猫膀胱中[3H]α,β-亚甲基ATP结合位点的表征及放射自显影定位

Characterization and autoradiographic localization of [3H] alpha,beta-methylene ATP binding sites in cat urinary bladder.

Zhao M, Bo X, Neely C F, Burnstock G

Department of Anatomy and Developmental Biology, University College London, UK.

Gen Pharmacol. 1996 Apr;27(3):509-12. doi: 10.1016/0306-3623(95)02062-4.

Abstract

The characteristics and distribution of [3H] alpha,beta-methylene ATP ([3H] alpha,beta-MeATP), a radioligand for P2X-purinoceptors, binding sites in cat urinary bladder detrusor were examined. 2. Saturation analysis revealed that, in cat bladder membrane preparations, only one population of binding sites with high affinity (Kd = 1.8 nM) was present, in contrast to other species where both high-and low-affinity binding sites are present. Another feature is that the density of the binding sites in the cat bladder (Bmax = 21.2 pmol/mg protein) is considerably higher (about 2-fold) than the high-affinity binding component in the rat bladder membrane preparations. 3. Displacement experiments with unlabelled purinoceptor ligands indicate that [3H] alpha,beta-MeATP mainly binds to P2X-purinoceptors. The order of binding displacement activity was: alpha,beta-methylene ATP, beta,gamma-methylene ATP > 2-methylthioATP > ATP > suramin and L-beta,gamma-methylene ATP > > adenosine. 4. Autoradiographic study demonstrated dense specific binding sites of [3H] alpha,beta-MeATP on detrusor smooth muscle of cat bladder. 5. The results of this study are consistent with pharmacological studies for the existence of P2X-purinoceptors in cat bladder.

摘要

对[3H]α,β-亚甲基ATP（[3H]α,β-MeATP）（一种P2X嘌呤受体的放射性配体）在猫膀胱逼尿肌中的结合位点的特征和分布进行了研究。2. 饱和分析表明，在猫膀胱膜制剂中，仅存在一群具有高亲和力（Kd = 1.8 nM）的结合位点，这与其他同时存在高亲和力和低亲和力结合位点的物种不同。另一个特点是，猫膀胱中结合位点的密度（Bmax = 21.2 pmol/mg蛋白质）比大鼠膀胱膜制剂中的高亲和力结合成分高得多（约2倍）。3. 用未标记的嘌呤受体配体进行的置换实验表明，[3H]α,β-MeATP主要与P2X嘌呤受体结合。结合置换活性的顺序为：α,β-亚甲基ATP、β,γ-亚甲基ATP＞2-甲硫基ATP＞ATP＞苏拉明，且L-β,γ-亚甲基ATP＞＞腺苷。4. 放射自显影研究表明，[3H]α,β-MeATP在猫膀胱逼尿肌平滑肌上有密集的特异性结合位点。5. 本研究结果与关于猫膀胱中存在P2X嘌呤受体的药理学研究一致。