The neurosteroid environment in the hippocampus exerts bi-directional effects on seizure susceptibility in mice

Abstract

The progesterone derivative allopregnanolone (ALLO) rapidly potentiates γ-aminobutyric acid A (GABA A ) receptor mediated inhibition. The present studies determined whether specific manipulation of neurosteroid levels in the hippocampus would alter seizure susceptibility in an animal model genetically susceptible to severe ethanol (EtOH) withdrawal, Withdrawal SeizureProne (WSP) mice. Male WSP mice were surgically implanted with bilateral guide cannulae aimed at the CA1 region of the hippocampus one week prior to measuring seizure susceptibility to the convulsant pentylenetetrazol (PTZ), given via timed tail vein infusion. Bilateral intra-hippocampal infusion of ALLO (0.1 g/side) was anticonvulsant, increasing the threshold dose of PTZ for onset to myoclonic twitch and face and forelimb clonus by 2-3 fold. In contrast, infusion of the 5α-reductase inhibitor finasteride (FIN; 2 g/side), which decreases endogenous ALLO levels, exhibited a proconvulsant effect. During withdrawal from chronic EtOH exposure, WSP mice were tolerant to the anticonvulsant effect of intra-hippocampal ALLO infusion, consistent with published results following systemic injection. Finally, administration of intra-hippocampal FIN given only during the development of physical dependence significantly increased EtOH withdrawal severity, measured by handling-induced convulsions. These findings are the first demonstration that bidirectional manipulation of hippocampal ALLO levels produces opposite behavioral consequences that are consistent with alterations in GABAergic inhibitory tone in drug naïve mice. Importantly, EtOH withdrawal rendered WSP mice less sensitive to ALLO's anticonvulsant effect and more sensitive to FIN's proconvulsant effect, suggesting an alteration in the sensitivity of hippocampal GABA A receptors in response to fluctuations in GABAergic neurosteroids during ethanol withdrawal.

Sex Differences in Ethanol’s Anxiolytic Effect and Chronic Ethanol Withdrawal Severity in Mice with a Null Mutation of the 5α-Reductase Type 1 Gene

“…Another potential explanation for the lack of genotype difference in chronic withdrawal is that the presence of 5α-reductase-2 in the KO mice compensated for any impact of Srd5a1 deletion on ethanol withdrawal severity. We have found that microinjection of finasteride into discrete brain regions, which would only inhibit Srd5a1 , was required to significantly increase chronic ethanol withdrawal severity (Gililland-Kaufman et al, 2008; Tanchuck et al, 2013), whereas systemic finasteride did not significantly alter chronic ethanol withdrawal severity in male and female C57BL/6J mice (Finn et al, 2004b). Future studies could pharmacologically inhibit the type 2 enzyme in the Srd5a1 mutants or test the double mutants that have been developed (mice lacking Srd5a1 and Srd5a2 ; Mahendroo et al, 2001) and determine the impact on chronic ethanol withdrawal severity.…”

“…The withdrawal-induced decrease in endogenous ALLO levels was accompanied by a significant reduction in activity and expression of 5α-reductase and 3α-HSD (Cagetti et al, 2004; Finn et al, 2004a; Tanchuck et al, 2009), and separate studies determined that administration of finasteride increased chronic ethanol withdrawal (Gililland-Kaufman et al, 2008; Tanchuck et al, 2013). These preclinical findings are consistent with results in small cohorts of alcoholic patients in which a decrease in plasma ALLO and THDOC levels corresponded to an increase in the subjective ratings of anxiety and depression during the early withdrawal phase (Romeo et al, 1996).…”

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

“…However, basal levels of 5 neuroactive steroids were lower in WSP-1 versus WSR-1 mice, so a withdrawal-induced decrease in WSP-1 mice may have a greater physiological consequence versus a similar decrease in WSR-1 mice (e.g., 3α,5α- and 3α,5β-androstanediols). Additionally, it should be considered that ethanol withdrawal renders WSP mice (but not WSR) from both genetic replicates tolerant to the anticonvulsant effects of exogenous ALLO administration (e.g., Finn et al, 2006; Gililland-Kaufman et al, 2008; Tanchuck et al, 2013). This behavioral tolerance corresponded to a rightward shift in the ability of ALLO to potentiate GABA-stimulated chloride flux in forebrain microsacs (Finn et al, 2006), consistent with a reduction in GABA A R sensitivity to GABA A R-active neuroactive steroids during ethanol withdrawal in WSP mice.…”

Ethanol withdrawal-induced dysregulation of neurosteroid levels in plasma, cortex, and hippocampus in genetic animal models of high and low withdrawal

“…Both WSP and DBA mice exhibit high WD severity, measured by increased handling-induced convulsions (HICs) and anxiety following 72 h of continuous ethanol vapor or CIE vapor (Crabbe et al, 1985; Crabbe, 1998; Finn and Crabbe, 1999; Finn et al, 2004; Gorin et al, 2005; Kosobud and Crabbe, 1986; McCool and Chappell, 2015; Metten and Crabbe, 2005; Metten et al, 2010). Importantly, both WSP and DBA mice exhibited tolerance to ALLO’s anticonvulsant effect during peak (8 h) WD (Beckley et al, 2008; Finn et al, 2000, 2004, 2006), and this tolerance in WSP mice also corresponded to a reduction in functional sensitivity of GABA A Rs to ALLO (Finn et al, 2006) and reduced anticonvulsant effect of intra-CA1 ALLO (Gililland-Kaufman et al, 2008). Thus, a reduction in GABA A R sensitivity to neurosteroids may contribute to a severe WD phenotype in genetic animal models of high WD.…”

“…Cortical and hippocampal regions also exhibit increased spike and sharp wave activity in the electroencephalogram (e.g., Veatch and Becker, 2002; Veatch and Gonzalez, 1996) and increased immediate early gene expression (Vilpoux et al, 2009) that was greater in DBA versus C57 mice (Chen et al, 2009) during WD, highlighting the importance of these brain regions in WD circuitry. Thus, one purpose of the present experiments was to use GC-MS to simultaneously quantify a panel of 10 neurosteroids in plasma, cortex and hippocampus in control-treated male WSP-1 and WSR-1 mice and over the time course of WD that corresponded to enhanced convulsive activity in WSP mice (i.e., 8 and 48 h; Finn et al, 2004; Gililland-Kaufman et al, 2008), and that also corresponded to the acute phase of WD in mice (i.e., 0 – 48 h in rodents; Heilig et al, 2010). We predicted that the WD-induced changes in brain neurosteroid levels would differ from the pattern detected in plasma and that there would be a line difference in hippocampal and cortical neurosteroid levels during WD.…”