Micturition In Rats Research Articles

Urinary and ejaculatory dysfunction induced by denervation of specific striated muscles anatomically related to the urethra in male rats

The purpose of the present study was to determine the contribution of the external urethral sphincter (EUS), the ischiocavernous (IC), or the bulbospongiosus (BS) on the control of micturition, copulatory behavior and semen expulsion in male rats. We hypothesized that the EUS contributes to maintain urinary continence, while all three muscles participate in expulsive urethral functions. In Experiment 1, it was analyzed the effects of bilateral denervation of IC, BS or EUS, or sham surgery, on voiding behavior and urinary parameters measured before surgery and 2 and 10 days post-surgery. In Experiment 2, copulatory behavior and the weight of the seminal plug expelled during ejaculation were recorded before and after sham surgery or bilateral denervation of the aforementioned muscles. Immediately after ejaculation, the animals were anesthetized to confirm the denervation and determine whether seminal material had accumulated in the lower urinary tract. In IC-denervated animals, voiding duration and the number of mounts was increased, and intromission or ejaculation patterns were absent. Denervation of BS induced signs of post-micturition dribble, decreased voiding frequency, increased urine volume and reduced the amount of semen ejaculated. Denervation of EUS induced signs of post-micturition dribble and urinary incontinence, as well as retrograde ejaculation. Striated muscles anatomically related to the urethra contribute differentially to the control of continence and expulsive urethral functions. Damages to the muscles or to their innervation, as may occur during pelvic surgery, would result in sexual and urinary dysfunctions.

Expression and function of CCL2/CCR2 in rat micturition reflexes and somatic sensitivity with urinary bladder inflammation

Chemokines are proinflammatory mediators of the immune response, and there is growing evidence for chemokine/receptor signaling involvement in pronociception. Bladder pain syndrome (BPS)/interstitial cystitis (IC) is a chronic pain syndrome characterized by pain, pressure, or discomfort perceived to be bladder-related with at least one urinary symptom. We have explored the expression and functional roles of CCL2 (monocyte chemoattractant protein-1) and its high-affinity receptor, CCR2, in micturition reflex function and somatic sensitivity in rats with urinary bladder inflammation induced by cyclophosphamide (CYP) treatment of varying duration (4 h, 48 h, chronic). Real-time quantitative RT-PCR, ELISAs, and immunohistochemistry demonstrated significant (P ≤ 0.01) increases in CCL2 and CCR2 expression in the urothelium and in Fast Blue-labeled bladder afferent neurons in lumbosacral dorsal root ganglia with CYP-induced cystitis. Intravesical infusion of RS504393 (5 μM), a specific CCR2 antagonist, reduced voiding frequency and increased bladder capacity and void volume in rats with CYP-induced cystitis (4 h), as determined with open outlet, conscious cystometry. In addition, CCR2 blockade, at the level of the urinary bladder, reduced referred somatic sensitivity of the hindpaw and pelvic region in rats with CYP treatment, as determined with von Frey filament testing. We provide evidence of functional roles for CCL2/CCR2 signaling at the level of the urinary bladder in reducing voiding frequency and somatic sensitivity following CYP-induced cystitis (4 h). These studies suggest that chemokines/receptors may be novel targets with therapeutic potential in the context of urinary bladder inflammation.

Selective control by posterior spinal nerve roots of micturition and erection in rats.

The posterior rootlets in L6 and S1 spinal cord of adult male Sprague-Dawley rats underwent electrostimulation. The bladder pressure, urethral perfusion pressure and intracavernous pressure were recorded. When some posterior rootlets of L6 and S1 were electrostimulated, the intracavernous pressure peaked rapidly, but the bladder pressure and the urethral perfusion pressure curve did not show great change. When other rootlets were stimulated, the bladder pressure changed greatly, but the urethral perfusion pressure and the intracavernous pressure did not show great change. When different rootlets were stimulated, the urethral perfusion pressure changed maximally, but there were no great changes in bladder pressure or intracavernous pressure. Furthermore, stimulation of some rootlets produced simultaneous changes in two or three different pressure measures mentioned above. The results demonstrate that regulation by L6 and S1 posterior rootlets of the rat bladder detrusor, external urethral sphincter and penis cavernous body are significantly distinct. Different rootlets can be distinguished by electrostimulation.

