Elucidation of the pathophysiology of interstitial cystitis/bladder pain syndrome via experimental autoimmune cystitis rat model.

Although the cause of interstitial cystitis/painful bladder syndrome (IC/PBS) remains unknown, autoimmune involvement has been strongly suggested to be a contributing factor. To elucidate the pathophysiology of IC/PBS, we characterized the experimental autoimmune cystitis (EAC) in rats. Adult female Sprague-Dawley rats were divided into the EAC and control groups. The EAC rats were generated by administrating a homogenate of donor rat bladder tissue as a bladder antigen. The characteristics of the two groups were determined by evaluating pain behavior and conducting cystometry, histopathology, and molecular analyses. The EAC rats showed: [1] a decreased paw withdrawal threshold, [2] a reduced intercontraction interval on cystometry, [3] the irregular surfaces of the umbrella cells of epithelium throughout the bladder wall, [4] accumulation of stress granules in the bladder and vascular endothelium, [5] the increased expression of genes related to inflammation and ischemia at the mRNA and protein levels, [6] a significantly increased paw withdrawal threshold with pain treatment, and [7] the induction of glomerulation of the bladder wall, epithelium denudation, and lymphocyte infiltration in the interstitium by bladder distension. These results suggest that the EAC rats showed pain and frequent urination with the overexpression of inflammatory chemokines, reflecting clinical IC/BPS, and the bladder epithelium and vascular endothelium may be the primary sites of IC/BPS, and bladder injury such as bladder distension can cause progression from BPS to IC with Hunner lesions.

Choreito, a Kampo medicine attenuates detrusor overactivity and bladder pain symptoms in rat tranilast-induced interstitial cystitis/bladder pain syndrome-like model.

AIMS: Interstitial cystitis/bladder pain syndrome (IC/BPS) is a chronic inflammatory condition of the bladder. However, there are only a few medicines that are of pharmaceutical grade and reliably effective for IC/BPS symptoms. Choreito (CRT) is a pharmaceutical-grade Kampo medicine and has been widely prescribed for patients of lower urinary tract symptoms (LUTS) and BPS in Japan. In this study, we exploratory investigated the effects of CRT on the IC/BPS-like symptoms induced by tranilast. METHODS: The rat IC/BPS-like model was induced by feeding administration with 0.4% tranilast. The rats were divided into the three following treatment groups: normal diet (Normal), tranilast treatment (Control), and the groups of 1% CRT (CRT) treatment for IC/BPS-like model. After 4 weeks, continuous cystmetry, locomotor, and vascular permeability was assessed. Furthermore, the cytokine levels in bladder were analyzed by the Bio-Plex suspension array system and plasma monoamine were measured. RESULTS: Control group exhibited 14.3% decrease of locomotor activity in the dark period, and which were 20.3% increase by 1%CRT treatment. The voiding interval was shorter in control than in other groups. 1%CRT suppressed the shortening of voiding interval. Evans blue leakage of bladder wall observed 44.8% higher in control group than in the normal group. The leakage of 1%CRT group was 33.3% less than in the control group. The cytokine level of IFNγ and VEGF were elevated in the control, and CRT treatment suppressed the elevation of IFNγ in the bladder. Plasma noradrenaline was significantly reduced by CRT treatment compared normal group. CONCLUSION: These results suggest that CRT can be an effective therapeutic agent for the treatment of IC/BPS-like symptoms.

CNOT1 regulates circadian behaviour through Per2 mRNA decay in a deadenylation-dependent manner.

Circadian clocks are an endogenous internal timekeeping mechanism that drives the rhythmic expression of genes, controlling the 24 h oscillatory pattern in behaviour and physiology. It has been recently shown that post-transcriptional mechanisms are essential for controlling rhythmic gene expression. Controlling the stability of mRNA through poly(A) tail length modulation is one such mechanism. In this study, we show that Cnot1, encoding the scaffold protein of the CCR4-NOT deadenylase complex, is highly expressed in the suprachiasmatic nucleus, the master timekeeper. CNOT1 deficiency in mice results in circadian period lengthening and alterations in the mRNA and protein expression patterns of various clock genes, mainly Per2. Per2 mRNA exhibited a longer poly(A) tail and increased mRNA stability in Cnot1+/- mice. CNOT1 is recruited to Per2 mRNA through BRF1 (ZFP36L1), which itself oscillates in antiphase with Per2 mRNA. Upon Brf1 knockdown, Per2 mRNA is stabilized leading to increased PER2 expression levels. This suggests that CNOT1 plays a role in tuning and regulating the mammalian circadian clock.

