In Silico Electrophysiological Investigation of Transient Receptor Potential Melastatin-4 Ion Channel Biophysics to Study Detrusor Overactivity.

Enhanced electrical activity in detrusor smooth muscle (DSM) cells is a key factor in detrusor overactivity which causes overactive bladder pathological disorders. Transient receptor potential melastatin-4 (TRPM4) channels, which are calcium-activated cation channels, play a role in regulating DSM electrical activities. These channels likely contribute to depolarizing the DSM cell membrane, leading to bladder overactivity. Our research focuses on understanding TRPM4 channel function in the DSM cells of mice, using computational modeling. We aimed to create a detailed computational model of the TRPM4 channel based on existing electrophysiological data. We employed a modified Hodgkin-Huxley model with an incorporated TRP-like current to simulate action potential firing in response to current and synaptic stimulus inputs. Validation against experimental data showed close agreement with our simulations. Our model is the first to analyze the TRPM4 channel's role in DSM electrical activity, potentially revealing insights into bladder overactivity. In conclusion, TRPM4 channels are pivotal in regulating human DSM function, and TRPM4 channel inhibitors could be promising targets for treating overactive bladder.

Low-Intensity Extracorporeal Shock Wave Therapy Ameliorates Detrusor Hyperactivity with Impaired Contractility via Transient Potential Vanilloid Channels: A Rat Model for Ovarian Hormone Deficiency.

This study explores low-intensity extracorporeal shock wave therapy (LiESWT)'s efficacy in alleviating detrusor hyperactivity with impaired contractility (DHIC) induced by ovarian hormone deficiency (OHD) in ovariectomized rats. The rats were categorized into the following four groups: sham group; OVX group, subjected to bilateral ovariectomy (OVX) for 12 months to induce OHD; OVX + SW4 group, underwent OHD for 12 months followed by 4 weeks of weekly LiESWT; and OVX + SW8 group, underwent OHD for 12 months followed by 8 weeks of weekly LiESWT. Cystometrogram studies and voiding behavior tracing were used to identify the symptoms of DHIC. Muscle strip contractility was evaluated through electrical-field, carbachol, ATP, and KCl stimulations. Western blot and immunofluorescence analyses were performed to assess the expressions of various markers related to bladder dysfunction. The OVX rats exhibited significant bladder deterioration and overactivity, alleviated by LiESWT. LiESWT modified transient receptor potential vanilloid (TRPV) channel expression, regulating calcium concentration and enhancing bladder capacity. It also elevated endoplasmic reticulum (ER) stress proteins, influencing ER-related Ca2+ channels and receptors to modulate detrusor muscle contractility. OHD after 12 months led to neuronal degeneration and reduced TRPV1 and TRPV4 channel activation. LiESWT demonstrated potential in enhancing angiogenic remodeling, neurogenesis, and receptor response, ameliorating DHIC via TRPV channels and cellular signaling in the OHD-induced DHIC rat model.

Predicting muscarinic receptor occupancy in human bladder mucosa from urinary concentrations of antimuscarinic agents for overactive bladder.

To assess the pharmacologically relevant and selective muscarinic receptor occupancy in the bladder mucosa, we considered not only plasma drug concentrations but also urinary drug concentrations. The purpose of this study was to predict muscarinic receptor occupancy in the human bladder mucosa based on urinary concentrations in response to clinical dosages of antimuscarinic agents used to treat overactive bladder. The calculated mean plasma or serum unbound steady state concentrations were 0.06-11 nM in clinical dosages of five antimuscarinic agents. Urinary concentrations calculated from the mean plasma or serum and renal clearance ranged between 19 nM and 2 µM, which were >10-fold higher than the Ki values for bladder muscarinic receptors excluding propiverine. Bladder mucosal muscarinic receptor occupancy estimated from the urinary concentrations and the Ki values was >90 % at a steady state in clinical dosages of five antimuscarinic agents. The bladder muscarinic receptor occupancy was higher than that in the parotid gland calculated based on the mean plasma or serum unbound concentrations and Ki values for muscarinic receptors in the parotid gland. These results suggest that sufficient and selective muscarinic receptor occupancy by antimuscarinic agents, to exert pharmacological effects, in the bladder mucosa can be predicted using urinary concentrations.