Spinal Cord FAAH in Normal Micturition Control and Bladder Overactivity in Awake Rats

We assessed whether spinal inhibition of the cannabinoid degrading enzyme FAAH would have urodynamic effects in normal rats and rats with bladder overactivity induced by partial urethral obstruction or prostaglandin E2. We also determined the expression of FAAH, and the cannabinoid receptors CB1 and CB2 in the sacral spinal cord. We used 44 rats for functional (cystometry) and Western blot experiments. The FAAH inhibitor oleoyl ethyl amide (3 to 300 nmol) was administered intrathecally (subarachnoidally) or intravenously. The expression of FAAH and CB1/CB2 receptors was determined by Western blot. Oleoyl ethyl amide given intrathecally affected micturition in normal rats and rats with bladder overactivity but effects were more pronounced in the latter. In normal rats oleoyl ethyl amide only decreased micturition frequency, while it decreased frequency and bladder pressures in rats with bladder overactivity. Intravenous oleoyl ethyl amide (3 to 300 nmol) had no urodynamic effect. FAAH and CB1/CB2 receptors were expressed in the rat sacral spinal cord. The expression of CB1/CB2 receptors but not FAAH was higher in obstructed than in normal rats. FAAH inhibition in the sacral spinal cord by oleoyl ethyl amide resulted in urodynamic effects in normal rats and rats with bladder overactivity. The spinal endocannabinoid system may be involved in normal micturition control and it appears altered when there is bladder overactivity.

The Effect of the 5-HT2A/2C Receptor Agonist DOI on Micturition in Rats with Chronic Spinal Cord Injury

We examined the effects of the 5-HT2A/2C receptor agonist DOI on micturition in chronic spinal cord injured rats. Female Sprague-Dawley® rats were used. Spinal cord injury was produced by transection at the T10 level. A cystometric study was performed 8 to 12 weeks after transection. Cystometrograms were done using urethane anesthesia in all rats. The selective 5-HT2A antagonist ketanserin was administered after each DOI dose-response curve. All drugs were administered intravenously. Compared to controls, spinal cord injured rats had higher bladder capacity and post-void residual urine volume, and lower voiding efficiency. In spinal cord injured rats DOI (0.01 to 0.3 mg/kg) induced significant dose dependent increases in micturition volume and decreases in residual volume, resulting in increased voiding efficiency. Cystometrogram measurements showed a dose dependent increase in high frequency oscillation activity, evident as an increased number of small oscillation per voiding. This correlated with the improved voiding efficiency. Ketanserin (0.1 mg/kg) partially or completely reversed the DOI induced changes. High frequency oscillation seems to reflect external urethral sphincter burst activity during voiding. Bladder voiding efficiency and high frequency oscillation activity were decreased in spinal cord injured rats. High frequency oscillation activity can be enhanced by 5-HT2A receptor agonism, improving voiding efficiency. To our knowledge it remains to be studied whether these results may have implications for the future treatment of voiding dysfunction in patients with spinal cord injury.

Results of a Randomized Phase III Trial of Mirabegron in Patients with Overactive Bladder

