Estrous cycle variations in GABAA receptor phosphorylation enable rapid modulation by anabolic androgenic steroids in the medial preoptic area

Anabolic androgenic steroids (AAS), synthetic testosterone derivatives that are used for ergogenic purposes, alter neurotransmission and behaviors mediated by GABAA receptors. Some of these effects may reflect direct and rapid action of these synthetic steroids at the receptor. The ability of other natural allosteric steroid modulators to alter GABAA receptor-mediated currents is dependent upon the phosphorylation state of the receptor complex. Here we show that phosphorylation of the GABAA receptor complex immunoprecipitated by β2/β3 subunit-specific antibodies from the medial preoptic area (mPOA) of the mouse varies across the estrous cycle; with levels being significantly lower in estrus. Acute exposure to the AAS, 17α-methyltestosterone (17α-MeT), had no effect on the amplitude or kinetics of inhibitory postsynaptic currents in the mPOA of estrous mice when phosphorylation was low, but increased the amplitude of these currents from mice in diestrus, when it was high. Inclusion of the protein kinase C (PKC) inhibitor, calphostin, in the recording pipette eliminated the ability of 17α-MeT to enhance currents from diestrous animals, suggesting that PKC-receptor phosphorylation is critical for the allosteric modulation elicited by AAS during this phase. In addition, a single injection of 17α-MeT was found to impair an mPOA-mediated behavior (nest building) in diestrus, but not in estrus. PKC is known to target specific serine residues in the β3 subunit of the GABAA receptor. Although phosphorylation of these β3 serine residues showed a similar profile across the cycle, as did phosphoserine in mPOA lysates immunoprecipitated with β2/β3 antibody (lower in estrus than in diestrus or proestrus), the differences were not significant. These data suggest that the phosphorylation state of the receptor complex regulates both the ability of AAS to modulate receptor function in the mPOA and the expression of a simple mPOA-dependent behavior through a PKC-dependent mechanism that involves the β3 subunit and other sites within the GABAA receptor complex.

► GABAA receptor phosphorylation was higher in diestrus/proestrus than in estrus.
► AAS elicited rapid enhancement of synaptic currents in diestrus, but not in estrus.
► Effects of AAS in diestrus were blocked by PKC inhibition.
► Levels of GABAA receptor subunit mRNAs did not vary across the estrous cycle.
► Single AAS injections impaired an mPOA-mediated behavior in diestrus, but not in estrus.