Increase In Bladder Mass Research Articles

Gender-dependent bladder response to one-day bladder outlet obstruction

Development of bladder fibrosis, loss of compliance, and voiding dysfunction are among the severe consequences of various lower urinary conditions, for example, bladder outlet obstruction (BOO), neurogenic bladder, and radiotherapy to the pelvic area. The bladder remodelling results in significant changes in bladder function and architecture, and may ultimately be deleterious for kidney function. The molecular signals underlying pathologic bladder remodelling, as well as the impact of gender, remain poorly understood. To investigate the bladder remodelling after one day BOO, whether the remodelling is different between different bladder sections, and whether genders may affect the remodelling. Thirty male and 30 female C57BL/6NRj mice were randomly divided into Control, Sham and BOO groups with ten mice per group. A 24-h total urethral obstruction was performed at the proximal urethra. Histological changes were observed via H&E, trichrome and immunohistochemistry staining. Harvested bladders were divided into upper and lower sections for analysis. Protein and gene expression were detected by Western blotting and qPCR. No significant changes in bladder wall thickness were observed following BOO, while increased bladder mass after BOO was found in female mice only. We detected FN and ⍺-SMA upregulation in the male upper bladder segment. Female BOO mice bladders showed increased α-SMA expression in both bladder segments, but no difference of FN was observed in either bladder segments. BOO-induced upregulation of TGF-β and Gremlin were detected in both male and female bladders, while downregulation of BMP-7 was detected only in male bladders. Furthermore, phosphorylation of both SMAD2/3 and SMAD1/5/9 were increased in male bladders following BOO, whereas female mice exhibited increased pSMAD2/3 in the upper and increased pSMAD1/5/9 in the lower bladder segment. Our data indicate that some specific proteins and growth factors were detected as early alterations of tissue which may lead to fibrosis. In addition, the males tended to have more pronounced response than females. However, the causes and consequences of the findings need to be further investigated.

New mouse model of underactive bladder developed by placement of a metal ring around the bladder neck.

To develop a new mouse model of underactive bladder (UAB) caused by chronic bladder outlet obstruction (BOO). BOO was created in 6-week-old male C57BL/6 mice using surgery to loosely place a silver jump ring around the bladder neck of each mouse. Micturition behavior (assessed with a metabolic cage) and cystometry were used to evaluate bladder function at 8 and 16 weeks after BOO. Following completion of the functional studies, the bladders of the mice were excised, weighed, and subjected to histological analysis. Micturition behavior analysis showed that mice subjected to BOO for 16 weeks had a lower frequency of micturition (7.3 ± 1.1 vs 12.5 ± 3.0 times/d, P < .05) and volume per void (106.0 ± 0.1 vs 133.9 ± 3.2 μL, P < .05) than mice subjected to BOO for 8 weeks. Cystometry revealed that mice subjected to BOO for 16 weeks had lower baseline pressure (8.4 ± 0.6 vs 14.0 ± 0.7 cmH2 O, P < .01) and micturition pressure (13.9 ± 1.1 vs 42.8 ± 1.7 cmH2 O, P < .05) than mice subjected to BOO for 8 weeks. BOO caused progressive increases in bladder mass and collagen deposition over time. We successfully established a novel mouse model of UAB using surgery to place a silver jump ring loosely on the bladder neck. BOO initially induced bladder overactivity but subsequently resulted in UAB due to deterioration of detrusor smooth muscle contractility and progressive deposition of collagen in the bladder wall.

Evaluation of combined therapeutic effects of hydrogen sulfide donor sodium hydrogen sulfide and phosphodiesterase type-5 inhibitor tadalafil on erectile dysfunction in a partially bladder outlet obstructed rat model.