CNOT7 Outcompetes Its Paralog CNOT8 for Integration into The CCR4-NOT Complex.

The CCR4-NOT deadenylase complex is a major post-transcriptional regulator of eukaryotic gene expression. CNOT7 and CNOT8 are both vertebrate homologs of the yeast CCR4-NOT catalytic subunit Caf1. They are highly similar and are sometimes considered redundant, but Cnot7 and Cnot8 knockout mice exhibit different phenotypes, implying distinct physiological functions. In this study, we reveal a non-reciprocal effect of CNOT7 on CNOT8, in which CNOT8 protein is increased in the depletion of CNOT7 without corresponding changes in mRNA levels whereas CNOT7 is not affected by the loss of CNOT8. Cnot8 mRNA may be bound by the CCR4-NOT complex, suggesting that CCR4-NOT might directly regulate CNOT8 expression. Cnot8 mRNA is relatively unstable, but Cnot7 knockdown did not stabilize Cnot8 mRNA, nor did it increase translation. CNOT8 protein was also less stable than CNOT7. CNOT7 showed greater affinity than CNOT8 for the CCR4-NOT scaffold protein CNOT1 and was able to block CNOT8 from binding to CNOT1. Depletion of CNOT7 increased CNOT8 incorporation into the CCR4-NOT complex and stabilized CNOT8. These data suggest that CNOT7 is the dominant paralog in CCR4-NOT and that CNOT7 and CNOT8 protein stability is regulated in distinct ways.

Regulation of CCR4-NOT complex deadenylase activity and cellular responses by MK2-dependent phosphorylation of CNOT2.

CCR4-NOT complex-mediated mRNA deadenylation serves critical functions in multiple biological processes, yet how this activity is regulated is not fully understood. Here, we show that osmotic stress induces MAPKAPK-2 (MK2)-mediated phosphorylation of CNOT2. Programmed cell death is greatly enhanced by osmotic stress in CNOT2-depleted cells, indicating that CNOT2 is responsible for stress resistance of cells. Although wild-type (WT) and non-phosphorylatable CNOT2 mutants reverse this sensitivity, a phosphomimetic form of CNOT2, in which serine at the phosphorylation site is replaced with glutamate, does not have this function. We also show that mRNAs have elongated poly(A) tails in CNOT2-depleted cells and that introduction of CNOT2 WT or a non-phosphorylatable mutant, but not phosphomimetic CNOT2, renders their poly(A) tail lengths comparable to those in control HeLa cells. Consistent with this, the CCR4-NOT complex containing phosphomimetic CNOT2 exhibits less deadenylase activity than that containing CNOT2 WT. These data suggest that CCR4-NOT complex deadenylase activity is regulated by post-translational modification, yielding dynamic control of mRNA deadenylation.

Phenotypic Characterization of the Endocannabinoid-Degrading Enzyme Alpha/Beta-Hydrolase Domain 6 Knockout Rat.

