Urethral Relaxation Research Articles

128 - The effects of a novel photo-reactive NO-donor “NORD-1” on internal urethral relaxation using an intrapelvic nerve injury rat model.

128 - The effects of a novel photo-reactive NO-donor “NORD-1” on internal urethral relaxation using an intrapelvic nerve injury rat model.

Time-dependent progress of lower urinary tract dysfunction in streptozotocin-induced diabetic rats with or without low-dose insulin treatment.

Objectives: To examine time-dependent functional and structural changes of the lower urinary tract in streptozotocin-induced diabetic rats with or without low-dose insulin treatment and explore the pathophysiological characteristics of insulin therapy on lower urinary tract dysfunction (LUTD) caused by diabetes mellitus (DM). Methods: Female Sprague-Dawley rats were divided into five groups: normal control (NC) group, 4 weeks insulin-treated DM (4-DI) group, 4 weeks DM (4-DM) group, 8 weeks insulin-treated DM (8-DI) group and 8 weeks DM (8-DM) group. DM was initially induced by i.p. injection of streptozotocin (65 mg/kg), and then the DI groups received subcutaneous implantation of insulin pellets under the mid dorsal skin. Voiding behavior was evaluated in metabolic cages. The function of bladder and urethra in vivo were evaluated by simultaneous recordings of the cystometrogram and urethral perfusion pressure (UPP) under urethane anesthesia. The function of bladder and urethra in vitro were tested by organ bath techniques. The morphologic changes of the bladder and urethra were investigated using Hematoxylin-Eosin and Masson's staining. Results: Both 4-and 8-weeks diabetic rats have altered micturition patterns, including increased 12-h urine volume, urinary frequency/12 hours and voided volume. In-vivo urodynamics showed the EUS bursting activity duration is longer in 4-DM group and shorter in 8-DM group compared to NC group. UPP change in 8-DM were significantly lower than NC group. While none of these changes were found between DI and NC groups. Organ bath showed the response to Carbachol and EFS in bladder smooth muscle per tissue weights was decreased significantly in 4- and 8-weeks DM groups compared with insulin-treated DM or NC groups. In contrast, the contraction of urethral muscle and maximum urethral muscle contraction per gram of the tissue to EFS stimulation were significantly increased in 4- and 8-weeks DM groups. The thickness of bladder smooth muscle was time-dependently increased, but the thickness of the urethral muscle had no difference. Conclusions: DM-induced LUTD is characterized by time-dependent functional and structural remodeling in the bladder and urethra, which shows the hypertrophy of the bladder smooth muscle, reduced urethral smooth muscle relaxation and EUS dysfunction. Low-dose insulin can protect against diuresis-induced bladder over-distention, preserve urethral relaxation and protect EUS bursting activity, which would be helpful to study the slow-onset, time-dependent progress of DM-induced LUTD.

The potential involvement of MRP5 pump in urethral dysfunction in streptozotocin-induced diabetic rats.

To examine the effects of i.v. administration of MK-571, a MRP4/5 pump inhibitor, on urethral function in the urethane-anesthetized rat, and the changes of urethral multidrug resistance protein 5 (MRP5) pump in streptozotocin (STZ)-induced diabetes mellitus (DM) rats. Isovolumetric cystometry and urethral perfusion pressure (UPP) measurements were carried out in normal control (NC) group and 8week DM groups under urethane anesthesia. When stable rhythmic bladder contractions were showed, UPP parameters were recorded after successive administration of various dose of MK-571. Additionally, urethral cyclic guanosine monophosphate (cGMP) protein level was evaluated by ELISA, and changes of MRP5 pump and neurogenic nitric oxide synthase (nNOs) in the urethra were examined with immunohistochemical staining and Western blot analysis. In NC group, UPPnadir was significantly decreased but UPP change increased after administration of MK-571, while no significant differences in UPP parameters were observed in 8-week DM group. Furthermore, urethral MRP5 protein level was up-regulated, whereas urethral cGMP and nNOS protein levels were down-regulated in 8-week DM group. MK-571 could not restore NO-mediated urethral relaxation dysfunction in DM rats, which may be attributed to the up-regulation of urethral MRP5 pump, and thus decrease of intracellular cGMP concentration in the urethra. These novel results would be useful for a better understanding of DM-related lower urinary tract dysfunction LUT (LUTD). Also, they could be helpful to study the importance of MRP pumps in the control of urethral relaxation mechanisms under physiological and pathological states.

Mild thermal stimulation of the buttock skin increases urinary voiding efficiency in anesthetized rats.

