latipes (Medaka) was our experimental animal. Brain aromatase activity was measured in the homogenates of brain sections from the three orthogonal planes using the tritiated water method. A three-dimensional computer representation of aromatase activity was built using those values and its overlay on a Medaka brain model permitted the localization of the highest levels of aromatase activity to a medial ventral region of the brain. The effects of sex steroids were investigated by measuring the activity of aromatase after in vivo exposure to estradiol or methyl-testosterone. Estradiol increased and methyl-testosterone decreased the activity of brain aromatase. Sexual dimorphism in brain aromatase activity was identified using coronal sections. In the brain of males the activity is higher in rostral areas containing the preoptic nuclei. In the brain of females the highest levels of activity are found in caudal areas containing hypothalamic periventricular nuclei. In vivo exposure of male Medaka to estradiol changed the localization of aromatase activity to a pattern that resembles that of the female. We established dose-response relationships and time-dependency for the effects of sex steroids on brain aromatase and characterize the kinetics of brain aromatase of Medaka.;In a parallel study of regulatory mechanisms, we investigated the role of the purinergic receptor P2X7/P2Z in the mediation of the toxic effects of the dinoflagellate Pfiesteria piscicida. Using fluorescent dyes in a digital imaging system we observed that the P2X 7/P2Z selective agonist BzATP and ATP elevated the cytosolic free calcium and induced permeability of to YO-PRO-1 in GH4C1 rat pituitary cells. The permeabilization of GH4C1 was inhibited by oxidizedATP. We then substituted the P2X7/P2Z agonists by a partially purified fraction of Pfiesteria toxic cultures. We found that GH4C1 cells have ionotropic purinergic receptors with pharmacologic and functional properties consistent with the P2X7 subtype and pPfTx mimics the kinetics of cell permeabilization by ATP.