Introduction: Alpha/beta-hydrolase domain 6 (ABHD6) is an enzyme that hydrolyzes 2-arachidonoylglycerol, a high-efficiency endogenous cannabinoid. Although the endocannabinoid system has been suggested to be involved in regulation of bladder function, the roles of ABHD6 in the control of micturition remain unknown. To elucidate the physiological and pathological roles of ABHD6 in vivo, we examined phenotypes of ABHD6 knockout rats (Abhd6-/-) generated by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated proteins system. Materials and Methods: Age-matched knockout and wild-type (WT) rats of both sexes were used. Results: Expression of ABHD6, assessed by quantitative real-time polymerase chain reaction and Western blot analysis, was clearly diminished in Abhd6-/- rats compared with WT rats. Mutant rats had a normal appearance, and the body weight and food consumption were similar to those of WT rats. The interval between bladder contractions assessed by continuous cystometry was significantly shorter in ABHD6 knockout rats than in WT rats when the bladder was stimulated with acetic acid. Mechanical paw withdrawal thresholds measured by von Frey testing were significantly lowered in the knockout rats than in WT rats. The plasma levels of prostaglandin E2 (PGE2) and the stable metabolite of PGE2 in Abhd6-/- rats were twice as high as that in WT rats. Conclusions: Deletion of the ABHD6 gene in rats causes more frequent urination in the stimulated bladder and hyperalgesia to non-noxious mechanical stimuli along with increased plasma PGE2.

Dietary glycine improves urine storage symptoms in urology outpatients.

OBJECTIVES: Spinal glycinergic mechanisms inhibit the micturition reflex, and administration of glycine inhibits bladder activity in rats. Therefore, we examined whether dietary glycine would improve storage symptoms in urological outpatients. METHODS: We enrolled 20 participants (16 men and four women) with an overactive bladder symptom score (OABSS) ≥ 3. All participants took 3 g of glucose (placebo) twice a day for the first four weeks, then 3 g of glycine twice a day for the next four weeks. We evaluated blood pressure, international prostate symptom score (IPSS), nocturia quality of life (N-QOL) score, OABSS, frequency of urination, sleep latency, time to first nighttime void, bladder pain, global self-assessment (GSA) evaluated urinary symptom improvement, and adverse events. RESULTS: Glucose administered as a placebo improved urinary frequency, urine force on the IPSS, and five of the 13 items on the N-QOL. However, compared to the results before and after glucose administration, glycine treatment decreased the number of nocturnal voids, urgency, and total score for urine storage items on the IPSS. It also reduced blood pressure and improved IPSS-QOL. For the OABSS, improvements with glycine were noted in the number of nocturnal urinations, urinary urgency, urge incontinence, and total score. For the N-QOL, eight of 13 items, and the total score, improved. The actual number of nighttime urinations, sleep latency, latency to first nighttime urination, bladder pain, and GSA also improved. There were no adverse events. CONCLUSIONS: Glycine might improve urine storage symptoms, cardiovascular function, pain, and sleep.

Deletion of the lysyl oxidase-like 1 gene induces impaired elastin fiber synthesis and inefficient urethral closure in rats.

We investigated the bladder and urethral function in a rat model lacking the protein lysyl oxidase-like 1 (Loxl1). Female nulliparous rats of Loxl1-/- or age-matched wild type (WT) rats had leak-point pressure testing, cystometry, histopathological analyses of lower urinary tract, and contractile response of isolated detrusor strips to carbachol and electric field stimulation. The Loxl1-/- rats showed increased looseness and redundancy of the skin, the decreased intercontraction interval and voided volume in cystometry, the lower leak-point pressure, thinner elastic fibers of the mesentery, bladder, urethra and vagina, and smaller contractile response of detrusor strips to carbachol when compared to the WT rats. Thus, the insufficient hydrostatic mechanism of urethra via submucosal impaired elastin synthesis might reduce the resting urethral closure pressure and the diminished cholinergic contractile response of detrusor smooth muscle might be involved in bladder activity in the Loxl1-/- rats.

Vanilla scent reduces frequency of urination in urethane-anesthetized rats.