To evaluate the impacts of hydrogen sulfide (H2 S) donor, sodium hydrogen sulfide (NaHS), and phosphodiesterase type-5 inhibitor (PDE5i), tadalafil per se and their combination treatment on partial bladder outlet obstruction (PBOO)-induced erectile dysfunction (ED). Sprague-Dawley rats were equally divided into five groups: (a) sham-operated control; (b) PBOO; (c) PBOO-treated with NaHS (5.6 mg/kg/day, ip); (d) PBOO-treated with tadalafil (2 mg/kg/day, oral); and (e) PBOO-treated with combination of NaHS and tadalafil. The obstruction was created by urethral ligation for 6 weeks. In vivo erectile responses, in vitro relaxant and contractile responses in penile tissue as well as protein expression of nitric oxide synthases (NOS), H2 S synthesis enzymes, oxidative stress, hypoxia, fibrosis markers, and the smooth muscle/collagen ratio and apoptosis were analyzed. Combined treatment entirely returned increased bladder mass, reduced erectile responses, relaxation responses to acetylcholine, and electrical field stimulation in obstructed rats, while partial amelioration was observed after mono-treatment. Decreased neuronal NOS and 3-mercaptopiruvate transferase enzyme expressions in penile tissue from obstructed rats were also entirely restored by the combined treatment. Mono-treatment partially improved increased hypoxia, oxidative stress, fibrosis markers, decreased smooth muscle mass, and H2 S levels, while combined therapy completely recovered. The combination therapy with H2 S donor and PDE5i had positive effects on erectile responses through the improvement of ischemia-induced morphological and functional penile alterations in obstruction. H2 S and NO may likely play a synergistic role in the regulation of erectile function and have constructive effects on clinical outcomes in male patients with ED and benign prostatic hyperplasia/lower urinary tract symptoms.

Murine social stress results in long lasting voiding dysfunction

Repeated exposure to social stress shifts the voiding phenotype in male mice leading to bladder wall remodeling and is associated with increased expression of the stress neuropeptide, corticotropin-releasing factor (CRF) in Barrington's nucleus neurons. In these studies, we set out to determine if the voiding phenotype could recover upon removal from the stressor. Male mice were exposed for 1h daily to an aggressor and the voiding phenotype was assessed at one month followed by randomization to three groups. One group underwent immediate sacrifice. Two groups were allowed a one month recovery from the social stress exposure with or without the addition of fluoxetine (1.2mg/ml) in their drinking water and repeat voiding patterns were measured prior to sacrifice. Social stress significantly increased bladder mass, bladder mass corrected for body weight, voided volumes, and decreased urinary frequency. The abnormal voiding phenotype persisted after a 1month recovery with no effect from the addition of fluoxetine. CRF mRNA in Barrington's nucleus was increased by social stress and remained elevated following recovery with no effect from the addition of fluoxetine. The mRNA and protein expression for the alpha 1 chains of type 1 and type III collagen was unchanged across all groups suggesting that changes in the extracellular matrix of the bladder are not responsible for the voiding phenotype. This persisting voiding dysfunction correlates with the persistent elevation of CRF mRNA expression in Barrington's nucleus.

Nerve-sparing Mid-urethral Obstruction (NeMO) in Female Small Rodents.

Partial bladder outlet obstruction (pBOO) has a high prevalence, causes significant patient burden, and immense health care costs. The most common animal model to investigate bladder remodeling in pBOO are female rodents undergoing partial obstruction at the proximal urethra. Variability in the degree of obstruction and animal mortality are major concerns with proximal obstruction. Furthermore, dissecting around the proximal urethra and bladder neck jeopardizes bladder innervation. We developed a nerve-sparing mid-urethral obstruction (NeMO) model for pBOO avoiding the disadvantages of the traditional model. We approached the urethra just inferior to the pubic symphysis, which obviated the need for laparotomy as well as for dissection in this area; also, the striated urethral sphincter remained untouched. We performed NeMO in female Sprague-Dawley rats (12 obstructions, 6 sham animals) as well as in female C57/bl6 mice (20 obstructions, 18 sham animals). After two weeks, we evaluated bladder function, bladder mass, and body mass. We had no mortalities among obstructed- or sham-operated female rats; as described for the traditional proximal pBOO-method, we tied the suture around the proximal urethra and a temporarily placed 0.9 mm metal rod. NeMO induced an 85% increase in bladder mass after two weeks, average residual urine volume was 0.4 mL in partially obstructed rats while only 0.03 mL in sham animals. In mice, we tested 3 sizes of cannulas that we placed along the urethra when tying the suture. We found that using a 27-gauge cannula resulted in over 50% animal mortality; placing the 25-gauge cannula did not yield the desired response in increasing bladder mass; utilizing a 26-gauge cannula yielded favorable results with minimal animal mortality (1/8) yet a significant 2-fold increase in bladder mass.