Many patients with overactive bladder discontinue pharmacotherapy due to suboptimal efficacy or side effects. Mirabegron, a β3-adrenoceptor agonist, may offer an effective and well tolerated alternative treatment for overactive bladder. A randomized, double-blind, placebo controlled trial was conducted in the United States and Canada. After a 2-week placebo run-in period, adults with overactive bladder symptoms for 3 or more months were randomized 1:1:1 to receive placebo, 50 or 100 mg mirabegron once daily for 12 weeks. Efficacy data were collected via patient completed diaries and quality of life assessments. Co-primary efficacy end points were changes from baseline to final visit in mean number of incontinence episodes per 24 hours and micturitions per 24 hours. Key secondary micturition and incontinence end points were also evaluated. Safety assessments included treatment emergent adverse events, laboratory assessments, vital signs, electrocardiograms and post-void residual volume. Compared to placebo, 50 and 100 mg mirabegron groups demonstrated statistically significantly greater mean decreases (95% CI) from baseline for incontinence episodes (-1.13 [-1.35, -0.91], -1.47 [-1.69, -1.25] and -1.63 [-1.86, -1.40]) and micturitions (-1.05 [-1.31, -0.79], -1.66 [-1.92, -1.40] and -1.75 [-2.01, -1.48]) per 24 hours (p <0.05). Significant improvements in all key secondary end points were observed for both mirabegron doses vs placebo. The incidence of frequently reported treatment emergent adverse events (hypertension, urinary tract infection, headache, nasopharyngitis) was similar in the mirabegron and placebo groups. Dry mouth was reported for 1.5%, 0.5% and 2.1% of patients in the placebo, 50 and 100 mg mirabegron groups, respectively. Once daily mirabegron in a 50 or 100 mg dose is an effective treatment for overactive bladder symptoms with a low occurrence of side effects.

Effect of Chemical Stimulation of the Medial Frontal Lobe on the Micturition Reflex in Rats

We assessed the influence of the medial frontal lobe on micturition after chemical stimulation. We also examined the relation between the medial frontal lobe and the rostral pontine reticular formation, which has a strong inhibitory effect on micturition. A total of 35 female rats underwent continuous cystometry. Bladder activity changes were examined after physiological saline, glutamate, the glutamate receptor antagonist MK-801, noradrenaline or the adrenergic α-1 receptor antagonist naftopidil was injected in the medial frontal lobe. When glutamate was injected in the medial frontal lobe, MK-801 was also injected in the rostral pontine reticular formation. Glutamate injection in the medial frontal lobe prolonged the interval between bladder contractions while injection of the glutamate antagonist MK-801 shortened the interval. Glutamate injection in the medial frontal lobe just after MK-801 injection in the ipsilateral rostral pontine reticular formation also prolonged the interval between bladder contractions. However, after prior injection of MK-801 in the bilateral rostral pontine reticular formation glutamate injection in the medial frontal lobe did not influence cystometric parameters. Noradrenaline injection in the medial frontal lobe shortened the interval between bladder contractions while injection of its antagonist naftopidil prolonged the interval. Medial frontal lobe neurons excited by glutamate inhibited the micturition reflex via activation of the rostral pontine reticular formation by glutamatergic projection while medial frontal lobe neurons excited by noradrenaline facilitated the micturition reflex. Thus, the medial frontal lobe may be an important integration center for the initiation of micturition and urine storage mechanisms.

Expression of Fatty Acid Amide Hydrolase (FAAH) in Human, Mouse, and Rat Urinary Bladder and Effects of FAAH Inhibition on Bladder Function in Awake Rats