Transient receptor potential ankyrin 1 channels in the bladder mediate low temperature elicited bladder overactivity in rats.

AIMS: This study aimed to investigate whether pathways involving transient receptor potential ankyrin 1 (TRPA1) channels in the urinary bladder mediate the bladder overactivity elicited by exposure to a low temperature in rats. METHODS: At postnatal week 10, female Sprague-Dawley (SD) rats were intraperitoneally injected with the TRPA1 channel antagonist, HC030031, at room temperature (RT) and subsequently exposed to low temperature (LT). Bladder specimens treated with HC030031 were evaluated for contractions through cumulative addition of the TRPA1 channel agonist trans-cinnamaldehyde. Two days before cystometric investigation, small interfering RNA (siRNA) targeting TRPA1 was transfected into urinary bladders. Then, cystometric investigations were performed on rats subjected to TRPA1 siRNA transfection at both RT and LT. Expression of TRPA1 channels in the urinary bladder was assessed through immunohistochemistry and real-time reverse transcription-polymerase chain reaction. RESULTS: At RT, micturition patterns were unaffected by HC030031 treatment. However, upon exposure to LT, rats treated with HC030031 exhibited a reduction of LT-elicited bladder overactivity, as evidenced by inhibited decreases in voiding interval, micturition volume, and bladder capacity. Additionally, HC030031 inhibited trans-cinnamaldehyde-induced contractions. Immunohistochemical analysis showed the presence of TRPA1 channels in the urinary bladder. Notably, rats with TRPA1 siRNA-transfected bladders could partially inhibit bladder overactivity during LT exposure. CONCLUSIONS: These findings indicate that pathways involving TRPA1 channels expressed in the urinary bladder could mediate the LT-elicited bladder overactivity.

Multi-omic approaches provide insights into the molecular mechanisms of Sojae semen germinatum water extract against overactive bladder.

Sojae semen germinatum (SSG) is derived from mature soybean seeds that have been germinated and dried, typically with sprouts measuring approximately 0.5 cm in length. SSG is traditionally known for its properties in clearing heat and moisture. Nevertheless, limited information was reported on the effects and mechanisms of SSG in alleviating urinary symptoms. This study employed urodynamic parameters to investigate the therapeutic effect of SSG water extract on overactive bladder (OAB) in the rat model with benign prostatic hyperplasia. Through a combination of transcriptomic and metabolomic analyses, the pathways and key proteins of the SSG treatment for OAB were identified and validated by ELISA and Western blotting. Furthermore, network pharmacology elucidated the roles of SSG's isoflavones acting on the target which was identified by above-mentioned multi-omics analysis. Our results indicate that SSG water extract significantly mitigated OAB by down-regulating the PGE2/EP1/PLCß2/p-MLC signaling pathway. It was speculated that the active ingredient in the SSG on EP1 was genistein. This study provided valuable insights into the molecular mechanisms of SSG water extract, emphasizing the multi-target characteristics and critical pathways in improving OAB. Furthermore, this study contributes to the potential utilization of SSG as a functional food.

The Assessment of the Efficacy of Imperatorin in Reducing Overactive Bladder Symptoms.

Overactive bladder syndrome (OAB) is a prevalent condition that affects the elderly population in particular and significantly impairs quality of life. Imperatorin, a naturally occurring furocoumarin, possesses diverse pharmacological properties that warrant consideration for drug development. The aim of this study was to investigate the potential of imperatorin (IMP) to attenuate the cystometric and biochemical changes typically associated with retinyl acetate-induced overactive bladder (OAB) and to assess its viability as a pharmacological intervention for OAB patients. A total of 60 rats were divided into four groups: I-control, II-rats with rapamycin (RA)-induced OAB, III-rats administered IMP at a dose of 10 mg/kg/day, and IV-rats with RA-induced OAB treated with IMP. IMP or vehicle were injected intraperitoneally for 14 days. The cystometry and assessment of bladder blood flow were performed two days after the last dose of IMP. The rats were then placed in metabolic cages for 24 h. Urothelial thickness measurements and biochemical analyses were performed. Intravesical infusion of RA induced OAB. Notably, intraperitoneal administration of imperatorin had no discernible effect on urinary bladder function and micturition cycles in normal rats. IMP attenuated the severity of RA-induced OAB. RA induced increases in urothelial ATP, calcitonin gene-related peptide (CGRP), organic cation transporter 3 (OCT3), and vesicular acetylcholine transporter (VAChT), as well as significant c-Fos expression in all micturition areas analyzed, which were attenuated by IMP. Furthermore, elevated levels of Rho kinase (ROCK1) and VAChT were observed in the detrusor, which were reversed by IMP in the context of RA-induced OAB in the urothelium, detrusor muscle, and urine. Imperatorin has a mitigating effect on detrusor overactivity. The mechanisms of action of IMP in the bladder appear to be diverse and complex. These findings suggest that IMP may provide protection against RA-induced OAB and could potentially develop into an innovative therapeutic strategy for the treatment of OAB.