In the present study, we examined the effects of mild thermal stimulation of the skin on voiding efficiency using urethane-anesthetized rats with reduced voiding efficiency. Spontaneous urination was induced by infusing saline. For each voiding, the voiding efficiency was calculated from the voided volume and the bladder capacity measured. A Peltier thermode was attached to the buttock skin to apply stimulation: cooling between to 25°C and 35°C, every 20s throughout the saline infusion. The voiding efficiency was 29±9% (mean±SD) before stimulation and increased significantly by 10-15% during stimulation. During thermal stimulation, the maximum vesical pressure during micturition was unchanged, but the urethral relaxation duration was significantly prolonged. Applying local anesthesia to the stimulated skin area abolished the changes in voiding efficiency in response to thermal stimulation. These results suggest that the excitation of cutaneous thermoreceptive afferents modulates urethral function during urination, thereby improving voiding efficiency.

Machine learning-assisted fluoroscopy of bladder function in awake mice.

Understanding the lower urinary tract (LUT) and development of highly needed novel therapies to treat LUT disorders depends on accurate techniques to monitor LUT (dys)function in preclinical models. We recently developed videocystometry in rodents, which combines intravesical pressure measurements with X-ray-based fluoroscopy of the LUT, allowing the in vivo analysis of the process of urine storage and voiding with unprecedented detail. Videocystometry relies on the precise contrast-based determination of the bladder volume at high temporal resolution, which can readily be achieved in anesthetized or otherwise motion-restricted mice but not in awake and freely moving animals. To overcome this limitation, we developed a machine-learning method, in which we trained a neural network to automatically detect the bladder in fluoroscopic images, allowing the automatic analysis of bladder filling and voiding cycles based on large sets of time-lapse fluoroscopic images (>3 hr at 30 images/s) from behaving mice and in a noninvasive manner. With this approach, we found that urethane, an injectable anesthetic that is commonly used in preclinical urological research, has a profound, dose-dependent effect on urethral relaxation and voiding duration. Moreover, both in awake and in anesthetized mice, the bladder capacity was decreased ~fourfold when cystometry was performed acutely after surgical implantation of a suprapubic catheter. Our findings provide a paradigm for the noninvasive, in vivo monitoring of a hollow organ in behaving animals and pinpoint important limitations of the current gold standard techniques to study the LUT in mice.

Therapeutic effects of silodosin and urapidil on underactive bladder associated with diabetic cystopathy.

Pharmacological treatment options for underactive bladder (UAB) syndrome are limited. Urapidil is the only alpha1 -adrenoceptor (AR) antagonist that can be used for urinary disorders in women in some countries. However, no studies have directly verified the effects of alpha1 -AR antagonists on the female urethra and UAB-like dysfunctions. We investigated the effects of silodosin (alpha1A -AR antagonist) and urapidil (nonselective alpha1 -AR antagonist) on the voiding function in female rats with diabetes mellitus (DM). Changes in intraurethral pressure (IUP) induced by midodrine (alpha1 -AR agonist) and mean blood pressure (MBP) were continuously measured in normal female rats to verify the pharmacological profiles of the drugs. To establish a DM model, rats were administered streptozotocin (STZ; 50 mg/kg, intravenous). Eight weeks after STZ administration, drugs were subcutaneously delivered through an osmotic pump. Four weeks after drug administration, emptied bladder blood flow (BBF), intravesical pressure, and the micturition volume were measured. Both silodosin and urapidil inhibited the midodrine-induced increase in IUP and decreased MBP in a dose-dependent manner. Silodosin had a more substantial effect on the lower urinary tract than on MBP. Twelve weeks after STZ administration, DM rats exhibited UAB-like dysfunction (increased bladder capacity/bladder weight and residual volume and decreased bladder voided efficiency) and decreased BBF. Both drug treatments controlled this dysfunction. Alpha1 -AR antagonists induced dose-dependent urethral relaxation in female rats. These drugs ameliorated UAB-like dysfunction in STZ-induced DM rats. In addition, alpha1A -AR antagonists such as silodosin, which have limited effects on blood pressure, appear to be useful for treating UAB.

Fowler's Syndrome-The Cause of Urinary Retention in Young Women, Often Forgotten, but Significant and Challenging to Treat.