Finding NeMO—Nerve-sparing Mid-urethral Obstruction: A Pathophysiologically Accurate Model of Rodent Partial Bladder Outlet Obstruction

To develop and evaluate a novel technique modeling partial bladder outlet obstruction (pBOO) using a nerve-sparing mid-urethral obstruction (NeMO) approach. Female unoperated rats were compared to rats after NeMO, NeMO sham, proximal urethral (PU) obstruction, or PU sham. Residual volume, bladder capacity, voiding volume, and bladder mass were recorded; the contractile characteristics of isolated bladder strips were also analyzed. Additionally, we quantitated nerve fibers at the bladder neck as well as the extracellular matrix in the bladder wall. NeMO yields a more predictable degree of obstruction vs PU, causes no animal mortality, and is easy to release. NeMO also results in a more moderate increase in bladder mass commensurate with human disease vs the exaggerated response to PU, and does not lead to the excessive bladder dilation observed after PU while showing increased residual urine and fibrosis over time, thus closely modeling human pBOO pathophysiology. Importantly, PU shams significantly incite both an undesirable mass increase as well as bladder dysfunction, correlating with a denervation injury making them unsuitable as controls when modeling a non-neurogenic pBOO. The bladder physiology and structure of NeMO-sham animals were indistinguishable from those of unoperated controls. The low complication rate and low variability of NeMO also can be applied to mice, opening the pBOO field to the full spectrum of transgenic manipulation. NeMO is a pathophysiologically accurate modeling approach, with low variability and mortality, and newly paves the way for realistic and robust interpretation of omics and sequencing analytical methodologies. We therefore suggest NeMO as a new standard model when investigating pBOO.

Estrogen Receptor-α is a Key Mediator and Therapeutic Target for Bladder Complications of Benign Prostatic Hyperplasia

Estrogens are important in prostate growth and have a role in benign prostatic hyperplasia. However, to our knowledge no current therapy directly targets estrogen action. Estrogens act primarily via estrogen receptors α and β. In a mouse model we evaluated the relative contribution of these receptors to bladder complications of benign prostatic hyperplasia. We also evaluated the prevention of these bladder complications using the selective estrogen receptor modulators raloxifene and tamoxifen (estrogen receptor-α selective antagonists), and R,R-THC (estrogen receptor-β selective antagonist). Adult male C57bl/6 mice received implants of 25 mg testosterone and 2.5 mg 17β-estradiol slow release pellets. Untreated controls underwent sham surgery. We evaluated the contributions of the estrogen receptor subtypes in ERαKO and ERβKO mice compared to their respective wild-type litter mates. Wild-type mice treated with testosterone plus 17β-estradiol were compared to mice treated with testosterone plus 17β-estradiol and 25 mg selective estrogen receptor modulators to evaluate the prevention of benign prostatic hyperplasia complications by selective estrogen receptor modulators. Large bladders with urinary retention developed in ERαWT and ERβWT litter mates treated with testosterone plus 17β-estradiol but such bladders did not develop in ERαKO mice treated with testosterone plus 17β-estradiol. ERβKO mice treated with testosterone plus 17β-estradiol had large bladders with urinary retention and increased bladder mass. Cotreatment with the estrogen receptor-α antagonist raloxifene resulted in decreased bladder mass compared to that in wild-type mice treated with testosterone plus 17β-estradiol. Bladders in mice treated with the estrogen receptor-β antagonist R,R-THC were similar to those in testosterone plus 17β-estradiol treated mice. Estrogen receptor-α but not β is a key mediator of bladder complications of benign prostatic hyperplasia and a potential target for future therapies.

Partial Bladder Outlet Obstruction in Mice May Cause E-Cadherin Repression through Hypoxia Induced Pathway

Posterior urethral valves are the most common cause of partial bladderoutlet obstruction in the pediatric population. Posterior urethral valves is a devastating clinical problem that ultimately results in urinary incontinence, neurogenic bladder and renal impairment. Despite improvements in medical and surgical management at least a third of patients with this condition progress toend stage renal disease and half will have problems with urinary incontinence. To achieve better understanding of the mechanism associated with clinical events we generated partial bladder outlet obstruction in male mice. In this model we investigated pathological consequences and underlying molecular mechanisms secondary to partial bladder outlet obstruction. Five to 8-week-old male C57BL/6 mice were divided intoa surgical obstruction group and a sham operated group that served as controls. Bladders and kidneys were harvested from each group 1, 2, 3, 5 and 7days postoperatively, respectively. We examined histological and biochemical alterations, and further investigated our hypothesis that partial bladder outlet obstruction induces hypoxia activated profibrotic signaling and changes in gene expression in the bladder. Mice with partial bladder outlet obstruction demonstrated significant increases in bladder mass and urinary retention compared to sham operated mice.Obstruction caused fibrosis in the bladder and induced up-regulation of profibrotic genes, hypoxia-inducible factors and epithelial-mesenchymal transition-inducing transcription factors, resulting in E-cadherin down-regulation. Obstruction induced significant histological and molecular alterations, including activation of the hypoxia-inducible factors pathway in the mouse bladder. Activation of epithelial-mesenchymal transition-inducing transcription factors by hypoxia-inducible factors might have an important role in thepathogenesis of partial bladder outlet obstruction.

