A 17β‐derivative of allopregnanolone is a neurosteroid antagonist at a cerebellar subpopulation of GABA A receptors with nanomolar affinity

Background and purpose: High‐affinity, subtype‐selective antagonists of the neurosteroid binding sites of GABA A receptors are not available. We have characterized an allopregnanolone derivative as an antagonist of cerebellar GABA A receptors with nanomolar affinity. Experimental approach: Receptor binding and electrophysiological methods were used for the allosteric modulation of cerebellar GABA A receptors by an allopregnanolone derivative, (20R)‐17β‐(1‐hydroxy‐2,3‐butadienyl)‐5α‐androstane‐3α‐ol (HBAO). GABA A receptors of rat cerebellar membranes were labelled with the chloride channel blocker [ 3 H]ethynylbicycloorthobenzoate (EBOB). The ionophore function of GABA A receptors was studied by whole‐cell patch clamp electrophysiology in cultured rat cerebellar granule and cortical cells. Key results: Partial displacement of cerebellar [ 3 H]EBOB binding by nanomolar HBAO was attenuated by 0.1 mM furosemide, an antagonist of α 6 and β 2‐3 subunit‐containing GABA A receptors. Displacement curves of HBAO were reshaped by 30 nM GABA and shifted to the right. However, the micromolar potency of full displacement by allopregnanolone was not affected by 0.1 mM furosemide or 30 nM GABA. The nanomolar, but not the micromolar phase of displacement of [ 3 H]EBOB binding by GABA was attenuated by 100 nM HBAO. Submicromolar HBAO did not affect [ 3 H]EBOB binding to cortical and hippocampal GABA A receptors. HBAO up to 1 μM did not affect chloride currents elicited by 0.3‐10 μM GABA, while it abolished potentiation by 1 μM allopregnanolone with nanomolar potency in cerebellar but not in cortical cells. Furosemide attenuated cerebellar inhibition by 100 nM HBAO. Conclusions and implications: HBAO is a selective antagonist of allopregnanolone, a major endogenous positive modulator via neurosteroid sites of cerebellar (probably α 6 β 2–3 δ) GABA A receptors. British Journal of Pharmacology (2007) 151 , 1078–1086; doi: 10.1038/sj.bjp.0707316