Activation of the adenosine A1 receptor in the lumbosacral spinal cord improves bladder overactivity in rats with cystitis induced by cyclophosphamide.

PURPOSE: To investigate the effect of intrathecal administration of CCPA, an adenosine A1 receptor agonist, on voiding function in rats with cystitis induced by cyclophosphamide (CYP). METHODS: Thirty 8-week-old Sprague Dawley rats were randomly divided into a control group (n = 15) and a cystitis group (n = 15). Cystitis was induced by a single intraperitoneal injection of CYP (200 mg/kg, dissolved in physiological saline) in rats. Control rats were injected intraperitoneally with physiological saline. The PE10 catheter reached the level of L6-S1 spinal cord through L3-4 intervertebral space for intrathecal injection. Forty-eight hours after intraperitoneal injection, urodynamic tests were conducted to observe the effect of intrathecal administration of 10% dimethylsulfoxide (vehicle) and 1 nmol CCPA on micturition parameters, including basal pressure (BP), threshold pressure (TP), maximal voiding pressure (MVP), intercontraction interval (ICI), voided volume (VV), residual volume (RV), bladder capacity (BC), and voiding efficiency (VE). Histological changes of the bladder of cystitis rats were studied through hematoxylin-eosin staining (HE staining). Moreover, Western blot and immunofluorescence were used to study the expression of adenosine A1 receptor in the L6-S1 dorsal spinal cord in both groups of rats. RESULTS: HE staining revealed submucosal hemorrhage, edema, and inflammatory cell infiltration in the bladder wall of cystitis rats. The urodynamic test showed significant increase in BP, TP, MVP and RV in cystitis rats, while ICI, VV, BC and VE decreased significantly, indicating bladder overactivity. CCPA inhibited the micturition reflex in both control and cystitis rats, and significantly increased TP, ICI, VV, BC, and VE, but had no significant effect on BP, MVP and RV. Western blot and immunofluorescence showed that there was no significant difference in the expression of adenosine A1 receptor in the L6-S1 dorsal spinal cord between the control and cystitis rats. CONCLUSION: The findings of this study suggest that intrathecal administration of the adenosine A1 receptor agonist CCPA alleviates CYP-induced bladder overactivity. Furthermore, our results indicate that the adenosine A1 receptor in the lumbosacral spinal cord may be a promising target for treatment of bladder overactivity.

Involvement of Mast-Cell-Tryptase- and Protease-Activated Receptor 2-Mediated Signaling and Urothelial Barrier Dysfunction with Reduced Uroplakin II Expression in Bladder Hyperactivity Induced by Chronic Bladder Ischemia in the Rat.

