Wegner Florian, Rassler Cornelia, Allgaier Clemens, Strecker Karl, Wohlfarth Kai
Department of Neurology, University of Leipzig, Liebigstrasse 22a, 04103 Leipzig, Saxonia, Germany.
Neuropharmacology. 2007 Feb;52(2):672-83. doi: 10.1016/j.neuropharm.2006.09.009. Epub 2006 Nov 3.
GABA(A) receptor function is modulated by various important drugs including neuroactive steroids that act on allosteric modulatory sites and can directly activate GABA(A) receptor channels at high concentrations. We used whole cell patch-clamp recordings and rapid applications of the neuroactive steroid alphaxalone to investigate repetitive steroid effects. Alphaxalone potentiation of submaximal GABA-evoked currents was enhanced significantly by repetitive coapplications at all investigated recombinant isoforms (alpha1beta3delta, alpha1beta3gamma2L, alpha6beta3delta, alpha6beta3gamma2L) and at GABA(A) receptors of differentiated human NT2 neurons. A similar increase of current amplitudes was induced by repetitive applications of a high steroid concentration without GABA. We refer to these reversible effects as auto-modulation because repeated interactions of steroids enhanced their own pharmacological impact at the receptor sites in a time and concentration dependent manner without affecting GABA controls. Pronounced auto-modulatory actions were also measured using the neurosteroid 5alpha-THDOC in contrast to indiplon, THIP, and pentobarbital indicating a steroid specificity. Protein kinase A inhibition significantly reduced alphaxalone auto-modulation at alpha1beta3gamma2L, alpha6beta3gamma2L, and alpha6beta3delta subtypes while it enhanced potentiation at alpha1beta3delta isoforms suggesting a crucial influence of receptor subunit composition and phosphorylation for steroid actions. Especially at extrasynaptic GABA(A) receptor sites containing the delta subunit steroid auto-modulation may have a critical role in enhancing potentiation of GABA-induced currents.
GABA(A)受体功能受到多种重要药物的调节，包括作用于变构调节位点的神经活性甾体，这些甾体在高浓度时可直接激活GABA(A)受体通道。我们使用全细胞膜片钳记录和神经活性甾体alphaxalone的快速应用来研究甾体的重复效应。在所有研究的重组亚型（α1β3δ、α1β3γ2L、α6β3δ、α6β3γ2L）以及分化的人NT2神经元的GABA(A)受体上，重复共同应用时，alphaxalone对亚最大GABA诱发电流的增强作用显著增强。在没有GABA的情况下，重复应用高浓度甾体也会诱导类似的电流幅度增加。我们将这些可逆效应称为自调节，因为甾体的重复相互作用以时间和浓度依赖的方式增强了它们在受体位点自身的药理作用，而不影响GABA的调控。与茚地普隆、THIP和戊巴比妥相比，使用神经甾体5α-THDOC也测量到了明显的自调节作用，表明存在甾体特异性。蛋白激酶A抑制显著降低了α1β3γ2L、α6β3γ2L和α6β3δ亚型上alphaxalone的自调节作用，而在α1β3δ亚型上增强了增强作用，这表明受体亚基组成和磷酸化对甾体作用有至关重要的影响。特别是在含有δ亚基的突触外GABA(A)受体位点，甾体自调节可能在增强GABA诱发电流的增强作用中起关键作用。
