Fig. 5

Anantin increases P2X3 serine phosphorylation. a Western immunoblotting shows the amount of P2X3 pSer and total amount of P2X3 receptors in control and after 1 h anantin (500 nM) application. β-tubulin was used as loading control of the total extract. Histograms on the right show statistically lower P2X3 pSer (relative optical density value) after anantin application compared to control (n = 4; *p < 0.05, Mann–Whitney rank sum test). b Representative example of Western immunoblotting, showing the amount of P2X3 pTyr and total amount of P2X3 receptors in control and after anantin application; β-tubulin used as loading control of the total extract. P2X3 pTyr does not change as shown in the plot on the right, summarizing pTyr P2X3 relative optical density in control and after anantin treatment. c Western immunoblotting shows the amount of P2X3 pSer and total amount of P2X3 receptors in control and after prolonged anantin application (500 nM, 24 h) (n = 3; *p < 0.05, Mann–Whitney rank sum test). β-tubulin used as loading control of the total extract. Note that anantin effects after 24 h or 1 h of treatment (panels c and a, respectively) are virtually indistinguishable. d Representative example of Western immunoblotting showing the amount of P2X3 pSer in control and after treatment with anantin (500 nM, 2–3 h), siRNA BNP (24 h), or their combination (n = 4); β-tubulin is used as loading control of the total extract. Histograms on the right quantify P2X3 pSer (relative optical density) according to the treatment (n = 4; *p < 0.05, Kruskal–Wallis test)