Activation of P2Y₆ receptors increases the voiding frequency in anaesthetized rats by releasing ATP from the bladder urothelium

Abstract

Background and purpose Despite the abundant expression of the UDP‐sensitive P2Y6 receptor in urothelial cells and sub‐urothelial myofibroblasts its role in the control of bladder function is not well understood. Experimental approach We compared the effects of UDP and of the selective P2Y6 receptor agonist, PSB0474, on bladder urodynamics in anaesthetized rats; the voided fluid was tested for ATP bioluminescence. The isolated urinary bladder was used for in vitro myographic recordings and [3H]‐ACh overflow experiments. Key results Instillation of UDP or PSB0474 into the bladder increased the voiding frequency (VF) without affecting the amplitude (A) and the duration (Δt) of bladder contractions; an effect blocked by the P2Y6 receptor antagonist, MRS2578. Effects mediated by urothelial P2Y6 receptors required extrinsic neuronal circuitry as they were not detected in the isolated bladder. UDP‐induced bladder hyperactvity was also prevented by blocking P2X3 and P2Y1 receptors, respectively, with A317491 and MRS2179 applied i.v.. UDP decreased [3H]‐ACh release from stimulated bladder strips with urothelium, but not in its absence. Inhibitory effects of UDP were converted into facilitation by the P2Y1 receptor antagonist, MRS2179. The P2Y6 receptor agonist increased threefold ATP levels in the voided fluid. Conclusions and Implications Activation of P2Y6 receptors increased the voiding frequency indirectly by releasing ATP from the urothelium and activation of P2X3 receptors on sub‐urothelial nerve afferents. Bladder hyperactivity may be partly reversed following ATP hydrolysis to ADP by E‐NTPDases, thereby decreasing ACh release from cholinergic nerves expressing P2Y1 receptors.

Impairment of ATP hydrolysis decreases adenosine A1 receptor tonus favoring cholinergic nerve hyperactivity in the obstructed human urinary bladder

“…In control experiments, aliquots of matched samples were immediately freeze-dried in liquid nitrogen and preserved at −80°C for subsequent ATP quantification by bioluminescence using the luciferin-luciferase ATP kit HS II (Roche Applied Science, Indianapolis, USA) according to manufacturer's instructions. Luminescence was determined using a multi-detection microplate reader (Synergy HT; BioTek Instruments) [23][24][25].…”

“…Therefore, the evoked [ 3 H]ACh release was calculated by subtracting the basal tritium outflow from the total tritium outflow during the stimulation period (see, e.g., [21][22][23]). Likewise, stimulation-evoked release of ATP was calculated by subtracting the basal release, measured in the sample collected before stimulation, from the total release of the nucleotide determined after stimulus application [23][24][25].…”

Attenuated Aortic Vasodilation and Sympathetic Prejunctional Facilitation in Epinephrine-Deficient Mice: Selective Impairment of β2-Adrenoceptor Responses

“…Vectashield with DAPI (49,6-diamidino-2-phenylindole) was used as mounting media (Vector Laboratories, Burlingame, CA). Observations and analyses were performed with a laser-scanning confocal microscope (FV1000; Olympus Fluoview, Tokyo, Japan) (Carneiro et al, 2014).…”

Investigating the associations of mucosal P2Y6 receptor expression and urinary ATP and ADP concentrations, with symptoms of overactive bladder

“…We investigated the expression of P2Y6 purinoceptors in the mucosa layer of the human bladder, given compelling evidence from animal studies of modulation of nonneuronal ATP release by P2Y6 9,10 . P2Y6 immunofluorescence was observed in urothelial and suburothelial layers of human mucosa (Figure 2).…”