We aimed to investigate the relationship between mast cell (MC) infiltration into the bladder with urothelial barrier dysfunction and bladder hyperactivity in a chronic bladder ischemia (CBI) rat model. We compared CBI rats (CBI group; n = 10) with normal rats (control group; n = 10). We measured the expression of mast cell tryptase (MCT) and protease-activated receptor 2 (PAR2), which are correlated with C fiber activation via MCT, and Uroplakins (UP Ia, Ib, II and III), which are critical to urothelial barrier function, via Western blotting. The effects of FSLLRY-NH2, a PAR2 antagonist, administered intravenously, on the bladder function of CBI rats were evaluated with a cystometrogram. In the CBI group, the MC number in the bladder was significantly greater (p = 0.03), and the expression of MCT (p = 0.02) and PAR2 (p = 0.02) was significantly increased compared to that of the control group. The 10 µg/kg FSLLRY-NH2 injection significantly increased the micturition interval of CBI rats (p = 0.03). The percentage of UP-II-positive cells on the urothelium with immunohistochemical staining was significantly lower in the CBI group than in the control group (p < 0.01). Chronic ischemia induces urothelial barrier dysfunction via impairing UP II, consequently inducing MC infiltration into the bladder wall and increased PAR2 expression. PAR2 activation by MCT may contribute to bladder hyperactivity.

Application of D-Optimal Mixture Design in the Development of Nanocarrier-Based Darifenacin Hydrobromide Gel.

BACKGROUND: Darifenacin hydrobromide, a BCS Class II drug, is poorly bioavailable due to extensive first-pass metabolism. The present study is an attempt to investigate an alternative route of drug delivery by developing a nanometric microemulsion-based transdermal gel for the management of an overactive bladder. METHODS: Oil, surfactant, and cosurfactant were selected based on the solubility of the drug, and surfactant: cosurfactant in surfactant mixture (Smix) was selected at a 1:1 ratio as inferred from the pseudo ternary phase diagram. The D-optimal mixture design was used to optimize the o/w microemulsion wherein the globule size and zeta potential were selected as dependable variables. The prepared microemulsions were also characterized for various physico-chemical properties like transmittance, conductivity, and TEM. The optimized microemulsion was gelled using Carbopol 934 P and assessed for drug release in vitro and ex vivo, viscosity, spreadability, pH, etc. RESULTS: Drug excipient compatibility studies showed that the drug was compatible with formulation components. The optimized microemulsion showed a globule size of less than 50 nm and a high zeta potential of -20.56 mV. The ME gel could sustain the drug release for 8 hours as reflected in in vitro and ex vivo skin permeation and retention studies. The accelerated stability study showed no significant change in applied storage conditions. CONCLUSION: An effective, stable, non-invasive microemulsion gel containing darifenacin hydrobromide was developed. The achieved merits could translate into increased bioavailability and dose reduction. Further confirmatory in vivo studies on this novel formulation, which is a cost-effective & industrially scalable option, can improve the pharmacoeconomics of overactive bladder management.

Development of Neurogenic Detrusor Overactivity after Thoracic Spinal Cord Injury Is Accompanied by Time-Dependent Changes in Lumbosacral Expression of Axonal Growth Regulators.

Thoracic spinal cord injury (SCI) results in urinary dysfunction, which majorly affects the quality of life of SCI patients. Abnormal sprouting of lumbosacral bladder afferents plays a crucial role in this condition. Underlying mechanisms may include changes in expression of regulators of axonal growth, including chondroitin sulphate proteoglycans (CSPGs), myelin-associated inhibitors (MAIs) and repulsive guidance molecules, known to be upregulated at the injury site post SCI. Here, we confirmed lumbosacral upregulation of the growth-associated protein GAP43 in SCI animals with bladder dysfunction, indicating the occurrence of axonal sprouting. Neurocan and Phosphacan (CSPGs), as well as Nogo-A (MAI), at the same spinal segments were upregulated 7 days post injury (dpi) but returned to baseline values 28 dpi. In turn, qPCR analysis of the mRNA levels for receptors of those repulsive molecules in dorsal root ganglia (DRG) neurons showed a time-dependent decrease in receptor expression. In vitro assays with DRG neurons from SCI rats demonstrated that exposure to high levels of NGF downregulated the expression of some, but not all, receptors for those regulators of axonal growth. The present results, therefore, show significant molecular changes at the lumbosacral cord and DRGs after thoracic lesion, likely critically involved in neuroplastic events leading to urinary impairment.

Vibegron inhibits enhanced spontaneous contractions induced by anoxia/reoxygenation in isolated whole bladder from rats.

