Marks Michael J, Whiteaker Paul, Collins Allan C
Institute for Behavioral Genetics, 447 UCB, University of Colorado, Boulder, CO 80309, USA.
Mol Pharmacol. 2006 Sep;70(3):947-59. doi: 10.1124/mol.106.025338. Epub 2006 May 25.
Diversity of neuronal nicotinic acetylcholine receptor binding was measured using [3H]epibatidine after deletion of alpha7, beta2, or beta4 subunits. [3H]Epibatidine binding is distinctly biphasic. Densities of higher (Kd approximately 0.02 nM) and lower (Kd approximately 5 nM) affinity sites in whole brains of wild-type mice are very similar. Relative sensitivity to inhibition by cytisine or alpha-bungarotoxin was used to evaluate pharmacological subsets of the higher- and lower-affinity sites, respectively. Deletion of each subunit had distinct effects on the binding sites. Deletion of alpha7 did not affect higher-affinity sites but reduced the numbers of lower-affinity sites. This reduction was confined to the [3H]epibatidine binding sites sensitive to inhibition by alpha-bungarotoxin. Deletion of the beta2 subunit had the largest effect. Higher-affinity sites sensitive to inhibition by cytisine were eliminated, and cytisine-resistant sites were reduced. Deletion of the beta2 subunit also significantly reduced the number of lower-affinity sites insensitive to alpha-bungarotoxin. beta4 Gene deletion partially reduced cytisine-resistant and alpha-bungarotoxin-resistant sites with lower and higher affinity for [3H]epibatidine, respectively. Gene deletion in four brain regions (thalamus, hippocampus, superior colliculus, and inferior colliculus) elicited changes generally similar to whole brain. However, relative expression of the binding sites differed among the regions. [3H]Cytisine and 125I-alpha-bungarotoxin binding sites were eliminated by beta2 and alpha7 gene deletion, respectively. These studies establish that the lower-affinity sites represent a structurally diverse set of sites that require expression of either alpha7, beta2, or beta4 subunits and extend and confirm previous classifications of the higher-affinity [3H]epibatidine binding sites.
在缺失α7、β2或β4亚基后，使用[3H]埃皮巴蒂啶测量神经元烟碱型乙酰胆碱受体结合的多样性。[3H]埃皮巴蒂啶结合明显呈双相性。野生型小鼠全脑中高亲和力（Kd约为0.02 nM）和低亲和力（Kd约为5 nM）位点的密度非常相似。分别使用对金雀花碱或α-银环蛇毒素抑制的相对敏感性来评估高亲和力和低亲和力位点的药理学亚组。每个亚基的缺失对结合位点有不同的影响。α7亚基的缺失不影响高亲和力位点，但减少了低亲和力位点的数量。这种减少仅限于对α-银环蛇毒素抑制敏感的[3H]埃皮巴蒂啶结合位点。β2亚基的缺失影响最大。对金雀花碱抑制敏感的高亲和力位点被消除，对金雀花碱耐药的位点减少。β2亚基的缺失也显著减少了对α-银环蛇毒素不敏感的低亲和力位点的数量。β4基因缺失分别部分降低了对[3H]埃皮巴蒂啶具有较低和较高亲和力的对金雀花碱耐药和对α-银环蛇毒素耐药的位点。四个脑区（丘脑、海马体、上丘和下丘）的基因缺失引起的变化通常与全脑相似。然而，结合位点的相对表达在不同区域之间有所不同。[3H]金雀花碱和125I-α-银环蛇毒素结合位点分别通过β2和α7基因缺失而被消除。这些研究表明，低亲和力位点代表了一组结构多样的位点，需要α7、β2或β4亚基的表达，并扩展和证实了先前对高亲和力[3H]埃皮巴蒂啶结合位点的分类。
