Quantification of ten neuroactive steroids in plasma in Withdrawal Seizure-Prone and -Resistant mice during chronic ethanol withdrawal

Rationale The rapid membrane actions of neuroactive steroids, particularly via an enhancement of γ-aminobutyric acid A receptors (GABA A Rs), participate in the regulation of central nervous system excitability. Prior evidence suggests an inverse relationship between endogenous GABAergic neuroactive steroid levels and behavioral changes in excitability during ethanol withdrawal. Objectives Previously, we found that ethanol withdrawal significantly decreased plasma allopregnanolone (ALLO) levels, a potent GABAergic neuroactive steroid, and decreased GABA A R sensitivity to ALLO in Withdrawal Seizure-Prone (WSP) but not in Withdrawal Seizure-Resistant (WSR) mice. However, the effect of ethanol withdrawal on levels of other endogenous GABA A R-active steroids is not known. Methods After validation of a gas chromatography-mass spectrometry method for the simultaneous quantification of ten neuroactive steroids, we analyzed plasma from control male WSP-1 and WSR-1 mice and during ethanol withdrawal. Results We quantified levels of nine neuroactive steroids in WSP-1 and WSR-1 plasma; levels of pregnanolone were not detectable. Basal levels of five neuroactive steroids were higher in WSR-1 versus WSP-1 mice. Ethanol withdrawal significantly suppressed five neuroactive steroids in WSP-1 and WSR-1 mice, including ALLO. Conclusions Due to lower basal levels of some GABA A R-active steroids in WSP-1 mice, a withdrawal-induced decrease in WSP-1 mice may have a greater physiological consequence than a similar decrease in WSR-1 mice. Because WSP-1 mice also exhibit a reduction in GABA A R sensitivity to neuroactive steroids during withdrawal, it is possible that the combined decrease in neuroactive steroids and GABA A R sensitivity during ethanol withdrawal in WSP-1 mice represents a neurochemical substrate for severe ethanol withdrawal.