Lazareno S, Farries T, Birdsall N J
MRC Collaborative Centre, Mill Hill, London, U.K.
Life Sci. 1993;52(5-6):449-56. doi: 10.1016/0024-3205(93)90301-i.
We have studied muscarinic agonist stimulated [35S]GTP gamma S binding and [gamma 32P]GTP hydrolysis (GTPase) in membranes from CHO cells stably transfected with human muscarinic m1-m4 receptors. 'Full' agonists were at least 10-fold more potent at m2 & m4 receptors than at m1 & m3. This pattern was less marked with 'partial' agonists, which had a greater maximal effect at m2 & m4 than at m1 & m3. McN-A343 uniquely was more potent and efficacious at m4 than at m2 receptors. Antagonist affinity constants were estimated by fitting the data from inhibition curves directly to the Schild model. Antagonist affinity estimates were very similar to those measured earlier in binding studies using animal tissues, and confirmed a small degree of m4 selectivity for tropicamide and secoverine. The receptor subtypes activated more than one G-protein subtype; m2 & m4 receptors activated only pertussis (PTX) sensitive G-proteins, while m1 & m3 coupled to both PTX sensitive and insensitive G-proteins. Acetylcholine (ACh) was more potent in stimulating guanine nucleotide exchange in PTX-treated m1 cells than in controls.
我们研究了用人类毒蕈碱型m1 - m4受体稳定转染的CHO细胞的膜中，毒蕈碱激动剂刺激的[35S]GTPγS结合和[γ32P]GTP水解（GTP酶）情况。“完全”激动剂对m2和m4受体的效力比对m1和m3受体至少高10倍。这种模式在“部分”激动剂中不太明显，“部分”激动剂在m2和m4上的最大效应比在m1和m3上更大。McN - A343在m4受体上比在m2受体上具有独特的更高效力和效能。通过将抑制曲线数据直接拟合到Schild模型来估计拮抗剂亲和力常数。拮抗剂亲和力估计值与早期使用动物组织进行的结合研究中测得的值非常相似，并证实了托吡卡胺和塞克维林对m4有一定程度的选择性。受体亚型激活不止一种G蛋白亚型；m2和m4受体仅激活对百日咳毒素（PTX）敏感的G蛋白，而m1和m3与对PTX敏感和不敏感的G蛋白都偶联。乙酰胆碱（ACh）在刺激经PTX处理的m1细胞中的鸟嘌呤核苷酸交换方面比在对照细胞中更有效。
