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

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Abstract

Rationale

The rapid membrane actions of neuroactive steroids, particularly via an enhancement of γ-aminobutyric acidA receptors (GABAARs), 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 GABAAR 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 GABAAR-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 GABAAR-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 GABAAR sensitivity to neuroactive steroids during withdrawal, it is possible that the combined decrease in neuroactive steroids and GABAAR sensitivity during ethanol withdrawal in WSP-1 mice represents a neurochemical substrate for severe ethanol withdrawal.

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Acknowledgments

We are indebted to Dr. Leslie Morrow and her laboratory, especially Mr. Todd O’Buckley, for their invaluable guidance and assistance at all stages of the GC-MS assay development and validation. We also are extremely grateful to Dr. Robert Purdy for his help in the early stages of the assay development, his willingness to share his comprehensive knowledge of neuroactive steroids, as well as his collaborative spirit, wisdom, and humor. He was a true pioneer in the field of neuroactive steroids, who will be missed. We also thank the Alcohol Dependence Core of the Portland Alcohol Research Center, and in particular Mr. Larry Huang, for assistance with the induction of physical dependence. Finally, we acknowledge that all experiments complied with NIH guidelines. This work was supported by NIH RO1 grant AA012439 (DAF and DJR, MPIs), P60 AA010760 (JCC), R24 AA020245 (JCC), and grants and resources from the Department of Veterans Affairs (DAF, JCC) and the Department of Defense (JCC). MMF was supported by KO1 AA016849; DKC was supported by T32 AA007468; and MJR was supported by F31 AA020716.

Conflict of interest

The authors have no conflict of interest to report. Authors have full control of all primary data and agree to allow the journal to review the data if requested.

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Author notes

  1. David J. Rossi

    Present address: Department of Integrative Physiology and Neuroscience, Washington State University, Pullman, WA, 99164-7620, USA

Authors and Affiliations

  1. Department of Behavioral Neuroscience, Oregon Health and Science University, Portland, OR, USA

    Christopher Snelling, Michelle A. Tanchuck-Nipper, Matthew M. Ford, Jeremiah P. Jensen, Debra K. Cozzoli, Marcia J. Ramaker, Melinda Helms, John C. Crabbe, David J. Rossi & Deborah A. Finn

  2. Division of Neuroscience, Oregon National Primate Research Center, Oregon Health and Science University, Beaverton, OR, USA

    Matthew M. Ford

  3. Department of Veterans Affairs Medical Center, Portland Alcohol Research Center, 3710 US Veterans Hospital Road (R&D-49), Portland, OR, 97239, USA

    John C. Crabbe & Deborah A. Finn

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  1. Christopher Snelling
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  2. Michelle A. Tanchuck-Nipper
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Correspondence to Deborah A. Finn.

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Snelling, C., Tanchuck-Nipper, M.A., Ford, M.M. et al. Quantification of ten neuroactive steroids in plasma in Withdrawal Seizure-Prone and -Resistant mice during chronic ethanol withdrawal. Psychopharmacology 231, 3401–3414 (2014). https://doi.org/10.1007/s00213-014-3618-y

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