Neurosteroids sensitize female mice to stress

Introduction

Episodic Ataxia type-1 (EA1) is a hereditary human neurological disorder that causes stress-induced loss of motor coordination. In EA1 mice, as in their human counterparts, stress results in ataxia and loss of motor coordination ¹ . At the cellular level EA1 mice have altered GABA synaptic activity in cerebellar Purkinje neurons that may underlie the behavioral phenotype ¹ . Our EA1 mice have revealed a striking gender bias, with females being much more sensitive to stress. Therefore, we tested the hypothesis that females are more sensitive to stress due to the presence of ovarian hormones that affect the GABAergic system.

Methods

Behavior: The duration the mice are able to run before falling off the accelerating rotarod (latency) was used to assess motor coordination. On the day of experimentation mice were given five baseline trials followed by our stress paradigm consisting of i.p. injection of 10 mg/kg isoproterenol (ISO) and 20 min. forced exercise, followed by 3 final rotarod trials.

Electrophysiology

Sagittal cerebellar slices (400 μM thick) were prepared from WT and EA1 littermates (8–12 weeks old). Whole-cell voltage clamp experiments were made from Purkinje cells visualized with infrared DIC on a Leica DMLFS upright microscope. Spontaneous Purkinje cell GABA currents (sIPSCs) were recorded at −60 mV using a standard CsCl based internal solution.

Results

We have found that only female EA1 mice experience impaired motor coordination in response to stress as measured by the accelerating rotarod, reducing their latency from 56. 1±3.8 s to 37.6±4.9 s (n=15; ANOVA p<0.05). Pre-treatment of wild-type mice with 1 mg/kg ALLO (a progesterone metabolite that is a potent positive modulator of GABAA receptors^{2, 3}) also resulted in a gender specific effect following stress, altering the latency of female mice from 61.6±4.1 s to 30.6±3.8 s (n=8; ANOVA p<0.05), while having a significantly smaller effect on male mice (n=8). Injection of picrotoxin (GABAA antagonist) prior to treatment with ALLO prevented the sensitizing effects of ALLO in female mice (n=7), indicating that ALLO caused its behavioral effects by enhancing GABAergic transmission. In addition, ovariectomy abolished the increased sensitivity observed in female mice (n=7). Consistent with our behavioral data, GABAA receptor currents recorded in female Purkinje cells are 10-fold more sensitive to potentiation by ALLO than GABAA receptor currents recorded from male Purkinje cells.

Conclusion

Increased GABA activity onto Purkinje cells increases sensitivity to stress-induced loss of motor coordination ¹ . The progesterone metabolite ALLO, a positive modulator of GABAA receptor activity, sensitizes female mice to stress to a greater degree than male mice. The presence of ovarian sex steroids is required to maintain the gender difference in sensitivity to ALLO. Behavior and electrophysiology data indicate that female GABA activity is more sensitive to ALLO than male GABA activity in the cerebellum.