Urinary retention in young women is a relatively rare clinical problem and is often underdiagnosed. In particular, functional causes of urinary retention pose a diagnostic challenge. One of them is Fowler’s syndrome, which is associated with impaired urethral relaxation. Fowler’s syndrome is characterized by a large bladder capacity, reduced sensation, increased maximal urethral closure pressure, and detrusor underactivity. Several hypotheses have arisen to explain the cause of urethral relaxation disorders: hormonal changes characteristic of Polycystic Ovary Syndrome (PCOS), causing abnormal stabilization of the muscle membrane, primary failure of relaxation of the striated muscle of the urethra sphincter, and increased urethral afferent activity, inhibiting the bladder afferent signals from reaching the brain by potentiating a spinal mechanism of urinary continence. Currently, sacral neuromodulation is the only intervention that can restore an atypical voiding pattern in women with Fowler’s syndrome. The therapeutic effectiveness exceeds 70%, although the revision rate is relatively high, exceeding 50%. Well-designed, long-term prospective studies comparing sacral neuromodulation (SNM) with other therapies such as pelvic floor muscle physiotherapy are warranted to offer the best patient-tailored treatment.

Can detomidine replace medetomidine for pharmacological semen collection in domestic cats?

Among the different methods used for semen collection from domestic cats, the pharmacological collection by urethral catheterization becomes disruptive. Medetomidine is the elected α2-adrenoceptor agonist for that, but in several countries, it is not commercially available. This study aimed to evaluate the efficacy of detomidine compared to medetomidine in collecting semen by urethral catheterization in domestic cats. Urethral catheterization was performed on 13 mongrel cats using a disposable semi-rigid tomcat urinary catheter. Of the 19 semen collections performed with medetomidine induction, 94.7% were successful, while with detomidine induction, only 56.3% of 16 were successful. The values semen samples variables were as follows for volume - 10.56 ± 0.4 vs 8.88 ± 0.5 mL, motility - 171.67 ± 0.79 vs 49.77 ± 3.45%, vigor – 4.1 ± 0.03 vs 3.10 ± 0.1 and concentration - 3.24 ± 0.19 vs 2.15 ± 0.13 ×109 sperm/mL respectively for medetomidine and detomidine group. The failure in semen collections with detomidine was mainly due to azoospermic samples, poor urethral relaxation, insufficient volume, or contamination of urine. The sperm concentration was also lower in the detomidine group (P <0.05) when compared to medetomidine. However, when the volume of semen collected was compared, we found no statistical differences. Despite its low performance in collecting semen from cats, detomidine may be an alternative when medetomidine is not accessible.

PD24-05 OXIDATIVE STRESS INDUCES DETRUSOR UNDERACTIVITY AND URETHRAL DYSFUNCTION IN AGING RATS

You have accessJournal of UrologyBladder & Urethra: Anatomy, Physiology & Pharmacology II (PD24)1 Apr 2020PD24-05 OXIDATIVE STRESS INDUCES DETRUSOR UNDERACTIVITY AND URETHRAL DYSFUNCTION IN AGING RATS Asato Ohtsubo*, Takuma Oshiro, Ryu Kimura, Minoru Miyazato, Tomohiro Matsuo, Yasuyoshi Miyata, and Hideki Sakai Asato Ohtsubo*Asato Ohtsubo* More articles by this author , Takuma OshiroTakuma Oshiro More articles by this author , Ryu KimuraRyu Kimura More articles by this author , Minoru MiyazatoMinoru Miyazato More articles by this author , Tomohiro MatsuoTomohiro Matsuo More articles by this author , Yasuyoshi MiyataYasuyoshi Miyata More articles by this author , and Hideki SakaiHideki Sakai More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000881.05AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Age-related symptoms, such as urinary retention and a poor urinary stream have been attributed to detrusor underactivity. Our recent studies showed that by simultaneously recording intravesical pressure and urethral perfusion pressure (UPP), urethral relaxation during bladder contractions was diminished in aged rats. Nevertheless, L-arginine (a substrate of nitric oxide (NO) synthase) could not restore a diminished urethral relaxation. Thus, we hypothesized that age-associated oxidative stress would impair NO/soluble guanylyl cyclase (sGC) system which induces urethral relaxation. This hypothesis was tested by measuring an oxidative stress marker in the bladder and also by simultaneously recording intravesical pressure and UPP using BAY 41-2272 as a novel NO-independent stimulator of sGC. METHODS: Female Sprague Dawley rats (young rats aged 3 months and old rats aged 12–15 months) were used. 1) Oxidative stress marker staining for 8-hydroxy-2’-deoxyguanosine (8-OHdG) and malondialdehyde (MDA) in the bladder was performed in both groups. 2) Urethral activity was evaluated by simultaneously recording the intravesical pressure under isovolumetric conditions and UPP under urethane anesthesia. 3) BAY 41-2272 (1 mg/kg) was administered intravenously in both groups under isovolumetric conditions. RESULTS: 1) Oxidative marker staining revealed that 8-OHdG and MDA levels were higher in aging rats than those in young rats. 2) UPP nadir during urethral relaxation was significantly higher (38%) in aging rats than in young rats (p < 0.05). The mean amplitude of high frequency oscillation (HFO) of the urethral striated muscle was also significantly lower (49%) in aging rats than in normal rats (p < 0.05). 3) BAY 41-2272 increased urethral relaxation in young rats while restoring impaired urethral relaxation in aged rats. CONCLUSIONS: These results indicate that aging induces a dysfunction in urethral relaxation during voiding, as evidenced by a decrease in HFO activity of the urethral striated muscle as well as NO mediated urethral relaxation of the urethral smooth muscle. Because NO-independent stimulator could restore impaired urethral relaxation in the aged bladder, age-associated oxidative stress in the NO/sGC system might be correlated with an age associated impairment in bladder and urethral coordination. This novel study coupled with physiological and morphological changes in the aged bladder would contribute to elucidating the process of aging in bladders. Source of Funding: This work was supported by a Grant-in-Aid for Scientific Research (18K17752, 18K10714 and 18K10749 ). © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e533-e534 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Asato Ohtsubo* More articles by this author Takuma Oshiro More articles by this author Ryu Kimura More articles by this author Minoru Miyazato More articles by this author Tomohiro Matsuo More articles by this author Yasuyoshi Miyata More articles by this author Hideki Sakai More articles by this author Expand All Advertisement PDF downloadLoading ...