OBJECTIVES: The scent of vanilla has a relaxing effect and is used to treat sleep disorders. Sleep disorders can both cause and be caused by nocturia. Therefore, we examined whether vanilla inhalation would reduce the frequency of urination in rats under light urethane anesthesia. METHODS: Twenty-four rats were anesthetized with 0.6 g/kg urethane subcutaneously (half the usual dose) to induce a sleep-like state. In 12 rats, continuous cystometry was performed via a transurethral catheter before, during and after inhalation of vanilla (n = 7) or the citrus fruit shiikuwasa (n = 5) for 60 minutes. The remaining 12 rats did not undergo cystometry but underwent vanilla inhalation treatment for 60 minutes (n = 6), or no inhalation treatment (n = 6); blood was then collected from these two groups and serum monoamine levels were compared. RESULTS: Intervals between bladder contractions were significantly longer after vanilla inhalation than before. However, baseline bladder pressure, maximum bladder contraction pressure, and residual volume remained unchanged. During shiikuwasa inhalation, the body movement of each rat increased but cystometric parameters did not change. Serum concentrations of adrenaline, noradrenaline and dopamine, but not serotonin, were significantly lower in rats that had inhaled vanilla than in those that had not. CONCLUSIONS: Vanilla scent decreased serum catecholamine levels and urination frequency in rats under light urethane anesthesia. These results suggest that the scent of vanilla may reduce nocturia.

[ASSOCIATION OF BREATH HYDROGEN CONCENTRATION WITH ORAL INTAKE AND URINARY DISEASES].

(Purpose) Ingestion of hydrogen is said to prevent oxidation in the body, but hydrogen is produced by intestinal bacterial flora and excreted in the exhaled breath. We investigated how breath hydrogen concentrations change with the diurnal cycle and under various conditions, including after consuming food or drink, and in people with urological disease. (Subjects and methods) Participants were healthy volunteers (40 men, 45 women; 30-83 years old) and urological outpatients (40 men with benign prostatic hyperplasia, 30 women with overactive bladder; 60 years or older). Breath hydrogen levels were measured before and after eating and drinking in three volunteers, and its diurnal variation was examined in one. The relationship between breath hydrogen and age or urological disease status was also analyzed by gender. Additional measurements were taken in the person with the highest breath hydrogen concentration and the person with the lowest; in these two people, breath hydrogen was measured at the same time for 10 or more days to determine the fluctuation range. (Results) Breath hydrogen concentration increased temporarily after ingestion of tap water, hydrogen water or food. It also increased with food intake and in cases of flatulence with intestinal gas accumulation, but decreased after defecation. In the person with the highest breath hydrogen, concentrations were 11.2-188.6 ppm, whereas in the person with the lowest, they were 0.4-2.3 ppm. Breath hydrogen increased significantly with age in healthy female volunteers. There was no association between breath hydrogen and benign prostatic hyperplasia, overactive bladder or constipation. (Conclusion) Breath hydrogen concentration increases with eating, drinking and aging, and is not associated with benign prostatic hyperplasia, overactive bladder or constipation. Breath hydrogen concentration varies widely between individuals, which may be due to differences in intestinal flora.

The CCR4-NOT deadenylase complex safeguards thymic positive selection by down-regulating aberrant pro-apoptotic gene expression.

A repertoire of T cells with diverse antigen receptors is selected in the thymus. However, detailed mechanisms underlying this thymic positive selection are not clear. Here we show that the CCR4-NOT complex limits expression of specific genes through deadenylation of mRNA poly(A) tails, enabling positive selection. Specifically, the CCR4-NOT complex is up-regulated in thymocytes before initiation of positive selection, where in turn, it inhibits up-regulation of pro-apoptotic Bbc3 and Dab2ip. Elimination of the CCR4-NOT complex permits up-regulation of Bbc3 during a later stage of positive selection, inducing thymocyte apoptosis. In addition, CCR4-NOT elimination up-regulates Dab2ip at an early stage of positive selection. Thus, CCR4-NOT might control thymocyte survival during two-distinct stages of positive selection by suppressing expression levels of pro-apoptotic molecules. Taken together, we propose a link between CCR4-NOT-mediated mRNA decay and T cell selection in the thymus.

Regulation of Fetal Genes by Transitions among RNA-Binding Proteins during Liver Development.

