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, 397, 127-137

Genotype Differences in Sensitivity to the Anticonvulsant Effect of the Synthetic Neurosteroid Ganaxolone During Chronic Ethanol Withdrawal

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Genotype Differences in Sensitivity to the Anticonvulsant Effect of the Synthetic Neurosteroid Ganaxolone During Chronic Ethanol Withdrawal

Michelle A Nipper et al. Neuroscience.

Abstract

Sensitivity to anticonvulsant effects of the γ-aminobutyric acidA receptor-active neurosteroid allopregnanolone (ALLO) during ethanol withdrawal varies across genotypes, with high sensitivity in genotypes with mild withdrawal and low sensitivity in genotypes with high withdrawal. The present studies determined whether the resistance to ALLO during withdrawal in mouse genotypes with high handling-induced convulsions (HICs) during withdrawal could be overcome with use of ganaxolone (GAN), the metabolically stable derivative of ALLO. In separate studies, male and female Withdrawal Seizure-Prone (WSP-1) and DBA/2J (D2) mice were exposed to air (controls) or 72-h ethanol vapor and then were scored for HICs during withdrawal (hourly for the first 12 h, then at hours 24 and 25). After the HIC scoring at hours 5 and 9, mice were injected with 10 mg/kg GAN or vehicle. Area under the HIC curve (AUC) for hours 5-12 was analyzed. In control WSP-1 mice, GAN significantly reduced AUC by 52% (males) and 63% (females), with effects that were absent or substantially reduced during withdrawal. In contrast, GAN significantly reduced AUC in both control and ethanol-withdrawing male and female D2 mice. AUC was decreased by 81% (males) and 70% (females) in controls and by 35% (males) and 21% (females) during withdrawal. The significant anticonvulsant effect of GAN during withdrawal in D2 but not WSP-1 mice suggests that different mechanisms may contribute to ALLO insensitivity during withdrawal in these two genotypes. Importantly, the results in D2 mice suggest that GAN may be a useful treatment for ethanol withdrawal-induced seizures.

Keywords: DBA/2J mice; GABA(A) receptors; Withdrawal Seizure-Prone mice; alcohol; allopregnanolone; sex.

Conflict of interest statement

Declarations of interest: None. The contents do not represent the views of the National Institute of Health, the US Department of Veterans Affairs, or the United States Government.

Figures

Figure 1.
Figure 1.. Insensitivity to the anticonvulsant effect of ganaxolone (GAN) during ethanol (EtOH) withdrawal in male WSP-1 mice, measured by hourly handling-induced convulsions (HICs; panel A) and area under the withdrawal curve (AUC, panel B).
Mice were exposed to 72 h EtOH vapor or air, and HICs were measured over the time course of withdrawal. Immediately following the HIC measurement at 5 h and 9 h, mice received an injection of 10 mg/kg GAN or equivalent volume of vehicle (VEH; see arrows on figure). Hourly HIC scores (panel A) and AUC (panel B) were significantly higher in the EtOH vs air exposed mice (p<0.001), and the significant interaction between GAN and EtOH treatment (p<0.001) indicated that the anticonvulsant effect of GAN was significantly reduced during withdrawal. Values are the mean ± SEM for the number of animals in parentheses. In cases where the SEM is not visible, the value is contained within the symbol for the mean. Panels A and B: +p=0.10, **p<0.01, ***p≤0.001 vs respective VEH treatment, planned comparisons (panel A) or post-hoc tests (panel B).
Figure 2.
Figure 2.. Minimal anticonvulsant effect of ganaxolone (GAN) during ethanol (EtOH) withdrawal in female WSP-1 mice, measured by hourly handling-induced convulsions (HICs; panel A) and area under the withdrawal curve (AUC, panel B).
Mice were treated as described in Figure 1. Hourly HICs (panel A) and AUC (panel B) were increased significantly during EtOH withdrawal (p<0.001) and were decreased significantly by GAN injection (p<0.01). The interaction between GAN and EtOH treatment was not significant, suggesting a similar anticonvulsant effect of GAN in the air- and EtOH-exposed mice. However, there was a marked reduction in efficacy during withdrawal. Depicted are the mean ± SEM for the number of mice in parentheses. In cases where the SEM is not visible, the value is contained within the symbol for the mean. Arrows indicate times when the mice were administered GAN or vehicle (VEH). Panel A: *p<0.05, **p<0.01, ***p<0.001 vs respective VEH treatment, planned comparisons. Panel B: **p<0.01 vs VEH, main effect of GAN.
Figure 3.
Figure 3.. Significant anticonvulsant effect of ganaxolone (GAN) in control male DBA/2J mice and during ethanol (EtOH) withdrawal, measured by hourly handling-induced convulsions (HICs; panel A) and area under the withdrawal curve (AUC, panel B).
Mice were treated as described in Figure 1. Hourly HICs (panel A) and AUC (panel B) were increased significantly during EtOH withdrawal (p<0.001) and were decreased significantly by GAN injection (p<0.001). The lack of an interaction between GAN and EtOH treatment suggested that GAN exerted a similar anticonvulsant effect in the air- and EtOH-exposed mice, and the slight reduction in efficacy appeared due to a more rapid return to values in vehicle (VEH)-treated mice during EtOH withdrawal. Mice received 2 injections of GAN or VEH, separated by 4 h (see arrows). Values are the mean ± SEM for the number of mice in parentheses. In cases where the SEM is not visible, the value is contained within the symbol for the mean. Panel A: *p<0.05, **p≤0.01, ***p≤0.001 vs respective VEH (air) or GAN (EtOH) treatment, planned comparisons. Panel B: ***p<0.001 vs VEH, main effect of GAN.
Figure 4.
Figure 4.. Despite low convulsive activity in control animals, there was a significant anticonvulsant effect of ganaxolone (GAN) in control female DBA/2J mice and during ethanol (EtOH) withdrawal, measured by hourly handling-induced convulsions (HICs; panel A) and area under the withdrawal curve (AUC, panel B).
Mice were treated as described in Figure 1. Hourly HIC (panel A) and AUC (panel B) were increased significantly during EtOH withdrawal (p<0.001) and were decreased significantly by GAN injection (p<0.05). The lack of an interaction between GAN and EtOH treatment suggested that GAN exerted a similar anticonvulsant effect in the air- and EtOH-exposed mice. Even with low convulsive activity in the controls, GAN suppressed HIC scores across time. During EtOH withdrawal, GAN had a pronounced anticonvulsant effect, and the more rapid return to values in vehicle (VEH)-treated mice likely contributed to the reduction in efficacy. Mice received 2 injections of GAN or VEH, separated by 4 h (see arrows). Values are the mean ± SEM for the number of mice in parentheses. In cases where the SEM is not visible, the value is contained within the symbol for the mean. Panel A: *p<0.05, ***p≤0.001 vs respective VEH (air) or GAN (EtOH) treatment, planned comparisons. Panel B: *p<0.05 vs VEH, main effect of GAN.

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