It has been recently proposed that repeated bladder ischemia/reperfusion induced by chronic pelvic ischemia may lead to detrusor overactivity, followed by lower urinary tract symptoms. Vibegron is a selective ß3-adrenoceptor agonist approved for the treatment of overactive bladder. Several studies have tested ß3-adrenoceptor agonists using animal models with detrusor overactivity related to bladder ischemia/reperfusion. However, whether ß3-adrenoceptor agonists directly affect ischemia/reperfusion-evoked detrusor overactivity is unclear. Therefore, we examined whether bladder anoxia/reoxygenation could enhance spontaneous bladder contractions (SBCs) and investigated the effect of vibegron on enhanced SBCs. Isolated whole bladders from rats were incubated with Krebs solution aerated with 95% N2 + 5% CO2 for 5 h (anoxia). Subsequently, the bathing solution was replaced with an oxygen-saturated solution (reoxygenation). Anoxia/reoxygenation caused enhancement of the amplitude but not the frequency of SBC compared with that before reoxygenation. Vibegron (0.3-30 µM) inhibited this increase in SBC amplitude, but not the frequency, in a dose-dependent manner. The inhibitory effect of vibegron was not affected by pretreatment with the adenylyl cyclase inhibitor SQ22536 (100 µM) or protein kinase A inhibitor KT5720 (1 µM) and was not accompanied by considerable changes in cyclic adenosine monophosphate (cAMP) content in the bladder. In contrast, the large conductance potassium channel inhibitor iberiotoxin (100 nM) suppressed the inhibitory effect of vibegron. These results suggest that bladder ischemia/reperfusion induces SBC enhancement and vibegron directly inhibits detrusor overactivity via the large conductance potassium channel, which involves ß3-adrenoceptor, rather than the cAMP signaling pathway.

Oxybutynin nanosuspension gel for enhanced transdermal treatment for overactive bladder syndrome.

Oxybutynin (OXY) is the most common drug to treat overactive bladder (OAB) syndrome. Transdermal administration is a more ideal route replacing oral administration to resolve problems of low bioavailability and severe side effects. However, commercial transdermal products of OXY frequently cause skin irritation and low permeation efficiency arising discontinued medication. Here, oxybutynin nanosuspension (OXY-NS) and its gel preparation (OXY-NG) were constructed to resolve these issues. In vitro permeation test and in vivo pharmacokinetics study confirmed that OXY-NG significantly enhanced the transdermal permeation of OXY, about 4-fold and 3-fold higher than oxybutynin coarse suspension (OXY-CG), respectively, and in vitro retention test certified that OXY-NG increased OXY concentration especially in viable epidermis (VE) and Dermis (about 3 times that of OXY-CG), consequently improving the bioavailability. Skin irritation assay demonstrated that OXY-NG would not trigger skin adverse effects. In addition, selectively blocking hair follicles test evidenced that hair follicles pathway played an important role in OXY-NS transdermal delivery. In general, by virtue of excellent drug loading, low toxicity and ease of scale-up, OXY-NG is a promising strategy to ameliorate skin permeation of insoluble OXY for better transdermal treatment for OAB, hence increasing its bioavailability, reducing adverse effects, and achieving good patient compliance.

Moxibustion attenuates neurogenic detrusor overactivity in spinal cord injury rats by inhibiting M2/ATP/P2X3 pathway.

PURPOSE: Activation of muscarinic receptors located in bladder sensory pathways is generally considered to be the primary contributor for driving the pathogenesis of neurogenic detrusor overactivity following spinal cord injury. The present study is undertaken to examine whether moxibustion improves neurogenic detrusor overactivity via modulating the abnormal muscarinic receptor pathway. MATERIALS AND METHODS: Female Sprague-Dawley rats were subjected to spinal cord injury with T9-10 spinal cord transection. Fourteen days later, animals were received moxibustion treatment for one week. Urodynamic parameters and pelvic afferents discharge were measured. Adenosine triphosphate (ATP) content in the voided cystometry fluid was determined. Expressions of M2, M3, and P2X3 receptors in the bladder mucosa were evaluated. RESULTS: Moxibustion treatment prevented the development of detrusor overactivity in spinal cord injury rats, with an increase in the intercontraction interval and micturition pressure threshold and a decrease in afferent activity during filling. The expression of M2 was markedly suppressed by moxibustion, accompanied by a reduction in the levels of ATP and P2X3. M2 receptor antagonist methoctramine hemihydrate had similar effects to moxibustion on bladder function and afferent activity, while the M2-preferential agonist oxotremorine methiodide abolished the beneficial effects of moxibustion. CONCLUSION: Moxibustion is a potential candidate for treating neurogenic bladder overactivity in a rat model of spinal cord injury, possibly through inhibiting the M2/ATP/P2X3 pathway.