BackgroundCannabinoid receptor (CB)–mediated functions may be involved in the regulation of bladder function, but information on endocannabinoid signals during micturition is scarce. ObjectiveInvestigate the expression of the endocannabinoid-degrading enzyme fatty acid amide hydrolase (FAAH) in human, rat, and mouse bladders and study the effects of inhibition of FAAH during urodynamics in awake rats. Design, setting, and participantsBladder tissue from humans, mice, and rats was used for measurements. Female Sprague-Dawley rats were administered the FAAH inhibitor oleoyl ethyl amide (OEtA) or vehicle intravenously (IV) or intravesically (IVES) with or without rimonabant (CB1 antagonist) or SR144528 (CB2 antagonist). MeasurementsReal-time transcriptase-polymerase chain reaction, Western blot, immunohistochemistry, and cystometry in awake rats. Results and limitationsMessenger RNA and protein for FAAH was expressed in the mucosa of human, mouse, and rat urinary bladders. Immunoreactivities for FAAH and CB2 were codistributed in rat and human urothelium. IV OEtA (0.3mg/kg) to rats increased intercontraction intervals (ICIs), micturition volume (MV), bladder capacity (BC), and threshold pressure (TP) by 17±1%, 16±1%, 17±1%, and 19±5%, respectively (all p<0.05 vs baseline). IVES OEtA (1 and 10mg/l) in rats dose-dependently increased (p<0.05 vs baseline) ICI (19±2% and 35±5%), MV (15±3% and 32±4%), BC (16±2% and 34±4%), and TP (15±1%, 21±3%). SR144528 (IVES 5mg/l) abolished all effects of OEtA, whereas rimonabant only counteracted effects of OEtA on TP. ConclusionsBladder mucosa of all species expressed FAAH. Rat and human urothelium coexpressed FAAH and CB2. The FAAH inhibitor OEtA altered urodynamic parameters that reflect sensory functions of micturition in rats. Suggesting a role for the endocannabinoid system in bladder mechanoafferent functions of rats, effects of IVES OEtA were abolished by an IVES CB2 antagonist and partly counteracted by an IVES CB1 antagonist.

The Effects of Acute and Chronic Psychological Stress on Bladder Function in a Rodent Model

Psychological stress plays a role in the exacerbation of functional lower urinary tract disorders, such as painful bladder syndrome and overactive bladder. To better understand the mechanism underlying this relationship, we characterized changes in micturition, anxiety-related behavior, and bladder pathology in rats exposed to repeated water avoidance (WA) stress. Twenty-four Wistar rats were subjected to WA stress or sham. Immediately after acute (day 1) and chronic (day 10) stress or sham, rats were placed in a metabolic cage for a 2-hour voiding behavior assessment. Voiding parameters were compared with baseline values obtained before stress. Four animals from each group were sacrificed on day 10 and bladders harvested for histologic and gene expression studies. The remaining 8 animals per group underwent repeated voiding assessment every 3 days for 1 month followed by 10 days of repeat WA stress or sham. Bladder histology and gene expression were studied. Rats exposed to WA stress developed a significant increase in micturition frequency and decrease in latency to void, voiding interval, and volume of first void compared with sham and baseline. Alterations in micturition persisted for approximately 1 month. Stressed rats showed increased fecal pellet excretion and anxiety-like behavior. In addition, bladder specimens from stressed animals revealed increased angiogenesis, and increased total and activated mast cells. In rats, repeated psychological stress results in lasting alterations in micturition frequency, interval, and volume. This rodent model may represent a valid tool for studying syndromes characterized by increased urinary frequency.

Improving voiding efficiency in the diabetic rat by a 5‐HT1A serotonin receptor agonist

Serotonin affects micturition in the normal rat through actions not only on ascending and descending spinal pathways and supraspinal centers but also on the lumbosacral spinal cord level. The selective 5-HT1A receptor agonist, 8-OH-DPAT((R)-(+)-8-hydroxy-2-(di-n-propylamino) tetralin), reversed detrusor-sphincter dyssynergia (DSD) in the spinal cord injury (SCI) rat. Rats with experimental diabetes mellitus (DM) have been shown to have both bladder and urethral dysfunction during reflex voiding. We therefore examined the effects of 8-OH-DPAT on micturition in DM rats. Female Sprague-Dawley rats were used. DM was induced by an intraperitoneal injection of streptozotocin (STZ, 65 mg/kg) and a cystometric study was performed 8 weeks post-injection. External urethral sphincter electromyography (EUS-EMG) was also measured. The 5-HT1A antagonist WAY-100635(N-tert-butyl-3-(4-(2-methoxyphenyl)-piperazin-1-yl)-2-phenylpropanamide) was administered after each 8-OH-DPAT dose-response. Compared to controls, DM rats had a higher bladder capacity, residual volume, and a lower voiding efficiency. In DM rats, 8-OH-DPAT (3-1,000 µg/kg, i.v.) induced significant dose-dependent increases in micturition volume, and decreases in residual volume, resulting in increases in voiding efficiency. During the micturition, there was a dose-dependent increased phasic EUS activity correlated with the improved voiding efficiency. WAY-100635 (300 µg/kg, i.v.) reversed the 8-OH-DPAT-induced changes. Both the bladder voiding efficiency and the periodic EUS activity were decreased in DM rats. 5-HT1A receptor agonism promoted periodic EUS activity, thereby improving voiding efficiency. Whether or not these results may have implications for the future treatment of voiding dysfunction in DM patients remains to be studied.

