Hosie Alastair M, Clarke Laura, da Silva Helena, Smart Trevor G
University College London, Department of Neuroscience, Physiology and Pharmacology, Gower Street, London WC1E 6BT, UK.
Neuropharmacology. 2009 Jan;56(1):149-54. doi: 10.1016/j.neuropharm.2008.07.050. Epub 2008 Aug 13.
This study addresses whether the potentiation site for neurosteroids on GABA(A) receptors is conserved amongst different GABA(A) receptor isoforms. The neurosteroid potentiation site was previously identified in the alpha1beta2gamma2S receptor by mutation of Q241 to methionine or leucine, which reduced the potentiation of GABA currents by the naturally occurring neurosteroids, allopregnanolone or tetrahydrodeoxycorticosterone (THDOC). By using heterologous expression of GABA(A) receptors in HEK cells, in combination with whole-cell patch clamp recording methods, a relatively consistent potentiation by allopregnanolone of GABA-activated currents was evident for receptors composed of one alpha subunit isoform (alpha2-5) assembled with beta3 and gamma2S subunits. Using mutant alphabetagamma receptors, the neurosteroid potentiation was universally dependent on the conserved glutamine residue in M1 of the respective alpha subunit. Studying wild-type and mutant receptors composed of alpha4beta3delta subunits revealed that the delta subunit is unlikely to contribute to the neurosteroid potentiation binding site and probably affects the efficacy of potentiation. Thus, in keeping with the ability of neurosteroids to potentiate GABA currents via a broad variety of GABA(A) receptor isoforms in neurons, the potentiation site is structurally highly conserved on this important neurotransmitter receptor family.
本研究探讨了神经甾体在GABA(A)受体上的增强位点在不同GABA(A)受体亚型中是否保守。神经甾体增强位点先前在α1β2γ2S受体中通过将Q241突变为甲硫氨酸或亮氨酸而得以确定，这降低了天然存在的神经甾体别孕烯醇酮或四氢脱氧皮质酮（THDOC）对GABA电流的增强作用。通过在HEK细胞中利用GABA(A)受体的异源表达，并结合全细胞膜片钳记录方法，对于由一个α亚基亚型（α2 - 5）与β3和γ2S亚基组装而成的受体，别孕烯醇酮对GABA激活电流具有相对一致的增强作用。使用突变的αβγ受体，神经甾体增强作用普遍依赖于各个α亚基M1中保守的谷氨酰胺残基。对由α4β3δ亚基组成的野生型和突变型受体的研究表明，δ亚基不太可能对神经甾体增强结合位点有贡献，并且可能影响增强的效力。因此，与神经甾体通过神经元中多种GABA(A)受体亚型增强GABA电流的能力一致，该增强位点在这个重要的神经递质受体家族中在结构上高度保守。
