SEX-SPECIFIC AROMATIZATION OF TESTOSTERONE IN MOUSE HYPOTHALAMIC NEURONS

Conversion of androgens to oestrogens by neural aromatase during brain development appears to be a prerequisite for sexual differentiation of the mammalian central nervous system. In order to investigate the pre- and perinatal patterns of testosterone (T) aromatization in the male and female mouse brain, aromatase activity (AA) was measured in hypothalamic and cerebral homogenates of embryonic day (ED) 17 fetuses and neonates using an in vitro (H2O)-H-3 product formation microassay. In addition, AA was examined in gender-specific neuronal cell cultures prepared from ED 15 mouse cerebral hemisphere and hypthalamus at 3 and 6 days in vitro (DIV), and this was compared with enzyme activities in homogenates. The aromatase has also been evaluated in glial-enriched cultures from ED 20 mouse hypothalamus and cortex as well as in ED 15 cultures treated with the neurotoxin kainic acid in order to localize AA to neurons and/or glial cells. Significant sex differencees in AA were observed in hypothalamic tissue homogenates as early as ED 17, becoming even more distinct in neonates, AA being always higher in males compared to females. Similar AA was also found in cells from both sexes from cultured ED 15 hypothalamus after 3 DIV. However, significantly higher AA was observed after 6 DIV in ED 15 male hypothalamic cultures compared to female. ED 20 glial-enriched hypothalamic culutures (purity >95%) from both brain regions exhibited very low AA after 6 DIV, and no sex differences were found. After treatment of ED 15 hypothalamic cultures with kainic acid, a 70-80% decrease in AA was observed in both sexes compared to non-treated cultures and sex differences were no longer present. Kinetic studies performed on aromatase in male and female hypothalamic cells from ED 15 cultures after 6 DIV, from ED 17 fetuses, and from neonatal homogenates with T as substrate indicate similar T binding affinities in cultures as in embryonic and neonatal homogenates from both genders, with an apparent K-m of similar to 40 nM. In cortex, from either ED 15 cultures, ED 20 cultures, ED 17 fetuses or neonates, AA was low and sex differences were not observed. Taken together, our results demonstrate: (1) sex differences in hypothalamic AA during pre- and perinatal development in the mouse with a higher capacity for oestrogen formation in male than in female; (2) no sex differences in AA from cortical tissues; (3) no sexual or developmental differences in hypothalamic aromatase T binding affinity; (4) aromatase is neuronal rather than astroglial, and (5) sex differences in AA are consequentially restricted to neurons.