Transplantation of human adipose-derived stem cells into the urethra ameliorates stress urinary incontinence and blunts the induction of c-Fos immunoreactivities in brain areas related to micturition in female rats

Stress urinary incontinence (SUI) is a common condition that primarily affects women. Here, we investigate the effects of human adipose-derived stem cells (ADSCs) in a rodent model of SUI. Female Sprague-Dawley rats at 7 weeks of age were randomly divided into three groups (n=8 per group): sham-operation, SUI-induction by transabdominal urethrolysis, and SUI-induction followed by transplantation of human ADSCs into the urethra. The abdominal leak point pressure at 8 weeks after the operation was markedly decreased by transabdominal urethrolysis, confirming successful induction of SUI. Interestingly, transplantation of human ADSCs into the urethra significantly blunted the decrease of abdominal leak point pressure in SUI-induced rats. Accordingly, we observed expression of α-smooth muscle actin in a significant proportion of transplanted ADSCs, indicating differentiation of ADSCs into smooth muscle cells in the urethra. Moreover, the SUI-induced elevations of c-Fos immunoreactivities in the pontine micturition center (PMC) and in the ventrolateral periaqueductal gray (vlPAG) were clearly suppressed by transplantation of human ADSCs. These results imply that human ADSCs can be an effective therapeutic modality to ameliorate the symptoms of SUI.

Selective innervation of sacral anterior rootlets to micturition and erection function in rats

To investigate the selective innervation of sacral anterior rootlets to micturition and erection function in SD rats. Forty male SD rats of clean grade, aged 6 weeks old, were selected. Ten rats received a retrograde nerve tract tracing study. Thirty rats were chosen for an electro-physiological study. The L6, S1 spinal cord segment anterior rootlets of anesthetic rats were electrostimulated respectively. The intravesical pressure, urethral perfusion pressure and intracavernous pressure were recorded simultaneously and innervation effectiveness was analyzed. CB-HRP labeled neurons were observed mainly in L6 and S1 spinal cords. When some anterior rootlets of L6 and S1 were electrostimulated, the intravesical pressure rose gradually, but the urethral perfusion pressure and the intracavernous pressure curve changed slightly; when other rootlets of the same anterior root were stimulated, the urethral perfusion pressure could reach the peak; while others were stimulated, the intracavernous pressure rose quickly, but there were no great changes in intravesical pressure and urethral perfusion pressure. Some other rootlets might lead to the simultaneous changes of 2 or 3 above-mentioned pressures. The innervations of L6 and S1 anterior rootlets to rats' bladder detrusor, external urethral sphincter and penile cavernous body are significantly distinct. Different rootlets may be distinguished by microanatomy and electrostimulation.

Melatonin Increases Bladder Capacity via GABAergic System and Decreases Urine Volume in Rats

