García-Lecea M, Delicado E G, Miras-Portugal M T, Castro E
Departamento de Bioquímica y Biología Molecular, Facultad de Veterinaria, Universidad Complutense de Madrid, Spain.
Neuropharmacology. 1999 May;38(5):699-706. doi: 10.1016/s0028-3908(98)00225-1.
P2X receptors present in cerebellar Purkinje cells have been studied by recording ATP-elicited [Ca2+]i signals from immuno-identified (calbindin+) cells in culture using fura-2 microfluorescence. The [Ca2+]i increases evoked by ATP were mimicked by 2MeSATP but not by alpha, beta-meATP and other purinoceptor agonists. The selective P2X1 antagonist diinosine pentaphosphate failed to inhibit ATP-elicited [Ca2+]i transients, but suramin and PPADS rapidly and reversibly blocked the [Ca2+]i responses to ATP and 2MeSATP. The IC50 values for suramin and PPADS inhibition were 48.7 +/- 4.4 and 5.9 +/- 0.3 microM, respectively. Both antagonists blocked completely the signal elicited by ATP, revealing that there was not a separate antagonist-insensitive P2X receptor population in Purkinje cells. The effect of ATP was potentiated by Zn2+ and H+ ions. A one unit acidification from pH 7.4 to 6.4 enhanced by 172% the [Ca2+]i transient elicited by an intermediate concentration of ATP. Conversely, alkalinization of the medium to pH 8.4 reduced the ATP response by 88%. This combination of pharmacological and modulatory properties indicates that endogenous P2X receptors present in Purkinje neurons are formed by P2X2 subunits, rather than the more abundantly expressed P2X4 purinoceptor subunits.
通过使用fura - 2微荧光技术记录培养的免疫鉴定（钙结合蛋白阳性）细胞中ATP引发的[Ca2+]i信号，对小脑浦肯野细胞中存在的P2X受体进行了研究。ATP引发的[Ca2+]i增加可被2MeSATP模拟，但不能被α,β - meATP和其他嘌呤受体激动剂模拟。选择性P2X1拮抗剂二肌苷五磷酸未能抑制ATP引发的[Ca2+]i瞬变，但苏拉明和PPADS能快速且可逆地阻断[Ca2+]i对ATP和2MeSATP的反应。苏拉明和PPADS抑制的IC50值分别为48.7±4.4和5.9±0.3微摩尔。两种拮抗剂都完全阻断了ATP引发的信号，表明浦肯野细胞中不存在单独的对拮抗剂不敏感的P2X受体群体。ATP的作用被Zn2+和H+离子增强。从pH 7.4到6.4一个单位的酸化使中等浓度ATP引发的[Ca2+]i瞬变增强了172%。相反，将培养基碱化至pH 8.4使ATP反应降低了88%。这种药理学和调节特性的组合表明，浦肯野神经元中存在的内源性P2X受体是由P2X2亚基形成的，而不是表达更丰富的P2X4嘌呤受体亚基。