Inhibition of Full Smooth Muscle Contraction in Isolated Human Detrusor Tissues by Mirabegron Is Limited to Off-Target Inhibition of Neurogenic Contractions.

Mirabegron is used for treatment of storage symptoms in overactive bladder (OAB) caused by spontaneous bladder smooth muscle contractions. However, owing to limitations in available studies using human tissues, central questions are still unresolved, including mechanisms underlying improvements by mirabegron and its anticontractile effects in the detrusor. Here, we assessed concentration-dependent mirabegron effects on contractions of human detrusor tissues in frequency-response curves and concentration-response curves for different cholinergic and noncholinergic agonists. Detrusor tissues were sampled from patients undergoing radical cystectomy. Contractions were induced by electric field stimulation (EFS) and by cumulative concentrations of cholinergic agonists, endothelin-1, and the thromboxane A2 analog U46619. EFS-induced contractions were inhibited using 10 µM mirabegron, but not using 1 µM. Inhibition by 10 µM mirabegron was resistant to the ß 3-adrenergic antagonist L-748,337. Concentration-dependent contractions by carbachol were not inhibited by 1 µM or 10 µM mirabegron. Concentration-response curves for methacholine were slightly right-shifted by 10 µM, but not 1 µM mirabegron. Concentration-dependent contractions by endothelin-1 or U46619 were not changed by mirabegron. In contrast, the muscarinic antagonist tolterodine right-shifted concentration-response curves for carbachol and methacholine and inhibited EFS-induced contractions. In conclusion, inhibition of neurogenic contractions in isolated detrusor tissues by mirabegron requires concentrations highly exceeding known plasma levels during standard dosing and the known binding constant (Ki values) for ß 3-adrenoceptors. Full contractions by cholinergic agonists, endothelin-1, and U46619 are not affected by therapeutic concentrations of mirabegron. Improvements of storage symptoms are most likely not imparted by inhibition of ß 3-adrenoceptors in the bladder wall itself. SIGNIFICANCE STATEMENT: Mirabegron is used for overactive bladder (OAB) treatment, but the underlying mechanisms are unclear, and preclinical and clinical findings are controversial due to limitations in available studies. Our findings suggest that inhibition of detrusor contractions by mirabegron is limited to neurogenic contractions, which requires unphysiologic concentrations and does not involve ß 3-adrenoceptors. Mechanisms accounting for improvements of OAB by mirabegron are located outside the urinary bladder.

Reviving Cav1.2 as an attractive drug target to treat bladder dysfunction.

Inhibition of bladder contraction with antimuscarinics is a common approach to treat bladder hyperactivity, and the L-type voltage-gated calcium channel α1C (Cav1.2) is crucial for bladder contractility. Therefore, strategies aimed at inhibiting Cav1.2 appear warranted. However, multiple clinical trials that attempted to treat bladder overactivity with calcium channel blockers (CCBs) have been unsuccessful, creating an unsolved mystery. In contrast, cardiologists and epidemiologists have reported strong associations between CCB use and bladder hyperactivity, opposing expectations of urologists. Recent findings from our lab offer a potential explanation. We have demonstrated that ketamine which can cause cystitis, functions, like nifedipine, as a Cav1.2 antagonist. We also show that a Cav1.2 agonist which potentiates muscle contraction, rather than antagonizing it, can increase the volume of voids and reduce voiding frequency. This perspective will discuss in detail the unsuccessful urological trials of CCBs and the promise of Cav1.2 agonists as potential novel therapies for bladder dysfunctions.

Increased transient receptor potential canonical 3 activity is involved in the pathogenesis of detrusor overactivity by dynamic interaction with Na+/Ca2+ exchanger 1.