Effects of Chronic Treatment With Cilostazol, a Phosphodiesterase 3 Inhibitor, on Female Rat Bladder in a Partial Bladder Outlet Obstruction Model

To investigate whether bladder dysfunction after bladder outlet obstruction (BOO) could be altered by treatment with cilostazol, a phosphodiesterase 3 inhibitor (PDE3i). Twelve-week-old female Sprague-Dawley rats were divided into 5 groups: groups 1 and 2, sham-operated rats and groups 3-5, BOO rats. Group 1 and 3 rats were given normal diet, group 2 and 5 rats were given high-dose PDE3i diet, and group 4 rats were given low-dose PDE3i diet. PDE3i was given within diet from the day of surgery. Four weeks after BOO, the bladder was excised and dissected into 4 longitudinal strips for isometric organ-bath assay. Contractile responses of bladder strips to electrical field stimulation (EFS), carbachol, and potassium chloride (KCl) were determined for each group. BOO induced a significant increase in bladder weight in groups 3-5 compared with groups 1 and2. PDE3i treatment did not affect bladder weight in sham or BOO rats. Contractile forces in response to EFS, carbachol, and KCl in group 3 were about 20%-40% of those in group 1. Contractile responses to EFS or KCl in PDE3i-treated BOO rats were not significantly different from those in group 3. Only high dose of PDE3i treatment in BOO rats caused a statistically significant increase in the response to carbachol compared with group3. PDE3i has a small but significant protective effect on the contractile dysfunction induced by a 4-week BOO in rats, although the increase in bladder mass was not altered. PDE3i could be auseful protection against contractile dysfunction of the obstructed bladder.

Large-conductance calcium-activated potassium channel activity, as determined by whole-cell patch clamp recording, is decreased in urinary bladder smooth muscle cells from male rats with partial urethral obstruction

To examine the effect of partial urethral obstruction (PUO) on bladder smooth muscle outward potassium current and the contribution of the large-conductance calcium-activated potassium (Maxi-K, BKCa) channel to this activity in smooth muscle cells isolated from bladders of sham-operated and PUO male rats using whole-cell patch clamp recording techniques. To determine the effect of PUO on the expression of the Maxi-K channel α and β1 subunits and in vitro detrusor contractility. Twenty adult male Sprague-Dawley rats were divided equally into two groups and subjected to surgical ligation of the urethra (PUO) or sham surgery (SHAM). After 2 weeks, the detrusors from PUO and SHAM rats were used for molecular analyses (mRNA and protein quantification of Maxi-K subunits) or organ bath contractility studies, or myocytes were isolated for conventional whole-cell patch clamp analyses. PUO increased bladder mass 2.5-fold and detrusor strips exhibited a more tonic-type contraction and increased contractility compared with controls (SHAM). Iberiotoxin (300 nM) sensitive Maxi-K channel current comprised about 40% of the outward whole-cell current in SHAM bladders but only about 8% in PUO bladders. Expression of the α subunit of the Maxi-K channel was significantly decreased ~40% while the expression of the β1 subunit was increased ~2-fold at the mRNA level. The increase in β1 expression was confirmed by Western blotting. Our findings show that obstruction of the rat bladder is associated with decreased Maxi-K channel activity of bladder smooth muscle cells, determined via direct current measurement. Increased expression of the β1 subunit points to a compensatory reaction to decreased Maxi-K channel activity. Maxi-K channel openers or gene therapy may therefore provide therapeutic benefit for the overactive bladder.

