The family of ATP-gated purinergic P2X receptors comprises seven subunits (P2 × 1-7) that are unevenly distributed in the central and peripheral nervous systems as well as other organs. Endogenous modulators of P2X receptors are phospholipids, steroids and neurosteroids. Here, we analyzed whether bile acids, which are natural products derived from cholesterol, affect P2X receptor activity. We examined the effects of primary and secondary bile acids and newly synthesized derivatives of lithocholic acid on agonist-induced responses in HEK293 T cells expressing rat P2 × 2, P2 × 4 and P2 × 7 receptors. Electrophysiology revealed that low micromolar concentrations of lithocholic acid and its structural analog 4-dafachronic acid strongly inhibit ATP-stimulated P2 × 2 but potentiate P2 × 4 responses, whereas primary bile acids and other secondary bile acids exhibit no or reduced effects only at higher concentrations. Agonist-stimulated P2 × 7 responses are significantly potentiated by lithocholic acid at moderate concentrations. Structural modifications of lithocholic acid at positions C-3, C-5 or C-17 abolish both inhibitory and potentiation effects to varying degrees, and the C3α-hydroxy group contributes to the ability of the molecule to switch between potentiation and inhibition. Lithocholic acid allosterically modulates P2 × 2 and P2 × 4 receptor sensitivity to ATP, reduces the rate of P2 × 4 receptor desensitization and antagonizes the effect of ivermectin on P2 × 4 receptor deactivation. Alanine-scanning mutagenesis of the upper halve of P2 × 4 transmembrane domain-1 revealed that residues Phe48, Val43 and Tyr42 are important for potentiating effect of lithocholic acid, indicating that modulatory sites for lithocholic acid and ivermectin partly overlap. Lithocholic acid also inhibits ATP-evoked currents in pituitary gonadotrophs expressing native P2 × 2, and potentiates ATP currents in nonidentified pituitary cells expressing P2 × 4 receptors. These results indicate that lithocholic acid is a bioactive steroid that may help to further unveil the importance of the P2 × 2, and P2 × 4 receptors in many physiological processes.

ATP门控的嘌呤能P2X受体家族包含七个亚单位（P2×1-7），它们在中枢神经系统，周围神经系统以及其他器官中分布不均。P2X受体的内源性调节剂是磷脂，类固醇和神经类固醇。在这里，我们分析了胆汁酸（胆固醇的天然产物）是否会影响P2X受体活性。我们检查了伯胆酸和伯胆酸以及新合成的石胆酸衍生物对激动剂诱导的表达大鼠P2×2，P2×4和P2×7受体的HEK293 T细胞反应的影响。电生理学发现，低微摩尔浓度的石胆酸及其结构类似物4-dafachronic acid强烈抑制ATP刺激的P2×2，但增强P2×4的反应，而伯胆汁酸和其他仲胆汁酸仅在较高浓度下不显示或减少影响。激动剂刺激的P2×7反应在中等浓度下可被石胆酸显着增强。C-3，C-5或C-17位的石胆酸的结构修饰在不同程度上消除了抑制作用和增强作用，并且C3α-羟基有助于分子在增强作用和抑制作用之间进行切换。胆酸可变构地调节P2×2和P2×4受体对ATP的敏感性，降低P2×4受体脱敏的速率，并拮抗伊维菌素对P2×4受体失活的作用。P2×4跨膜结构域1上半部分的丙氨酸扫描诱变显示，残基Phe48，Val43和Tyr42对于增强石胆酸的作用很重要，表明石胆酸和伊维菌素的调节位点部分重叠。胆酸还抑制表达天然P2×2的垂体促性腺激素中ATP诱发的电流，并增强表达P2×4受体的未确认垂体细胞中的ATP电流。这些结果表明，石胆酸是一种生物活性类固醇，可能有助于进一步揭示P2×2和P2×4受体在许多生理过程中的重要性。