Differential effects of ethanol on serum GABAergic 3alpha,5alpha/3alpha,5beta neuroactive steroids in mice, rats, cynomolgus monkeys, and humans.

BACKGROUND: Acute ethanol administration increases plasma and brain levels of progesterone and deoxycorticosterone-derived neuroactive steroids (3alpha,5alpha)-3-hydroxypregnan-20-one (3alpha,5alpha-THP) and (3alpha,5alpha)-3,21-dihydroxypregnan-20-one (3alpha,5alpha-THDOC) in rats. However, little is known about ethanol effects on GABAergic neuroactive steroids in mice, nonhuman primates, or humans. We investigated the effects of ethanol on plasma levels of 3alpha,5alpha- and 3alpha,5beta-reduced GABAergic neuroactive steroids derived from progesterone, deoxycorticosterone, dehydroepiandrosterone, and testosterone using gas chromatography-mass spectrometry. METHODS: Serum levels of GABAergic neuroactive steroids and pregnenolone were measured in male rats, C57BL/6J and DBA/2J mice, cynomolgus monkeys, and humans following ethanol administration. Rats and mice were injected with ethanol (0.8 to 2.0 g/kg), cynomolgus monkeys received ethanol (1.5 g/kg) intragastrically, and healthy men consumed a beverage containing 0.8 g/kg ethanol. Steroids were measured after 60 minutes in all species and also after 120 minutes in monkeys and humans. RESULTS: Ethanol administration to rats increased levels of 3alpha,5alpha-THP, 3alpha,5alpha-THDOC, and pregnenolone at the doses of 1.5 g/kg (+228, +134, and +860%, respectively, p < 0.001) and 2.0 g/kg (+399, +174, and +1125%, respectively, p < 0.001), but not at the dose of 0.8 g/kg. Ethanol did not alter levels of the other neuroactive steroids. In contrast, C57BL/6J mice exhibited a 27% decrease in serum 3alpha,5alpha-THP levels (p < 0.01), while DBA/2J mice showed no significant effect of ethanol, although both mouse strains exhibited substantial increases in precursor steroids. Ethanol did not alter any of the neuroactive steroids in cynomolgus monkeys at doses comparable to those studied in rats. Finally, no effect of ethanol (0.8 g/kg) was observed in men. CONCLUSIONS: These studies show clear species differences among rats, mice, and cynomolgus monkeys in the effects of ethanol administration on circulating neuroactive steroids. Rats are unique in their pronounced elevation of GABAergic neuroactive steroids, while this effect was not observed in mice or cynomolgus monkeys at comparable ethanol doses.

Differential hypothalamic-pituitary-adrenal activation of the neuroactive steroids pregnenolone sulfate and deoxycorticosterone in healthy controls and alcohol-dependent subjects.

Ethanol and the neuroactive steroids have interactive neuropharmacological effects and chronic ethanol administration blunts the ethanol-induced increase in neuroactive steroid levels in rodent plasma and brain. Few studies have explored neuroactive steroid regulation in alcohol-dependent human subjects. In fact, the regulation of adrenal neuroactive steroids has not been well defined in healthy controls. We thus explored the regulation of two neuroactive steroids, pregnenolone sulfate (PREG-S) and deoxycorticosterone, by pharmacological challenges to the hypothalamic-pituitary-adrenal (HPA) axis in healthy controls and 1-month abstinent alcohol-dependent patients with co-occurring nicotine dependence. Plasma levels of PREG-S and deoxycorticosterone were measured by radioimmunoassay in controls and alcohol-dependent patients after challenges of naloxone, ovine corticotrophin releasing hormone (oCRH), dexamethasone, cosyntropin, and cosyntropin following high-dose dexamethasone. In addition, basal diurnal measures of both hormones were obtained. PREG-S plasma levels in healthy controls were increased by cosyntropin challenge (with and without dexamethasone pretreatment) and decreased by dexamethasone challenge. However, PREG-S concentrations were not altered by naloxone or oCRH challenges, suggesting that PREG-S is not solely regulated by hypothalamic or pituitary stimulation. Deoxycorticosterone, in contrast, is regulated by HPA challenge stimulation in a manner similar to cortisol. Alcohol-dependent patients had a blunted PREG-S response to cosyntropin (with and without dexamethasone pretreatment). Furthermore, the time to peak deoxycorticosterone response following oCRH was delayed in alcohol-dependent patients compared to controls. These results indicate that plasma PREG-S and deoxycorticosterone levels are differentially regulated by HPA axis modulation in human plasma. Further, alcohol-dependent patients show a blunted PREG-S response to adrenal stimulation and a delayed deoxycorticosterone response to oCRH challenge.

Antagonism of neurosteroid modulation of native gamma-aminobutyric acid receptors by (3alpha,5alpha)-17-phenylandrost-16-en-3-ol.

Endogenous pregnane neurosteroids are allosteric modulators at gamma-aminobutyric acid type-A (GABAA) receptors at nanomolar concentrations. There is direct evidence for multiple distinct neurosteroid binding sites on GABAA receptors, dependent upon subunit composition and stoichiometry. This view is supported by the biphasic kinetics of various neuroactive steroids, enantioselectivity of some neurosteroids, selective mutation studies of recombinantly expressed receptors and the selectivity of the neurosteroid antagonist (3alpha,5alpha)-17-phenylandrost-16-en-3-ol (17PA) on 5alpha-pregnane steroid effects on recombinant GABAA receptors expressed in Xenopus oocytes and native receptors in dissociated neurons. However, it is unclear whether this antagonist action is present in a mature mammalian system. The present study evaluated the antagonist activity of 17PA on neurosteroid agonists both in vivo and in vitro by examining the effects of 17PA on 5alpha-pregnane-induced sedation in rats, native mature GABAA receptor ion channels utilizing the chloride flux assay and further studies in recombinant alpha1beta2gamma2 receptors. The data show that 17PA preferentially inhibits 3alpha,5alpha-THP vs. alphaxalone in vivo, preferentially inhibits 3alpha,5alpha-THDOC vs. alphaxalone potentiation of GABA-mediated Cl- uptake in adult cerebral cortical synaptoneurosomes, but shows no specificity for 3alpha,5alpha-THDOC vs. alphaxalone in recombinant alpha1beta2gamma2 receptors. These data provide further evidence of the specificity of 17PA and the heterogeneity of neurosteroid recognition sites on GABAA receptors in the CNS.

Ethnic differences in allopregnanolone concentrations in women during rest and following mental stress.

The neuroactive steroid allopregnanolone (ALLO) is stress sensitive, negatively modulates the HPA axis, and has been implicated in mood disorders. We examined ethnic differences in plasma ALLO at rest and following mental stress in African American (AA) men (n = 21) and women (n = 24) and non-Hispanic White men (n = 24) and women (n = 25). Overall, AA women had lower ALLO concentrations than non-Hispanic White women (p < .05), especially following mental stress (p < .01). Only 20% of AA women showed the expected stress-induced increase in ALLO compared with 59% of non-Hispanic White women (p < .01). No ethnic differences were seen in men. For both ethnic groups, poststress ALLO was negatively correlated with poststress cortisol (p < .05). Results are interpreted to reflect dysregulation in ALLO mechanisms in AA women and may have implications for ethnic differences in mood disorders.