Abstrakt
The family of ATP-gated purinergic P2X receptors comprises seven bunits (P2X1-7) that are unevenly distributedin the central and peripheral nervous systems as well as other organs. Endogenous modulators of P2X receptorsare phospholipids, steroids and neurosteroids.
Here, we analyzed whether bile acids, which are natural productsderived from cholesterol, affect P2X receptor activity. We examined the effects of primary and secondary bileacids and newly synthesized derivatives of lithocholic acid on agonist-induced responses in HEK293T cells ex-pressing rat P2X2, P2X4 and P2X7 receptors.
Electrophysiology revealed that low micromolar concentrations oflithocholic acid and its structural analog 4-dafachronic acid strongly inhibit ATP-stimulated P2X2 but potentiateP2X4 responses, whereas primary bile acids and other secondary bile acids exhibit no or reduced effects only athigher concentrations. Agonist-stimulated P2X7 responses are significantly potentiated by lithocholic acid atmoderate concentrations.
Structural modifications of lithocholic acid at positions C-3, C-5 or C-17 abolish bothinhibitory and potentiation effects to varying degrees, and the 3α-hydroxy group contributes to the ability of themolecule to switch between potentiation and inhibition. Lithocholic acid allosterically modulates P2X2 andP2X4 receptor sensitivity to ATP, reduces the rate of P2X4 receptor desensitization and antagonizes the effect ofivermectin on P2X4 receptor deactivation.
Alanine-scanning mutagenesis of the upper halve of P2X4 trans-membrane domain-1 revealed that residues Phe48, Val43 and Tyr42 are important for potentiating effect oflithocholic acid, indicating that modulatory sites for lithocholic acid and ivermectin partly overlap. Lithocholicacid also inhibits ATP-evoked currents in pituitary gonadotrophs expressing native P2X2, and potentiates ATPcurrents in nonidentified pituitary cells expressing P2X4 receptors.
These results indicate that lithocholic acid isa bioactive steroid that may help to further unveil the importance of the P2X2, and P2X4 receptors in manyphysiological processes.