[Estrogen formation in the central nervous system and characteristics of aromatase of rat hypothalamus].
Estrogen formation from androst-4-ene-3,17-dione and its kinetics were studied using microsomes from rat hypothalamus. [4-14C] androst-4-ene-3,17-dione and a homogenate of rat hypothalamus were incubated in the presence of NADPH at 37 degrees C for 3 hrs. The estrogen fraction was extracted from the incubation mixture with ethyl acetate, purified by column chromatography on Sephadex LH-20 and Bond Elut C18, and separated into estrone and estradiol fractions by HPLC. In analysis of the trimethylsilyl (TMS) derivatives of each fraction by gas chromatography-mass spectrometry (GC-MS), the molecular ion peak of the estrone fraction appeared at m/z 344, within 2 amu of that for the TMS derivative of natural estrone. The retention time of the estrone fraction derivative was 11.6 min, the same as that of natural estrone. 14C-estrone was thus concluded to be biosynthesized from [4-14C]-androst-4-ene-3,17-dione in rat hypothalamus. The kinetics of the aromatase of rat hypothalamic tissue was studied by measuring 3H2O released from [1 beta-3H]-androst-4-ene-3,17-dione and estrone as the estrogen product by measured gas chromatography selected ion monitoring (GC-SIM). High correlation was found between 3H2O release and estrone measured by GC-SIM (r = 0.97). Aromatase activity was linear with respect to incubation time and quantity of tissue. Km and Vmax were 30.3 nM and 7.98 fmol estrogen/h/mg of wet tissue, respectively. 4-hydroxyandrostenedione (4-OH-A) suppressed the activity of aromatase in both rat hypothalamic and human placental tissue in a concentration-dependent manner. Polyclonal IgG to human placental aromatase also suppressed aromatase activity of human placental tissue, but only slightly suppressed that of rat hypothalamus. The molecular structure of aromatase in rat hypothalamus was thus concluded to differ from that in human placenta.
Takezawa
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Y
Department of Urology, Gunma University School of Medicine, Maebashi, Japan.
Fukabori
Y
Y
Honma
S
S
D000735:Androstenedione / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001141:Aromatase / Q000378:metabolism*; D047072:Aromatase Inhibitors; D046508:Culture Techniques; D004305:Dose-Response Relationship, Drug; D004967:Estrogens / Q000032:analysis / Q000096:biosynthesis*; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D007031:Hypothalamus / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D010920:Placenta / Q000201:enzymology; D051381:Rats; D011919:Rats, Inbred Strains
D047072:Aromatase Inhibitors; D004967:Estrogens; D000735:Androstenedione; D001141:Aromatase; C014594:formestane