Transcripts of alpha-fetoprotein (Afp), H19, and insulin-like growth factor 2 (Igf2) genes are highly expressed in mouse fetal liver, but decrease drastically during maturation. While transcriptional regulation of these genes has been well studied, the post-transcriptional regulation of their developmental decrease is poorly understood. Here, we show that shortening of poly(A) tails and subsequent RNA decay are largely responsible for the postnatal decrease of Afp, H19, and Igf2 transcripts in mouse liver. IGF2 mRNA binding protein 1 (IMP1), which regulates stability and translation efficiency of target mRNAs, binds to these fetal liver transcripts. When IMP1 is exogenously expressed in mouse adult liver, fetal liver transcripts show higher expression and possess longer poly(A) tails, suggesting that IMP1 stabilizes them. IMP1 declines concomitantly with fetal liver transcripts as liver matures. Instead, RNA-binding proteins (RBPs) that promote RNA decay, such as cold shock domain containing protein E1 (CSDE1), K-homology domain splicing regulatory protein (KSRP), and CUG-BP1 and ETR3-like factors 1 (CELF1), bind to 3' regions of fetal liver transcripts. These data suggest that transitions among RBPs associated with fetal liver transcripts shift regulation from stabilization to decay, leading to a postnatal decrease in those fetal transcripts.

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.

Insufficient liver maturation affects murine early postnatal hair cycle.

Abnormal hair loss results from a variety of factors, such as metabolic dysfunctions, immunodeficiency, and environmental stressors. Here, we report that mutant mice having defects in liver function, develop alopecia. We have shown previously that in mice lacking a Cnot3 gene, which encodes an essential component of the CCR4-NOT deadenylase complex in liver (Cnot3-LKO mice), the liver does not mature properly, resulting in various pathologies such as hepatitis, hepatic necrosis, and anemia. Unexpectedly, Cnot3-LKO mice start to lose hair around postnatal day 17 (P17). The region of hair loss expands all across their backs and symptoms persist until around P28-30. Afterward, hair re-grows, and Cnot3-LKO mice show complete hair recovery by P40. The phenotype is dependent on mouse genotype, indicating that hair follicle morphogenesis and cycling are influenced by abnormal liver development. By performing histological, quantitative PCR, and immunoblot analyses, we detected sebaceous gland (SG) hypertrophy accompanied by an increase of peroxisome proliferator-activated receptor γ (PPARγ). Collectively, these findings suggest that paracrine signaling related to liver function influences hair growth, at least in part, by altering lipid metabolism.

Effects of combined treatment with fesoterodine and mirabegron in a pelvic congestion rat model: Results from in vitro and in vivo functional studies.

OBJECTIVES: To examine the effect of combining a nonselective muscarinic receptor antagonist, 5-hydroxymethyl tolterodine (an active metabolite of fesoterodine), with a ß3 adrenoceptor agonist, mirabegron, in a rat model of pelvic congestion. METHODS: The rat pelvic congestion model used female Sprague-Dawley rats with their bilateral common iliac and uterine veins ligated. Expressions of M2 and M3 receptor subtypes in the urothelium and detrusor were detected by real-time polymerase chain reaction assays. The effects of both drugs were investigated on isolated bladder strips contracted by electrical field stimulation. in vivo single cystometry was used to assess the effects of 5-hydroxymethyl tolterodine and mirabegron independently or in combination on bladder capacity, micturition pressure, and threshold pressure. RESULTS: Pelvic congestion rats showed decreased bladder capacity compared with controls, but micturition pressure and threshold pressure were unchanged. Pelvic congestion model rats also demonstrated an approximately two-fold increase in expression of both M2 and M3 receptor subtypes in the urothelium. Additive relaxant effects of 5-hydroxymethyl tolterodine and mirabegron were observed in vitro in the electrical field stimulation-induced contractions of bladder strips from pelvic congestion rats. In vivo, bladder capacity was increased significantly by a combination of 5-hydroxymethyl tolterodine and mirabegron, with the combined effect exceeding the sum of the effects of monotherapies. Micturition pressure and threshold pressure did not significantly differ between groups. CONCLUSIONS: The combination of 5-hydroxymethyl tolterodine with mirabegron suggests the potential of synergistic effects in a rat pelvic congestion model.