MP48-03 THE EFFECTS OF L-ARGININE ON URETHRAL FUNCTION IN STREPTOZOTOCIN-INDUCED AND INSULIN-TREATED DIABETIC RATS

You have accessJournal of UrologyUrodynamics/Lower Urinary Tract Dysfunction/Female Pelvic Medicine: Neurogenic Voiding Dysfunction (MP48)1 Apr 2020MP48-03 THE EFFECTS OF L-ARGININE ON URETHRAL FUNCTION IN STREPTOZOTOCIN-INDUCED AND INSULIN-TREATED DIABETIC RATS Nailong Cao, Baojun Gu*, Eduardo C Alexandre, Daisuke Gotoh, Masahiro Kurobe, Shinsuke Mizeguchi, and Naoki Yoshimura Nailong CaoNailong Cao More articles by this author , Baojun Gu*Baojun Gu* More articles by this author , Eduardo C AlexandreEduardo C Alexandre More articles by this author , Daisuke GotohDaisuke Gotoh More articles by this author , Masahiro KurobeMasahiro Kurobe More articles by this author , Shinsuke MizeguchiShinsuke Mizeguchi More articles by this author , and Naoki YoshimuraNaoki Yoshimura More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000903.03AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Normal lower urinary tract (LUT) function is dependent on the coordination between bladder and urethra, which is sensitive to a host of disorders, including diabetes mellitus (DM). Although STZ-induced diabetic rat model has been extensively used as a rodent model of the LUT dysfunction, it’s not a good physiological condition for diabetic patients. Our purpose is therefore to establish a physiological DM animal model with low dose insulin treatment, and examine the effects of L-arginine on urethral function in this animal model. METHODS: Thirty female Sprague-Dawley rats were divided into three groups. DM was initially induced by i. p. injection of STZ (65 mg/kg), and then received subcutaneous implantation of insulin under the mid dorsal skin. Isovolumetric cystometry and urethral perfusion pressure (UPP) measurements were performed under urethane anesthesia. L-arginine (100 mg/kg), was administered intravenously. Subsequently, N-nitro-L-arginine methyl ester hydrochloride (L-NAME) (50 mg/kg) was injected intravenously to inhibit NO synthase activity. The changes of urethral function among different groups in vitro were tested by organ bath. RESULTS: In vivo study, UPP nadir was only decreased by L-arginine both in Normal control (NC) and insulin-treated DM groups, but inhibited by L-NAME in all the groups (Fig.1). Furthermore, 5 out of 6 DM rats showed a similar detrusor-sphincter dyssynergia pattern after L-NAME (Fig.1). In vitro study, the ratio, namely urethral relaxation difference divided by relaxation before L-NAME administration, was significantly decreased in DM group (Fig.2). CONCLUSIONS: Low dose insulin can protest against diuresis-induced bladder overdistention and increase urethral relaxation, and this may be attributed to the protection of NO synthase and urethral NO sensitivity, but the longer effects should be further explored. Source of Funding: NIH (U54 DK112079 and R01 DK115476) © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e695-e695 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Nailong Cao More articles by this author Baojun Gu* More articles by this author Eduardo C Alexandre More articles by this author Daisuke Gotoh More articles by this author Masahiro Kurobe More articles by this author Shinsuke Mizeguchi More articles by this author Naoki Yoshimura More articles by this author Expand All Advertisement PDF downloadLoading ...

