Ligand Recognition Sites on P_2XReceptors Studied by Quantitative Autoradiography of [³H]α,β-Methylene-ATP Binding in Rat Brain

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• 作者：Worthington ; Rebecca A. ; Hansen ; Mitchell A. ; Bennett ; Maxwell R. ; Barden ; Julian A. ; Balcar ; Vladimir J.
• 刊名：Biochemical and Biophysical Research Communications
• 出版年：1998
• 出版时间：August 10, 1998
• 年：1998
• 卷：249
• 期：1
• 页码：166-171
• 全文大小：134 K

文摘

The specificity of α,β-methylene-ATP for P_2Xreceptor binding sites in the CNS has been examined by testing the effects of several ATP analogues and other ATP-related substances on the binding of 10 nM [³H]α,β-methylene-ATP to 20 μm thick sections of fresh-frozen rat brain. The labelling of the putative P_2Xreceptor binding sites by [³H]α,β-methylene-ATP was evaluated by quantitative densitometry. [³H]α,β-Methylene-ATP binding was strongly inhibited by two close ATP analogues, 3′-O-(trinitrophenyl)-adenosine-5′-triphosphate and β,γ-imido-ATP (IC₅₀2-5 μM). β,γ-Methylene-ATP was, however, less potent (<50 % inhibition at 25 μM). Inosine-5′-triphosphate, guanosine-5′-triphosphate, uridine-5′-triphosphate, and cytidine-5′-triphosphate were practically inactive up to concentrations of 100 μM. Periodate oxidised ATP and 1,N⁶-etheno-ATP produced <50 % inhibition at 100 and 500 μM concentrations, respectively. Cations (K⁺, Rb⁺, Cs⁺, and Mg²⁺at 5 mM and Na⁺at 150 mM) reduced [³H]α,β-methylene-ATP binding by no more than 50 % . Several agents known to interact with Ca²⁺- and/or ATP-related cationic channels (Cd²⁺, glibenclamide, dantrolene, nifedipine, and thapsigargin) had no effect. We conclude that [³H]α,β-methylene-ATP at low nanomolar concentrations binds to a site that has very strict structural requirements and is pharmacologically similar to ATP P_2Xreceptors.