The multidrug resistance protein (MRP) 4 is expressed and functionally active in isolated bladder from pig.

Multidrug resistance proteins type 4 (MRP4) and 5 (MRP5) play pivotal roles in the transport of cyclic nucleotides in various tissues. However, their specific functions within the lower urinary tract remain relatively unexplored. This study aimed to investigate the effect of pharmacological inhibition of MRPs on cyclic nucleotide signaling in isolated pig bladder. The relaxation responses of the bladder were assessed in the presence of the MRP inhibitor, MK571. The temporal changes in intra- and extracellular levels of cAMP and cGMP in stimulated tissues were determined by mass spectrometry. The gene (ABCC4) and protein (MRP4) expression were also determined. MK571 administration resulted in a modest relaxation effect of approximately 26% in carbachol-pre-contracted bladders. The relaxation induced by phosphodiesterase inhibitors such as cilostazol, tadalafil, and sildenafil was significantly potentiated in the presence of MK571. In contrast, no significant potentiation was observed in the relaxation induced by substances elevating cAMP levels or stimulators of soluble guanylate cyclase. Following forskolin stimulation, both intracellular and extracellular cAMP concentrations increased by approximately 15.8-fold and 12-fold, respectively. Similarly, stimulation with tadalafil + BAY 41-2272 resulted in roughly 8.2-fold and 3.4-fold increases in intracellular and extracellular cGMP concentrations, respectively. The presence of MK571 reduced only the extracellular levels of cGMP. This study reveals the presence and function of MRP4 transporters within the porcine bladder and paves the way for future research exploring the role of this transporter in both underactive and overactive bladder disorders.

Inhibition of multidrug resistance proteins by MK571 restored the erectile function in obese mice through cGMP accumulation.

BACKGROUND: Intracellular levels of cyclic nucleotides can also be controlled by the action of multidrug resistance protein types 4 (MRP4) and 5 (MRP5). To date, no studies evaluated the role of their inhibition in an animal model of erectile dysfunction (ED). OBJECTIVES: To evaluate the effect of a 2-week treatment with MK571, an inhibitor of the efflux of cyclic nucleotides in the ED of obese mice. MATERIALS AND METHODS: Mice were divided in three groups: (i) lean, (ii) obese, and (iii) obese + MK571. The corpus cavernosum (CC) were isolated, and concentration-response curves to acetylcholine (ACh), sodium nitroprusside (SNP), and tadalafil in addition to electrical field stimulation (EFS) were carried out in phenylephrine pre-contracted tissues. Expression of ABCC4 and ABCC5, intracellular levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), the protein levels for pVASPSer157 and pVASPSer239 , and the intracavernous pressure (ICP) were also determined. The intracellular and extracellular (supernatant) ratios in CC from obese and lean stimulated with a cGMP-increasing substance (BAY 58-2667) in the absence and presence of MK571 (20 µM, 30 min) were also assessed. RESULTS: The treatment with MK571 completely reversed the lower relaxing responses induced by EFS, ACh, SNP, and tadalafil observed in obese mice CC in comparison with untreated obese mice. Cyclic GMP and p-VASPSer239 expression were significantly reduced in CC from obese groups. MK571 promoted a sixfold increase in cGMP without interfering in the protein expression of p-VASPSer239 . Neither the cAMP levels nor p-VASPSer157 were altered in MK571-treated animals. The ICP was ∼50% lower in obese than in the lean mice; however, the treatment with MK571 fully reversed this response. Expressions of ABCC4 and ABCC5 were not different between groups. The intra/extracellular ratio of cGMP was similar in CC from obese and lean mice stimulated with BAY 58-2667. CONCLUSIONS: The MRPs inhibition by MK571 favored the accumulation of cGMP in the smooth muscle cells, thus improving the smooth muscle relaxation and the erectile function in obese mice.

TRPA1 channel mediates methylglyoxal-induced mouse bladder dysfunction.

Introduction: The transient receptor potential ankyrin 1 channel (TRPA1) is expressed in urothelial cells and bladder nerve endings. Hyperglycemia in diabetic individuals induces accumulation of the highly reactive dicarbonyl compound methylglyoxal (MGO), which modulates TRPA1 activity. Long-term oral intake of MGO causes mouse bladder dysfunction. We hypothesized that TRPA1 takes part in the machinery that leads to MGO-induced bladder dysfunction. Therefore, we evaluated TRPA1 expression in the bladder and the effects of 1 h-intravesical infusion of the selective TRPA1 blocker HC-030031 (1 nmol/min) on MGO-induced cystometric alterations. Methods: Five-week-old female C57BL/6 mice received 0.5% MGO in their drinking water for 12 weeks, whereas control mice received tap water alone. Results: Compared to the control group, the protein levels and immunostaining for the MGO-derived hydroimidazolone isomer MG-H1 was increased in bladders of the MGO group, as observed in urothelium and detrusor smooth muscle. TRPA1 protein expression was significantly higher in bladder tissues of MGO compared to control group with TRPA1 immunostaining both lamina propria and urothelium, but not the detrusor smooth muscle. Void spot assays in conscious mice revealed an overactive bladder phenotype in MGO-treated mice characterized by increased number of voids and reduced volume per void. Filling cystometry in anaesthetized animals revealed an increased voiding frequency, reduced bladder capacity, and reduced voided volume in MGO compared to vehicle group, which were all reversed by HC-030031 infusion. Conclusion: TRPA1 activation is implicated in MGO-induced mouse overactive bladder. TRPA1 blockers may be useful to treat diabetic bladder dysfunction in individuals with high MGO levels.