Urethral dysfunction and alterations of nitric oxide mechanisms in streptozotocin-induced diabetic rats with or without low-dose insulin treatment

AimsTo establish an animal model of diabetes mellitus (DM) with moderately elevated blood glucose levels, and to examine the nitric oxide (NO) mechanism controlling urethral function in streptozotocin (STZ)-induced DM rats. Main methodsFemale Sprague-Dawley rats were used. DM was induced by intraperitoneal injection of STZ (65 mg/kg) and some of them received subcutaneous implantation of a low-dose insulin pellet. Voiding behavior was evaluated in metabolic cages. Isovolumetric cystometry and urethral perfusion pressure (UPP) were then evaluated under urethane anesthesia, during which L-arginine (100 mg/kg) and N-nitro-L-arginine methyl ester hydrochloride (L-NAME) (50 mg/kg) were administered intravenously. In vitro urethral activity was also tested by organ bath muscle strip studies. Key findingsUPP changes and high-frequency oscillation (HFO) were significantly (P < 0.05) smaller in 8-weeks DM rats vs. normal control (NC) rats or insulin-treated DM rats, which showed reductions in urine overproduction and voided volume per micturition vs. untreated DM rats. UPP nadir was decreased by L-arginine in NC and insulin-treated DM groups, and decreased by L-NAME in all groups. Five of 6 untreated DM rats showed a detrusor-sphincter dyssynergia pattern after L-NAME. In in vitro studies, the relative ratio of L-NAME-induced reductions of urethral relaxation against pre-drug urethral relaxation was significantly smaller in DM vs. NC rats (P < 0.05). SignificanceLow-dose insulin-treated DM rats would be a useful model for studying natural progression of DM-induced lower urinary tract dysfunction. The impaired NO-mediated urethral relaxation mechanisms play an important role in DM-induced urethral dysfunction, which could contribute to DM-induced inefficient voiding.

Pathophysiological changes of the lower urinary tract behind voiding dysfunction in streptozotocin-induced long-term diabetic rats

We evaluated pathophysiological characteristics of the lower urinary tract dysfunction in a streptozotocin (STZ)-induced diabetic rat model. STZ (60 mg/kg) was injected intraperitoneally into male Wistar rats. In vitro bladder muscle strip experiments, in vivo cystometry, and simultaneous recordings of bladder pressure + urethral perfusion pressure (BP + UPP) with or without intravenous administration of L-arginine (300 mg/kg) or tadalafil (0.03 mg/kg) were performed at several time points. In vitro muscle strip experiments demonstrated that diabetic rats had significantly higher contractile responses to carbachol at 4–16 weeks, and a tendency for higher contractile responses to electrical field stimulation at 4–12 weeks, but this was reversed at 16 weeks. Diabetic rats had significant increases in voided volume, residual volume, bladder capacity, maximal voiding pressure, and amplitude and frequency of non-voiding contractions at 16 weeks. Tadalafil decreased the residual volume in diabetic rats. Diabetic rats had significantly higher UPP nadir and mean UPP during high-frequency oscillation at 16 weeks, which were reversed by tadalafil or L-arginine administration. The present results suggest that urethral relaxation failure, probably related to impairment of the NO/cGMP signalling pathway, rather than bladder contractile dysfunction may be a prominent cause for voiding dysfunction in STZ-induced chronic diabetic rats.

Urethral dysfunction and therapeutic effects of a PDE 5 inhibitor (tadalafil) in a rat model of detrusor underactivity induced by pelvic nerve crush injury.