Spinal glycinergic and gamma-aminobutyric acid-ergic neurons inhibit the micturition reflex after electrical stimulation of the perineum in rats with pelvic venous congestion.

OBJECTIVES: To examine whether electrical stimulation of the perineum inhibited urinary frequency in rats with pelvic venous congestion, and whether electrical stimulation influences spinal glycinergic/gamma-aminobutyric acid-ergic neurons. METHODS: Bilateral common iliac veins and bilateral uterine veins were ligated to create pelvic venous congestion rats. At 4 weeks after ligation, cystometry was carried out before and after electrical stimulation with/without intrathecal injection of strychnine (a glycine receptor antagonist) and/or bicuculline (a gamma-aminobutyric acid type A receptor antagonist). In addition, measurement of amino acid levels in the lumbosacral cord was carried out with/without electrical stimulation, and cystometry was carried out after oral administration of glycine. RESULTS: Continuous cystometry showed that the interval between bladder contractions was shorter in pelvic venous congestion rats than in sham rats. Electrical stimulation did not change cystometric parameters in sham rats, but the interval between bladder contractions was increased by electrical stimulation in pelvic venous congestion rats. Electrical stimulation increased the levels of glutamic acid, glycine, gamma-aminobutyric acid, and taurine in the lumbosacral cord of pelvic venous congestion rats. Intrathecal strychnine abolished the effects of electrical stimulation in pelvic venous congestion rats, and intrathecal administration of both strychnine and bicuculline shortened the interval between bladder contractions more than before electrical stimulation. Oral administration of glycine (3%) to pelvic venous congestion rats increased bladder capacity. CONCLUSIONS: Electrical stimulation of the perineum inhibits urinary frequency mainly through activation of spinal glycinergic neurons, and partly through activation of gamma-aminobutyric acid-ergic neurons in a rat model of pelvic venous congestion.

Mechanisms underlying the effects of propiverine on bladder activity in rats with pelvic venous congestion and urinary frequency.

We investigated the mechanisms by which propiverine hydrochloride influenced bladder activity in rats with pelvic venous congestion (PC) and urinary frequency. To create PC rats, female rats were anesthetized with isoflurane and the bilateral common iliac veins and bilateral uterine veins were ligated. At 4 weeks after ligation, we assessed voiding behaviour, locomotor activity, and urinary 8-hydroxydeoxyguanosine (8-OHdG) and nitric oxide metabolites (NOx). We also performed cystometry and measured mRNAs for nitric oxide synthase (NOS) and several receptors in the bladder wall. PC rats showed a decrease in locomotor activity and an increased frequency of urination. There was a decrease in endothelial NOS (eNOS), M3, and TRPV1 mRNA expression in the bladder wall, as well as an increase in inducible NOS (iNOS) mRNA. Administration of propiverine to PC rats increased locomotor activity to the level in sham rats, improved bladder function, decreased urinary 8-OHdG excretion, and increased urinary NOx excretion. In addition, propiverine increased neuronal NOS (nNOS) mRNA expression, and decreased expression of iNOS, M3 and TRPV1 mRNA in the bladder wall. Therefore, propiverine not only improved bladder dysfunction through its previously reported actions (anti-muscarinic effect, Ca antagonist effect, and inhibition of noradrenaline re-uptake), but also by reducing inflammation.

Evaluation of a rat model of functional urinary bladder outlet obstruction produced by chronic inhibition of nitric oxide synthase.