Impaired melatonin production is involved in disruption of the normal circadian pattern of sleep, which leads to nocturia in older adults. Melatonin improves nocturia. We hypothesized that melatonin could alleviate urinary frequency by suppressing the brain micturition center. We investigated the central contribution of melatonin to bladder function and urine volume. Cystometry was done in conscious female Sprague-Dawley rats (Japan SLC, Hamamatsu, Japan). We examined the effect of melatonin alone (4.3 x 10(-1) to 4.3 x 10 pmol intracerebroventricularly) or with the gamma-aminobutyric acid(A) antagonist bicuculline (5.0 x 10(-5) mg/kg intravenously) on bladder function. The influence of melatonin (4.3 x 10 pmol intracerebroventricularly) on urine volume was investigated in water loaded rats. Blood samples were collected to determine antidiuretic hormone, atrial natriuretic peptide and brain natriuretic peptide 4 hours after melatonin administration. Melatonin significantly increased bladder capacity dose dependently by 27.0%, 40.8% and 63.5% at 4.3 x 10(-1), 4.3 and 4.3 x 10 pmol, respectively, but had no significant effect on bladder contraction pressure. Bicuculline inhibited the melatonin induced increases in bladder capacity. Melatonin decreased urine volume in water loaded rats but plasma antidiuretic hormone, atrial natriuretic peptide and bladder contraction pressure were not changed. Results suggest that melatonin increases bladder capacity via gamma-aminobutyric acid(A) receptor in the brain and decreases urine volume. Thus, melatonin could be beneficial for nocturia via a central nervous system effect.

Organization of external urethral sphincter recruitment during micturition in the female rat.

The goal of this study was to examine the activation patterns of individual external urethral sphincter (EUS) motor units (MUs) during micturition in rats. To this end, bladder intravesical pressure (BP) and EUS EMG activity were recorded in urethane anesthetized adult female Sprague Dawley rats during continuous infusion of saline into the bladder. Individual EUS MU activity was recorded using a tungsten microelectrode (1 MΩ) inserted in the EUS or by placing an L6 ventral root filament, dissected in continuity, on platinum electrodes. Four different EUS MU recruitment patterns were observed. (1) Low threshold/sustained MUs were recruited prior to bladder contraction and exhibited sustained firing. (2) Medium threshold/not sustained MUs were recruited at the beginning of bladder contraction and stopped firing at the end of bladder contraction. (3) Medium threshold/sustained MUs were recruited at the beginning of bladder contraction and exhibited sustained firing. (4) High threshold/burst only MUs were recruited shortly before EUS bursting and stopped firing shortly after EUS bursting. Every EUS MU exhibited EUS bursting at the peak of bladder contraction. Among medium threshold MUs, derecruitment varied inversely with conduction velocity with slowly conducting units exhibiting longer sustained activity. This project is supported by the New York State Spinal Cord Injury Research Board.

Effects of Cannabinor, a Novel Selective Cannabinoid 2 Receptor Agonist, on Bladder Function in Normal Rats

BackgroundCannabinoid (CB) receptors may be involved in the control of bladder function; the role of CB receptor subtypes in micturition has not been established. ObjectivesOur aim was to evaluate the effects of cannabinor, a novel CB2 receptor agonist, on rat bladder function. Design, setting, and participantsSprague Dawley rats were used. Distribution of CB2 receptors in sensory and cholinergic nerves of the detrusor was studied. Selectivity of cannabinor for human and rat CB receptors was evaluated. Effects of cannabinor on rat detrusor and micturition were investigated. MeasurementsImmunohistochemistry, radioligand binding, tritium outflow assays, organ bath studies of isolated bladder tissue, and cystometry in awake rats were used. Results and limitationsCB2 receptor immunoreactivity was expressed in the urothelium and in sensory and cholinergic bladder nerves. Cannabinor exhibited similar binding at human and rat CB2 receptors and a 321-fold functional selectivity for the CB2 receptor versus the CB1 receptor. Cannabinor had no effect on isolated detrusor muscle function. In vivo, cannabinor 3.0mg/kg increased micturition intervals and volumes by 52% (p<0.05) and 96% (p<0.01), respectively, and increased threshold and flow pressures by 73% (p<0.01) and 49% (p<0.001), respectively. Cannabinor 0.3 or 1.0mg/kg or vehicle did not affect urodynamic parameters. ConclusionsConsidering that CB2 receptors are localized on sensory nerves and on the urothelium and that cannabinor had effects on “afferent” urodynamic parameters, peripheral CB2 receptors may be involved in sensory functions of rat micturition. Effects of cannabinor on cholinergic nerve activity in normal bladder tissue appear to be limited.