The urethral dysfunction produced by a rat model of peripheral neurogenic detrusor underactivity (DU) using pelvic nerve crush (PNC) injury was characterized and then tested with the administration of tadalafil, a phosphodiesterase type 5 (PDE 5) inhibitor. Ten days after producing PNC rats, awake cystometrograms (CMGs) and isovolumetric cystometrograms with urethral perfusion pressure (IC-UPP) measurements were performed. Also, in control rats, IC-UPP was recorded before and after intravenous atropine administration to determine if the reduction of bladder contraction pressure affects urethral relaxation during voiding. Then, CMG and IC-UPP measurements in PNC rats were recorded after intravenous administration of tadalafil. Lastly, real-time polymerase chain reaction was used to measure transcript levels of neuronal nitric oxide synthases (nNOS), endothelial nitric oxide synthases, and PDE 5 in urethral specimens from PNC and control rats. PNC rats demonstrated the characteristics of DU in CMG. Also, PNC rats exhibited significant decreases in isovolumetric bladder contraction amplitudes and urethral relaxation. Atropine attenuated the amplitude of isovolumetric bladder contractions; however, atropine did not affect urethral relaxation in control rats. Tadalafil decreased postvoid residual and increased voiding efficiency without changing bladder contraction amplitude in PNC rats. Also, tadalafil improved the amplitude of urethral relaxation during bladder contraction in PNC rats. Urethral nNOS transcript levels were upregulated in PNC rats compared to control rats. PNC rats revealed both DU and impaired urethral relaxation. PDE 5 inhibition in PNC rats enhanced urethral relaxation during voiding, resulting in improved voiding efficiency. Thus, urethral dysfunction could be a potential target for the treatment of inefficient voiding associated with neurogenic DU.

Urethral dysfunction in a rat model of chemically induced prostatic inflammation: potential involvement of the MRP5 pump.

Prostate inflammation (PI) is a clinical condition associated with infection and/or inflammation of the prostate. It is a common disease frequently associated to lower urinary tract (LUT) symptoms. The urethra is an understudied structure in the LUT and plays a fundamental role in the urinary cycle. Here, we proposed to evaluate the effect of PI on the urethra tissue. Male Sprague-Dawley rats were used, and PI was induced by formalin injection into the ventral lobes of the prostate. The pelvic urethra at the prostatic level was harvested for histological analysis, contraction (electrical field stimulation and phenylephrine), and relaxation (sodium nitroprusside/MK-571) experiments. Various gene targets [cytochrome c oxidase subunit 2, transforming growth factor-β1, interleukin-1β, hypoxia-inducible factor-1α, α1A-adrenoceptor, inositol 1,4,5-trisphosphate receptor type 1, voltage-gated Ca2+ channel subunit-α1D, neuronal nitric oxide synthase, soluble guanylyl cyclase, phosphodiesterase 5A, protein kinase CGMP-dependent 1, and multidrug resistance-associated protein 5 (MRP5; ATP-binding cassette subfamily C member 5)] were quantified, and cGMP levels were measured. No histological changes were detected, and functional assays revealed decreased contraction and increased relaxation of urethras from the PI group. The addition of MK-571 to functional assays increased urethral relaxation. Genes associated with inflammation were upregulated in urethras from the PI group, such as cytochrome oxidase c subunit 2, transforming growth factor-β1, interleukin-1β, and hypoxia-inducible factor-1α. We also found increased expression of L-type Ca2+ channels and the neuronal nitric oxide synthase enzyme and decreased expression of the MRP5 pump. Finally, cGMP production was enhanced in urethral tissue of PI animals. The results indicate that PI is associated with proinflammatory gene expression in the urethra without histologically evident inflammation and that PI produces a dysfunctional urethra and MRP5 pump downregulation, which results in cGMP accumulation inside the cell. These findings would help to better understand LUT dysfunctions associated with PI and the role of MRP pumps in the control of LUT function.

Further characterization of a novel EP2 and EP3 receptor dual agonist, ONO-8055, on lower urinary tract function in normal and lumbar canal stenosis rats.

To further explore the effects of a novel EP2 and EP3 dual agonist, ONO-8055, on detrusor contractility, we investigated the responses of bladder strips from sham and lumbar canal stenosis (LCS) rats to this agonist, its effects on lower urinary tract function in normal rats, and mRNA expression of EP2 and EP3 receptors in the sham and LCS rats. The responses of bladder strips from sham and LCS rats to ONO-8055 were measured. The effects of ONO-8055 on LUT function of normal rats were investigated with awake cystometry and intraurethral perfusion pressure (Pura) measurements. The relative mRNA of bladder and urethral tissue of the sham and LCS rats was quantified using specific probes for EP1, EP2, EP3, and EP4 genes. Compared with the vehicle, the muscle tensions of both the sham and LCS rats were significantly increased after adding this agonist. On awake cystometry of normal rats, bladder capacity and Pura were decreased in the ONO-8055 groups, but a statistically significant difference in mean changes was demonstrated only between the vehicle group and the group receiving the highest dose. Compared with the sham rats, mRNA expressions of the four EP receptors in the lower urinary tract of the LCS rats did not show a statistically significant difference. This agonist did not augment bladder contractility or urethral relaxation in normal rats.