Transient receptor potential canonical 3 (TRPC3) is a nonselective cation channel, and its dysfunction is the basis of many clinical diseases. However, little is known about its possible role in the bladder. The purpose of this study was to explore the function and mechanism of TRPC3 in partial bladder outlet obstruction (PBOO)-induced detrusor overactivity (DO). We studied 31 adult female rats with DO induced by PBOO (the DO group) and 40 sham-operated rats (the control group). Here we report that the expression of TRPC3 in the bladder of DO rats increased significantly. Furthermore, PYR10, which can selectively inhibit the TRPC3 channel, significantly reduced bladder excitability in DO and control rats, but the decrease of the bladder excitability of DO rats was more obvious. PYR10 significantly reduced the intracellular calcium concentration in smooth muscle cells (SMCs) in DO and control rats. Finally, Na+/Ca2+ exchanger 1 (NCX1) colocalizes with TRPC3 and affects its expression and function. Collectively, these results indicate that TRPC3 plays an important role in the pathogenesis of DO through a synergistic effect with NCX1. TRPC3 and NCX1 may be new therapeutic targets for DO.

Therapeutic effects of nerve growth factor-targeting therapy on bladder overactivity in rats with prostatic inflammation.

BACKGROUND: The present study examined the effect of liposomes conjugated with antisense oligonucleotide of nerve growth factor (NGF-OND) on local overexpression of NGF and bladder overactivity using rats with prostatic inflammation (PI). METHODS: Male Sprague-Dawley rats were divided into three groups: (1) Control group; intact rats, (2) PI-NS group; rats with PI and intravesical instillation of normal saline (NS), (3) PI-OND group; rats with PI and intravesical instillation of NGF-OND. On Day 0, PI was induced by intraprostatic 5%-formalin injection. On Day 14, NGF-OND or NS was instilled directly into the bladder after laparotomy. On Day 28, therapeutic effects of NGF-OND were evaluated by awake cystometry and histological analysis as well as reverse-transcription polymerase chain reaction measurements of messenger RNA (mRNA) levels of NGF in the bladder and prostate, inflammatory markers in the prostate, C-fiber afferent markers, and an A-type K+ channel α-subunit (Kv 1.4) in L6-S1 dorsal root ganglia (DRG). RESULTS: Intravesical NFG-OND treatment reduced PI-induced overexpression of NGF in both bladder and prostate, and reduced PI-induced bladder overactivity evident as longer intercontraction intervals in association with reductions of TRPV1 and TRPA1 mRNA expression levels in DRG. mRNA expression of Kv1.4 in DRG was reduced after PI, but improved in the PI-OND group. CONCLUSIONS: These results indicate that NGF locally expressed in the bladder is an important mediator inducing bladder overactivity with upregulation of C-fiber afferent markers and downregulation of an A-type K+ channel subunit in DRG following PI, and that liposome-based, local NGF-targeting therapy could be effective for not only bladder overactivity and afferent sensitization, but also PI. Thus, local blockade of NGF in the bladder could be a therapeutic modality for male LUTS due to BPH with PI.

Tadalafil ameliorates bladder overactivity by restoring insulin-activated detrusor relaxation via the bladder mucosal IRS/PI3K/AKT/eNOS pathway in fructose-fed rats.

The pathophysiologies of metabolic syndrome (MS) and overactive bladder (OAB) might overlap. Using fructose-fed rats (FFRs) as a rodent model of MS we investigated the effects of tadalafil (a phosphodiesterase type 5 inhibitor) on the dysregulated insulin signalling in the bladder mucosa and bladder overactivity. Micturition behaviour was evaluated. Concentration-response curves on detrusor relaxation to insulin stimulation were examined. Expression and phosphorylation of proteins in the insulin signalling pathway were evaluated by Western blotting. Levels of detrusor cGMP and urinary nitrite and nitrate (NOx) were measured. We observed FFRs exhibited metabolic traits of MS, bladder overactivity, and impaired insulin-activated detrusor relaxation in organ bath study. A high-fructose diet also impeded insulin signalling, reflected by overexpression of IRS1/pIRS1Ser307 and pIRS2Ser731 and downregulation of PI3K/pPI3KTyr508, AKT/pAKTSer473, and eNOS/peNOSSer1177 in the bladder mucosa, alongside decreased urinary NOx and detrusor cGMP levels. Tadalafil treatment restored the reduced level of mucosal peNOS, urinary NOx, and detrusor cGMP, improved the insulin-activated detrusor relaxation, and ameliorated bladder overactivity in FFRs. These results suggest tadalafil may ameliorate MS-associated bladder overactivity by restoring insulin-activated detrusor relaxation via molecular mechanisms that are associated with preservation of IR/IRS/PI3K/AKT/eNOS pathway in the bladder mucosa and cGMP production in the bladder detrusor.