Influence of glutamatergic projections to the rostral pontine reticular formation on micturition in rats

Aims We examined the effect of injecting glutamate or a glutamate receptor antagonist into the rostral pontine reticular formation (RPRF) on the micturition reflex in anesthetized rats and conscious rats. Main method Forty-eight female rats were divided into an isovolumetric cystometry group and a continuous cystometry group. Under urethane anesthesia or while conscious, physiological saline, glutamate, or MK-801 (a glutamate receptor antagonist) was injected into the RPRF, and then the changes of bladder activity were examined. Key findings There was no significant change of bladder activity after injection of physiological saline. In anesthetized rats, the injection of either glutamate or MK-801 into the RPRF transiently inhibited bladder contractions. There was a complete recovery of bladder activity 10–20 min after glutamate or MK-801 injection and there were no significant changes of cystometry parameters after the recovery of bladder contractions. In conscious rats, injection of glutamate into the RPRF prolonged the interval between bladder contractions and decreased the baseline bladder pressure. On the other hand, injection of MK-801 into the RPRF caused numerous small bladder contractions, some of which were accompanied by a leakage of a small amount of fluid from around the urethral catheter. Significance RPRF neurons receive glutamatergic projections, possibly from the forebrain, and the RPRF inhibits the micturition reflex pathway. RPRF neurons are also regulated by inhibitory interneurons, which receive glutamatergic projections as well. Therefore, the RPRF plays an important role in the regulation of urine storage.

Mechanisms by which the serotonergic system inhibits micturition in rats

Aims Serotonergic neurons and amino acid neurons are involved in the central nervous control of lower urinary tract function. We investigated the role of the serotonergic system in the central regulation of micturition, as well as the relationship between serotonergic neurons and amino acid neurons in the lumbosacral cord of rats. Main methods Under urethane anesthesia, bladder and urethral activity were recorded before and after intrathecal injection of serotonin (5-hydroxytryptamine: 5-HT), a 5-HT 2A receptor antagonist (ketanserin: KET), or KET + 5-HT by isovolumetric cystometry and measurement of the urethral pressure in intact rats and rats with hypogastric nerve transection (HGNT). Amino acid levels in the lumbosacral cord were also measured after intrathecal injection of 5-HT in intact rats. Key findings In intact rats, intrathecal injection of 5-HT transiently abolished rhythmic bladder contractions, decreased the maximal bladder contraction pressure, and increased the intravesical baseline pressure and the urethral baseline pressure. Intrathecal injection of KET + 5-HT also transiently abolished rhythmic bladder contractions. In HGNT rats, intrathecal injection of 5-HT transiently abolished rhythmic bladder contractions and increased the urethral baseline pressure. Intrathecal injection of 5-HT decreased the level of glycine in the lumbosacral cord. Significance The serotonergic system may be involved in blocking the afferent pathway of the micturition reflex, increasing sympathetic activity, and secondary promotion of urethral contraction through inhibition of glycinergic neurons in the lumbosacral cord. 5-HT 2A receptors may be involved in these effects on the bladder and urethra. Therefore, the serotonergic system may play a role of the maintenance of urine storage.

Dynamic Performance Evaluation on the Synergy of Micturition in Spinal Cord-Injured Female Rats under Pharmacological Effects

Recently, a temporal "coherent" fractal structure and synchronization of rhythms were proposed as two essential indicators for efficient voiding during micturition in female rats. The former was correlated with the intensity and the latter the frequency of physiological signals embedded in random noise. Studies using both indices confirmed that synergic co-activations of bladder and external urethral sphincter (EUS) of female rats were present during the voiding of urine. Therefore, it would be interesting to investigate if these two criteria could be used in the performance evaluation of pharmacological effects on spinal cord-injured rats during micturition. In this paper, the primary goals were to (1) examine if the involved muscles in the lower urinary tract would be under similar synergic co-activations during the administration of capsaicin (CAP) and resiniferatoxin (RTX), and (2) characterize quantitatively the differences of their nervous responses simultaneously. A total of 62 micturition experiments were performed on sixteen spinal cord-injured adult female Sprague-Dawley rats, and then the electromyograms of EUS and cystometrograms of bladder were analyzed. Results based on the aforementioned criteria indicated that the synergy of bladder and EUS during micturition by using RTX was better than that of the CAP. Furthermore, the residue urine volumes for rats under the former treatment were smaller than those of the rats under the latter treatment. Consequently, we concluded that the administration of RTX was more effective than CAP in facilitating voiding in the spinal cord-injured rats.