AIMS: Ligation of the urethra to create partial bladder outlet obstruction has widely been used as an animal model of bladder obstruction, although obstructive bladder dysfunction may be due to both mechanical and functional obstruction. Previous studies in rodents have demonstrated that long-term nitric oxide (NO) deficiency can lead to detrusor overactivity, and lack of NO may thus cause impairment of bladder outlet relaxation. The aim of this study was to define the characteristics of bladder and urethral dysfunction induced by chronic NO deficiency through both in vivo and in vitro investigations. MAIN METHODS: Rats were divided into two groups, and one group received an NO synthase inhibitor (Nω-nitro-L-arginine methyl ester hydrochloride: L-NAME) in the drinking water for 4 weeks. Bladder and urethral function were evaluated by continuous cystometry and isovolumetric cystometry. In vitro functional studies of detrusor strips and measurement of the mRNA and protein expression of an ischemic marker and a gap junction protein were also performed in separate rats. KEY FINDINGS: L-NAME administration raised blood pressure and decreased plasma nitrite/nitrate level compared to the control group. L-NAME treatment increased the frequency of bladder contractions and the residual volume, and elevated urethral pressure and bladder contraction pressure. In addition, carbachol-induced contraction was reduced in isolated detrusor strips from the L-NAME group, and bladder expression of HIF-1 and connexin 43 showed upregulation. SIGNIFICANCE: These findings suggest that chronic administration of L-NAME to rats induces bladder hyperactivity with residual urine, and may provide a useful model of functional bladder obstruction.

Emotional Stress Facilitates Micturition Reflex: Possible Inhibition by an α1-Adrenoceptor Blocker in the Conscious and Anesthetized State.

PURPOSE: To test the hypothesis that naftopidil prolongs intercontraction intervals in rats undergoing chronic stress as observed in previous animal models, voiding behavior and bladder function were measured and analyzed. METHODS: Female Sprague-Dawley rats weighing 200-230 g were exposed to repeated variate stress (RVS) for 1 week, chronic variable mild stress for 2 weeks, or simple mild stress for 1 week. Voiding behavior was assessed in metabolic cages. Voiding frequency and urine output were measured, and changes of these values were compared for the different types of stress. Micturition reflex was analyzed using unconscious cystometry. Naftopidil was administered orally at 30 mg/kg/day for 2 weeks. RESULTS: Unexpectedly, no stress-exposed rats exhibited increased micturition frequency compared to the normal nonstressed control. However, intercontraction intervals were shortened with each type of stress in the unconscious condition, especially by RVS (P<0.01). Naftopidil prolonged the shortened intervals. CONCLUSION: Although voiding behavior appears approximately normal in rats chronically exposed to emotional stress, internal bladder function can be affected. With anesthesia, micturition intervals were moderately shortened by emotional stress and clearly improved by naftopidil. Therefore, naftopidil appears to act at the spinal level at least.

Tadalafil improves bladder dysfunction and object recognition in rats with pelvic venous congestion.

OBJECTIVES: To examine the effects of tadalafil on bladder function and object recognition ability in rats with alterations in urinary frequency and locomotor activity as a result of pelvic venous congestion. METHODS: A total of 48 female rats were divided into three groups (sham, pelvic venous congestion and pelvic venous congestion/tadalafil groups). In the pelvic venous congestion and pelvic venous congestion/tadalafil groups, the bilateral common iliac veins and uterine veins were ligated under anesthesia. Rats in the pelvic venous congestion/tadalafil group received a diet containing tadalafil, and the other rats were fed a normal diet. After 4 weeks, rats underwent analysis of voiding behavior, locomotor activity, a novel object recognition test, continuous cystometry, measurement of plasma monoamines, and measurement of plasma and urinary nitric oxide metabolites. Expression of nitric oxide synthase messenger ribonucleic acid in the bladder wall was also assessed, along with histological examination of the bladder. RESULTS: Rats with pelvic venous congestion showed a higher urinary frequency, lower locomotor activity, and lower plasma and urinary nitric oxide levels than sham rats. The bladder wall endothelial nitric oxide synthase messenger ribonucleic acid level was low and object recognition was impaired. Pelvic venous congestion/tadalafil rats showed improvement in locomotor activity, bladder function and object recognition compared with pelvic venous congestion rats, as well as elevation of plasma and urinary nitric oxide, plasma monoamines, and bladder neuronal nitric oxide synthase messenger ribonucleic acid expression. Bladder wall vascularity was greater in pelvic venous congestion/tadalafil rats compared with sham rats. CONCLUSIONS: In rats with pelvic venous congestion, tadalafil might improve bladder function and the general condition by increasing blood flow to the bladder and brain, and by increasing dopamine levels.