Mirabegron causes vesical and urethral relaxation in rats with spinal cord injury.

The effects of solifenacin and mirabegron on vesical and urethral function were compared in rats with or without spinal cord injury (SCI). Isovolumetric cystometry and urethral pressure recording were initially performed in intact rats. Then, the bladder neck was ligated under urethane anesthesia, after which a catheter was inserted through the bladder dome for isovolumetric cystometry and another catheter was inserted into the urethra to measure urethral pressure. Solifenacin (0.03-3 mg/kg) or mirabegron (0.03-3 mg/kg) was injected intravenously, and bladder and urethral activity were recorded. To create rats with SCI, the spinal cord was transected at the lower thoracic level under isoflurane anesthesia. After 2 weeks, a catheter was inserted through the bladder dome for single cystometry and bladder activity was recorded without anesthesia following intravenous injection of solifenacin or mirabegron. Isovolumetric cystometry revealed a larger decrease in maximum bladder contraction pressure after injection of solifenacin, whereas prolongation of the interval between bladder contractions was greater with mirabegron. In SCI rats, single cystometry showed that solifenacin and mirabegron both increased bladder volume at the first non-voiding bladder contraction and decreased the maximum bladder contraction pressure. Mirabegron also increased the voided volume and decreased the percentage residual volume without altering bladder capacity. Solifenacin and mirabegron both inhibited bladder contractility, and mirabegron possibly also induced urethral relaxation. Mirabegron may be suitable for patients with overactive bladder and residual urine.

Tadalafil, a phosphodiesterase type 5 inhibitor, restores urethra and detrusor function in the initial phase of diabetes in rats.

This study investigated the effects of tadalafil on the urethra and detrusor in the initial phase of diabetes in rats. Thirty-six female Sprague-Dawley rats were assigned to a non-diabetes (ND), diabetes (D), or tadalafil-treated diabetes (DT) group (n = 12 per group), with the DT group receiving oral tadalafil (2 mg/kg/d) for 7 days before the experiments. Seven weeks after diabetes induction (by a single intraperitoneal injection of streptozotocin), urethral and intravesical pressure were simultaneously recorded in vivo, whereas responses of detrusor strips to potassium chloride (30 mM), electrical field stimulation (EFS) and carbachol were measured in vitro. The intravesical pressure at which the urethra started to relax was significantly lower in the DT than D group (mean [± s.d.] 18.9 ± 2.9 vs 29.1 ± 6.6 cm H2 O; P < .05). In addition, the reduction in urethral pressure was significantly larger in the DT than D group (-10.9 ± 4.0 vs -4.0 ± 2.9 cm H2 O; P < .05). Detrusor stimulation revealed that the mean contractile responses to EFS and carbachol were significantly lower in the ND and DT groups than in the D group (120.7 ± 26.5% and 130.8 ± 15.8% vs 200.1 ± 47.9% of the 30 mM KCl-induced contraction, respectively, in response to 50 Hz EFS [P < .05]; 211.1 ± 35.4% and 208.4 ± 25.3% vs 425.7 ± 125.0% of the 30 mM KCl-induced contraction, respectively, in response to 10-3 M carbachol [P < .05]). Tadalafil restored urethral relaxation function and detrusor responses to EFS and carbachol during the initial phase of diabetes.

Effect of ASP6432, a Novel Type 1 Lysophosphatidic Acid Receptor Antagonist, on Urethral Function and Prostate Cell Proliferation.