PACAP-Mediated ATP Release from Rat Urothelium and Regulation of PACAP/VIP and Receptor mRNA in Micturition Pathways after Cyclophosphamide (CYP)-Induced Cystitis

Pituitary adenylate cyclase-activating peptide (PACAP) peptides are expressed in micturition pathways, and PACAP expression is regulated by urinary bladder inflammation. Previous physiological studies have demonstrated roles for PACAP27 and PACAP38 in detrusor smooth muscle (DSM) contraction and a PAC1 receptor antagonist reduced cyclophosphamide (CYP)-induced bladder hyperreflexia. To gain insight into PACAP signaling in micturition and regulation with cystitis, receptor characterization by real-time quantitative polymerase chain reaction and physiological assays were performed. PACAP receptors were identified in tissues of rat micturition pathway, including DSM, urothelium (U), and dorsal root ganglia (DRG) after acute (4 h), intermediate (48 h) or chronic (8 days) CYP-induced cystitis. PAC1 messenger RNA expression significantly (p < or = 0.05) increased in U and DSM after 48 h and chronic CYP-induced cystitis after an initial decrease at 4 h. VPAC1 and VPAC2 transcripts increased in U and DSM after acute and intermediate CYP-induced cystitis followed by a decrease in VPAC2 expression with chronic cystitis. Application of PACAP27 (100 nM) to cultured urothelial cells evoked adenosine triphosphate (ATP) release that was blocked by the PAC1 specific antagonist, M65 (1 microM). PACAP38 (100 nM) also evoked ATP release from cultured urothelial cells, but ATP release was less than that observed with PACAP27. PACAP transcripts were increased in the U with intermediate and chronic cystitis, whereas vasoactive intestinal polypeptide (VIP) expression in both tissues was very low and showed no regulation with cystitis. Regulation of PACAP, galanin, and substance P transcripts expression was observed in lumbosacral DRG, but no regulation for VIP was observed. The current data demonstrate PACAP and PAC1 regulation in micturition pathways with inflammation and PACAP-mediated ATP release from urothelium.

Voiding Pattern Analysis as a Surrogate for Cystometric Evaluation in Uroplakin II Knockout Mice

Previous study has shown that the absence of uroplakin II can cause urinary tract dysfunction, including vesicoureteral reflux and renal abnormalities, as well as micturition pattern changes. We developed a simple surrogate measure of bladder function using ultraviolet visualization of urinary voiding patterns in a uroplakin II knockout mouse animal model. Three male and 3 female WT mice, and 3 male and 3 female uroplakin II knockout mice were evaluated by cystometric analysis and voiding pattern markings. Voiding pattern markings were graded by independent observers on a scale of 1 to 5 according to the degree of dispersion of voided urine. Statistical analysis was then used to correlate voiding dispersion grades with cystometric parameters in the same mice. The degree of dispersion of voiding pattern markings correlated with several measures of bladder function. Specifically the Pearson correlation coefficients for the observed voiding patterns highly correlated with baseline pressure, threshold pressure and intermicturition pressure measurements made during conscious cystometry in these mice (p <0.05). Ultraviolet visualization of urinary voiding patterns of mice correlated well with certain measures of standard cystometric evaluations. As such, this method provides a simple, noninvasive method of evaluating mouse bladder function. Implementation of this methodology, which can potentially be automated for high throughput analysis, can accelerate the development of novel therapy for certain important aspects of bladder disease/dysfunction.