Halonen Lauri M, Sinkkonen Saku T, Chandra Dev, Homanics Gregg E, Korpi Esa R
Institute of Biomedicine, Pharmacology, Biomedicum Helsinki, University of Helsinki, FI-00014 Helsinki, Finland.
Neurochem Int. 2009 Nov;55(6):389-96. doi: 10.1016/j.neuint.2009.04.008. Epub 2009 May 4.
The major inhibitory neurotransmitter in the brain, gamma-aminobutyric acid (GABA), has only partial efficacy at certain subtypes of GABA(A) receptors. To characterize these minor receptor populations in rat and mouse brains, we used autoradiographic imaging of t-butylbicyclophosphoro[(35)S]thionate ([(35)S]TBPS) binding to GABA(A) receptors in brain sections and compared the displacing capacities of 10mM GABA and 1mM 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP), a competitive GABA-site agonist. Brains from GABA(A) receptor alpha1, alpha4, delta, and alpha4+delta subunit knockout (KO) mouse lines were used to understand the contribution of these particular receptor subunits to "GABA-insensitive" (GIS) [(35)S]TBPS binding. THIP displaced more [(35)S]TBPS binding than GABA in several brain regions, indicating that THIP also inhibited GIS-binding. In these regions, GABA prevented the effect of THIP on GIS-binding. GIS-binding was increased in the cerebellar granule cell layer of delta KO and alpha4+delta KO mice, being only slightly diminished in that of alpha1 KO mice. In the thalamus and some other forebrain regions of wild-type mice, a significant amount of GIS-binding was detected. This GIS-binding was higher in alpha4 KO mice. However, it was fully abolished in alpha1 KO mice, indicating that the alpha1 subunit was obligatory for the GIS-binding in the forebrain. Our results suggest that native GABA(A) receptors in brain sections showing reduced displacing capacity of [(35)S]TBPS binding by GABA (partial agonism) minimally require the assembly of alpha1 and beta subunits in the forebrain and of alpha6 and beta subunits in the cerebellar granule cell layer. These receptors may function as extrasynaptic GABA(A) receptors.
大脑中的主要抑制性神经递质γ-氨基丁酸（GABA），在某些GABA(A)受体亚型上仅具有部分效力。为了表征大鼠和小鼠大脑中的这些次要受体群体，我们使用了叔丁基双环磷硫代酸酯([(35)S]TBPS)与脑切片中GABA(A)受体结合的放射自显影成像，并比较了10mM GABA和1mM 4,5,6,7-四氢异恶唑并[5,4-c]吡啶-3-醇（THIP，一种竞争性GABA位点激动剂）的置换能力。使用来自GABA(A)受体α1、α4、δ和α4+δ亚基敲除（KO）小鼠品系的大脑，以了解这些特定受体亚基对“GABA不敏感”（GIS）[(35)S]TBPS结合的贡献。在几个脑区中，THIP比GABA置换了更多的[(35)S]TBPS结合，表明THIP也抑制了GIS结合。在这些区域中，GABA阻止了THIP对GIS结合的影响。在δ KO和α4+δ KO小鼠的小脑颗粒细胞层中，GIS结合增加，在α1 KO小鼠的小脑颗粒细胞层中仅略有减少。在野生型小鼠的丘脑和其他一些前脑区域，检测到大量的GIS结合。这种GIS结合在α4 KO小鼠中更高。然而，在α1 KO小鼠中它完全消失，表明α1亚基对于前脑中的GIS结合是必需的。我们的结果表明，在脑切片中，显示GABA对[(35)S]TBPS结合的置换能力降低（部分激动作用）的天然GABA(A)受体，在前脑中至少需要α1和β亚基的组装，在小脑颗粒细胞层中至少需要α6和β亚基的组装。这些受体可能作为突触外GABA(A)受体发挥作用。