Current pharmacotherapies for lower urinary tract symptoms associated with benign prostate hyperplasia (LUTS/BPH) are in need of improvement. Lysophosphatidic acid (LPA) is a phospholipid with various biologic functions. However, its exact role in the lower urinary tract and its target receptor subtype have not been fully elucidated. We investigated the role of LPA and the type 1 LPA receptor (LPA1) in urethral/prostatic contractile function and prostate cell proliferation by pharmacologically characterizing ASP6432 (potassium 1-(2-{[3,5-dimethoxy-4-methyl-N-(3-phenylpropyl)benzamido]methyl}-1,3-thiazole-4-carbonyl)-3-ethyl-2,2-dioxo-2λ6-diazathian-1-ide), a novel LPA1 antagonist. ASP6432 exhibited potent and selective antagonistic activity against LPA1 in cells expressing LPA receptor subtypes. In isolated rat tissue strips and anesthetized rats, ASP6432 concentration-/dose-dependently inhibited LPA-induced urethra and prostate contractions. In addition, in anesthetized rats, ASP6432 maximally decreased the urethral perfusion pressure (UPP) in the absence of exogenous LPA stimulation by 43% from baseline, whereas tamsulosin, an α1-adrenoceptor antagonist, reduced UPP by 22%. Further, in human prostate stromal cells, ASP6432 significantly and concentration-dependently suppressed LPA-induced bromodeoxyuridine incorporation. These results demonstrate a pivotal role for LPA and LPA1 in the regulation of urethral tonus and prostate cell proliferation. The potent urethral relaxation and inhibition of prostatic stromal cell growth indicate the potential of ASP6432 as a novel therapeutic agent for LUTS/BPH.

Grape seed proanthocyanidin extract alleviates urethral dysfunction in diabetic rats through modulating the NO-cGMP pathway.

Oxidative stress is closely associated with the onset of diabetes mellitus (DM). Diabetic urethropathy is one of the most common complications of DM, but few studies have been conducted to investigate the role of oxidative stress in diabetic urethropathy. Grape seed proanthocyanidin extract (GSPE) has been previously reported to reduce oxidative injury. The present study aimed to investigate the role of oxidative stress and the protective effects of GSPE on urethral dysfunction using a streptozotocin-induced DM rat model. Female Wistar rats were divided into a control group (n=36), a DM group (n=36) and a DM + GSPE group (n=36). Urodynamic testing was performed using a PowerLab data acquisition device. The expression of neuronal nitric oxide synthase (nNOS), 3-nitrotyrosine and nuclear factor erythroid 2-related factor 2 (Nrf2) was determined using western blot analysis. The expression of 3-nitrotyrosine was also determined using immunohistochemistry. Nitric oxide (NO), cyclic guanosine monophosphate (cGMP), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) were measured using commercial ELISA kits. A significant increase was observed in the intravesical pressure thresholds for inducing urethral relaxation and the urethral perfusion pressure nadir in DM rats compared with the control group. GSPE was observed to reverse the increase of these parameters compared with the DM group. In addition, GSPE could reverse the downregulation of nNOS, NO and cGMP expression, and the decreased activities of antioxidant enzymes (SOD and GSH-Px). GSPE reversed the upregulation of 3-nitrotyrosine and MDA in DM rats. GSPE also activated Nrf2, which is a key antioxidative transcription factor. The findings of the present study demonstrated that GSPE protects urethra function in DM rats through modulating the NO-cGMP signaling pathway. The protective roles of GSPE may be associated with activation of the Nrf2 defense pathway.

OAB without an overactive bladder in the acute prostaglandin E2 rat model.

Intravesical prostaglandin E2 (PGE2) was previously used to induce overactive bladder (OAB) symptoms, as it reduces bladder capacity in rats and causes a "strong urgency sensation" in healthy women. However, the mechanism by which this occurs is unclear. To clarify how PGE2 reduces bladder capacity, 100 µM PGE2 was administered intravesically during open, single-fill cystometry with simultaneous measurement of sphincter EMG in the urethane-anesthetized female Wistar rat. PGE2 was also applied to the urethra or bladder selectively by use of a ligature at the bladder neck before (urethra) or during (bladder) closed-outlet, single-fill cystometry. Additional tests of urethral perfusion with PGE2 were made. PGE2 decreased bladder capacity, increased voiding efficiency, and increased sphincter EMG during open cystometry compared with saline controls. The number of nonvoiding contractions did not change with PGE2; however, bladder compliance decreased. During closed-outlet cystometry, PGE2 applied only to the bladder or the urethra did not decrease bladder capacity. Urethral infusion of PGE2 decreased urethral perfusion pressure. Taken together, these results suggest that intravesical PGE2 may decrease bladder capacity by targeting afferents in the proximal urethra. This may occur through urethral relaxation and decreased bladder compliance, both of which may increase activation of proximal urethra afferents from distension of the proximal urethra. This hypothesis stands in contrast to many hypotheses of urgency that focus on bladder dysfunction as the primary cause of OAB symptoms. Targeting the urethra, particularly urethral smooth muscle, may be a promising avenue for the design of drugs and devices to treat OAB.