Aging Changes in Bladder Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels Are Associated With Increasing Heterogeneity of Adrenergic/Mucosal Influence on Detrusor Control in the Mouse.

A geroscience-informed approach to the increasing prevalence of bladder control problems in older adults requires understanding the impact of aging on dynamic mechanisms that ensure resilience in response to stressors challenging asymptomatic voluntary control over urine storage and voiding. Bladder control is predicated on sensory neural information about bladder volume. Modulation of volume-induced bladder wall tensions by autonomic and mucosal factors controls neural sensitivity to bladder volume. We hypothesized that hyperpolarization-activated cyclic nucleotide-gated (HCN) channels integrate these factors and thereby mediate adrenergic detrusor tension control. Furthermore, loss of HCN expression compromises that integration and could result in loss of precision of detrusor control. Using a life-span mouse model, reverse transcription quantitative real-time PCR and pharmacologic studies in pretensioned intact and mucosa-denuded bladder strips were made. The dominant hcn1 expression declines with maturation and aging; however, aging is also associated with increased variance around mean values. In strips from Mature animals, isoproterenol had less effect in denuded muscle strips than in intact strips, and HCN blockade diminished isoproterenol responsiveness. With aging, variances about mean response values significantly increased, paralleling hcn1 expression. Our findings support a role for HCN in providing neuroendocrine/paracrine integration and suggest an association of increased heterogeneity of HCN expression in aging with reductions in response precision to neuroendocrine control. The functional implication is an increased risk of dysfunction of brainstem/bladder regulation of neuronal sensitivity to bladder volume. This supports the clinical model of the aging bladder phenotype as an expression of loss of resilience, and not as emerging bladder pathology with aging.

Efficacy of the combination of KPR-5714, a novel transient receptor potential melastatin 8 (TRPM8) antagonist, and ß3-adrenoceptor agonist or anticholinergic agent on bladder dysfunction in rats with bladder overactivity.

Transient receptor potential melastatin 8 (TRPM8) channels may contribute to the pathophysiological bladder afferent hyperactivity, thus a TRPM8 antagonist would be a promising therapeutic target for the bladder hypersensitive disorders including urinary urgency in overactive bladder (OAB). We aimed to investigate a pharmacological effect of KPR-5714, a novel selective TRPM8 antagonist, on TRPM8 channels, M3 receptors and ß3-adrenoceptors using the transfected cells of each gene at first. Then, combination effects of KPR-5714 and mirabegron, a ß3-adrenoceptor agonist, or tolterodine tartrate, an anticholinergic agent, were studied on rhythmic bladder contractions (RBCs) in normal rats and bladder function in frequent-voiding rats. In vitro measurements showed that KPR-5714 acts on neither ß3-adrenoceptor nor M3 receptor. In normal rats, KPR-5714 and mirabegron significantly reduced the frequency of RBCs, and a combined administration showed an additive effect. In rats with cerebral infarction, KPR-5714 and mirabegron significantly reduced the voiding frequency, and a combined administration showed an additive effect. In rats exposed to cold temperature, KPR-5714 and tolterodine tartrate significantly reduced the voiding frequency accompanied by the increased mean voided volume, and a combined administration showed additive effects. The present study demonstrated that the combined administration of KPR-5714 and mirabegron or tolterodine tartrate showed the additive effects on bladder dysfunction in different animal models, suggesting that the combination therapy of TRPM8 antagonist and ß3-adrenoceptor agonist or anticholinergic agent can be the potential treatment option for obtaining additive effects in comparison with monotherapy for OAB.