GABA receptors as broadcasters of sexually differentiating signals in the brain

Abstract

Epileptic seizures are more common in males than in females. One of the areas that has recently been implicated in the higher susceptibility of males to seizures is the substantia nigra reticulata (SNR). Several studies support the existence of phenotypic differences between male and female infantile SNR neurons, and particularly in several aspects of the GABAergic system, including its ability to control seizures. We have recently found that at postnatal day 14-17 (PN14-17) rats, which are equivalent to infants, activation of GABA(A) receptors has different physiological effects in male and female SNR neurons. This is likely due to the differences in the expression of the neuronal-specific potassium-chloride co-transporter KCC2, which regulates the intracellular chloride concentration. In male PN14-17 SNR neurons, GABA(A)-receptor activation with muscimol causes depolarization and increments in intracellular calcium concentration and the expression of calcium regulated genes, such as KCC2. Blockade of L-type voltage-sensitive calcium channels (L-VSCC) by nifedipine decreases KCC2 mRNA expression. However, in PN14-17 females, muscimol hyperpolarizes the SNR neurons, does not increase intracellular calcium, and decreases KCC2 mRNA expression. In PN15 females, nifedipine has no effect on KCC2 mRNA expression in the SNR. This sexually dimorphic function of GABA(A) receptors also creates divergent patterns of estradiol signaling. In male PN15 rats, estradiol decreases KCC2 mRNA expression in SNR neurons. Pretreatment with the GABA(A)-receptor antagonist bicuculline or with nifedipine, prevents the appearance of estradiol-mediated downregulation of KCC2 mRNA expression. In contrast, in PN15 females, estradiol does not influence KCC2 expression. These findings show that, in infantile rats, drugs or conditions that modulate the activity of GABA(A) receptors or L-VSCCs have different effects on the differentiation of the SNR. As a result, they have the potency of causing long-term changes in the function of the SNR in the control of seizures, movement, and the susceptibility to and course of epilepsy and movement disorders.

Figure 1. Sex differences in KCC2 and GABA_A receptor function in PN15 rat SNR

Panels A and B: KCC2-specific digoxigenin labeled in situ hybridization of PN15 SNR sections. Male SNR neurons (panel A) express lower KCC2 mRNA than female (panel B) SNR neurons. Panel C: In PN15 male SNR, the low KCC2 expression neurons results in high intracellular chloride concentration. Upon GABA-mediated activation of GABA_A receptors, chloride efflux ensues, followed by depolarization, activation of voltage-sensitive calcium channels (VSCCs) and increase in intracellular calcium. Panel D: In PN15 female SNR neurons, the high KCC2 expression is sufficient to lower intracellular chloride. As a result, activation of GABA_A receptors leads to influx of chloride, hyperpolarization, without affecting the activity of VSCCs.

Figure 2. Sexually dimorphic regulatory pathways affecting KCC2 mRNA expression in PN15 male and female rat SNR neurons

In male PN15 rats, GABA_A receptor or L-type VSCC (L-VSCC) activation increases KCC2 mRNA expression, whereas estradiol downregulates KCC2 mRNA expression. The asterisk indicates that estradiol-mediated downregulation of KCC2 mRNA occurs only when GABA-mediated activation of L-VSCCs is present. In female PN15 rats however, GABA_A receptor activation downregulates KCC2 mRNA, whereas estradiol has no effect.