Fig 2: Inhibition of P2X3 alleviated bladder overactivity evoked by TNBS-induced colitis. (A) Representative tracers of cystometry patterns from three groups responded to A-317491 by intrathecal injection. The rats with colitis exhibited bladder overactivity, manifested by increased micturition frequency and bladder pressure. n = 4 for each group. (B) Histogram showed that the ICI of rats treated with TNBS was significantly shorter than that of the control group, and intrathecal injection with A-317491 significantly prolonged the ICI of both control and colitis groups. (C) Histogram showed that colitis significantly increased the maximum BP of rats at day 3 post-TNBS but not at day 7 post-TNBS. Intrathecal injection with A-317491 significantly inhibited the increased maximum BP at day 3 post-TNBS. *p < 0.05, **p < 0.01 by Student’s t-test or one-way ANOVA followed by Newman–Keuls post hoc tests.

Fig 3: TNBS-induced colitis increased P2X3 expression in the bladder. (A–B) Western blotting showed that TNBS-induced colitis significantly increased the protein expression of the P2X3 receptor in the bladder. n = 3 for each group. (C–D) P2X3 immunoreactivity in the bladder showed that colitis significantly increased P2X3 expression of the urothelium of the bladder at days 3 and 7 post-TNBS compared to the control group. Bar = 100 µm; n = 4 for each group. *p < 0.05 by one-way ANOVA followed by Newman–Keuls post hoc tests.

Fig 4: Spontaneous contractions of the detrusor strip isolated from the bladder of three groups had no sensitivity to the inhibition of P2X3 receptors. (A) Representative tracers of detrusor strip contractility responded to A-317491. The detrusor strip isolated from colitis rats exhibited the characteristic of detrusor overactivity. n = 4 for each group. (B) Histogram showed that TNBS-induced colitis significantly increased the phasic amplitude of detrusor strip contractility compared to the control group, but treatment with A-317491 had no effect on the detrusor strip contractility isolated from both control and colitis groups. *p < 0.05 by one-way ANOVA followed by Newman–Keuls post hoc tests.

Fig 5: TNBS-induced colitis increased the P2X3 expression in the distal colon. (A–B) Western blotting showed that TNBS-induced colitis significantly increased the protein expression of the P2X3 receptor in the distal colon. n = 3 for each group. (C–D) P2X3 immunoreactivity in the submucosal plexus of the distal colon showed a significant increase in colitis at days 3 and 7 post-TNBS compared to the control group. Bar = 100 µm; n = 4 for each group. (E–F) P2X3 immunoreactivity in the muscular layer of the distal colon showed that TNBS-induced colitis significantly increased P2X3 expression compared to the control group. Bar = 50 µm; n = 4 for each group. *p < 0.05 and **p < 0.01 by one-way ANOVA followed by Newman–Keuls post hoc tests.