Bladder outlet obstruction: progression from inflammation to fibrosis

To investigate the progression of urodynamic changes, as well as histological and biochemical outcomes over a prolonged period of partial bladder outlet obstruction (pBOO) in an animal model with physiologically relevant pBOO. Healthy, adult, female Fischer rats underwent surgical creation of a pBOO for either 2, 4, 8, or 13 weeks and were compared with sham-operated rats. Urodynamic measurements were used to compare bladder volumes and pressure. Tissue was grossly analysed with light microscopy and bladder weights and thicknesses were compared. Reverse transcription-polymerase chain reaction for collagen, transforming growth factor β (TGF-β), connective tissue growth factor (CTGF), hypoxia inducible factor 1α (HIF-1α), and platelet-derived growth factor (PDGF-A) was performed on all samples, as well as immunohistochemistry (IHC) for α-smooth muscle actin (α-SMA). Finally, mass spectrometry was used to quantify the collagen content of the bladders as a measure of fibrosis. After induction of pBOO, all rats remained healthy. Initial urodynamics showed an increase in capacity while maintaining normal pressures, but then deteriorated into small capacity, high-pressure bladders. Haematoxylin and eosin (H&E) staining showed an initial inflammatory response, and this was confirmed with significantly increased mRNA levels of TGF-β, CTGF, HIF-1α, and PDGF. The progression to smooth muscle hypertrophy was evident on H&E and confirmed with increased bladder mass and thickness. IHC for α-SMA showed a progressive increase associated with the elevated bladder pressures. Masson's trichrome and mass spectrometry showed a progressive increase in collagen to 13 weeks. With this model, we have effectively replicated the clinical scenario, with significant pathophysiological changes occurring insidiously in otherwise healthy rats. We believe that our observed changes represent distinct phases of bladder decompensation; with an initial inflammatory response to the stress of the pBOO, smooth muscle hypertrophy to overcome the increased urethral resistance, and eventual decompensation to fibrosis. The time course of the inflammatory markers implies the need for early intervention to prevent this cascade. Novel strategies targeting these observed physiological responses could lead to improved preventative strategies, with respect to biochemical pathways and the time course of their initiation.

Bladder Dysfunction and Altered Somatic Sensitivity in PACAP −/− Mice

PACAP and receptors are expressed in micturition pathways. Studies show that PACAP has a role in detrusor smooth muscle contraction to facilitate adenosine triphosphate release from urothelium and PACAP antagonism decreases cyclophosphamide induced bladder hyperreflexia. PACAP contributions to micturition and somatic sensation were studied in PACAP knockout (PACAP(-/-)), litter mate heterozygote (PACAP(+/-)) and WT mice by conscious cystometry with continuous intravesical saline or acetic acid (0.5%) instillation, urination patterns, somatic sensitivity testing of hind paw and pelvic regions with calibrated von Frey filaments, and morphological bladder assessments. PACAP(-/-) mice had an increased bladder mass with fewer but larger urine spots. In PACAP(-/-) mice the lamina propria and detrusor smooth muscle were significantly thicker but the urothelium was unchanged. PACAP(-/-) mice had increased bladder capacity, voided volume and intercontraction interval with significantly increased detrusor contraction duration and large residual volume. WT mice responded to acetic acid (0.5%) with a decrease in voided volume and intercontraction interval but PACAP(+/-) and PACAP(-/-) mice did not respond. PACAP(-/-) mice were less responsive to somatic stimulation. PACAP(+/-) mice also had bladder dysfunction, and somatic and visceral sensory abnormalities but to a lesser degree. PACAP gene disruption contributes to changes in bladder morphology and function, and somatic and visceral hypoalgesia.

Cyclical estrogen and free radical damage to the rabbit urinary bladder

There are a number of lower urinary tract dysfunctions (LUTD) that occur primarily in women. Our hypothesis is that cyclical estrogen will produce LUTD in part by the generation of free radicals and oxidative damage to cellular and subcellular membranes. Twenty female rabbits were divided into five groups: control, ovariectomized (Ovx), Ovx receiving continuous estrogen, Ovx receiving cyclical estrogen ending off estrogen, and Ovx receiving cyclical estrogen ending on estrogen. Statistical analyses used ANOVA followed by the Tukey analysis for individual differences. High estrogen increased bladder mass, contraction, compliance, and blood flow and decreased oxidative damage. Low estrogen decreased bladder mass, contraction, compliance, and blood flow and increased oxidative damage. The decreased blood flow associated with increased oxidative damage demonstrates that cyclical damage to cellular membranes occurs. This supports the hypothesis that cycling estrogen may play a role in the etiology of LUTD of women.

Underlying Mechanisms of Detrusor Overactivity Following Bladder Outlet Obstruction

Several studies have demonstrated that many pathophysiological mechanisms can contribute to the development of detrusor overactivity associated with bladder outlet obstruction (BOO). BOO induces a rapid hypertrophy characterized by increased bladder mass and collagen deposition, which play an important role in the intercellular transmission of active force. Detrusor overactivity may be attributed to an increase in afferent activation and neuroplasticity, and nerve growth factor and transient receptor potential vanilloid 1 (TRPV1) are involved in this process. The change in the urothelium and urothelial signaling can also lead to increased afferent nerve activity and contribute to irritative symptoms accompanying outlet obstruction. This is because afferent neuronal fibers are located near the urothelium, and urothelial cells release neurotransmitters in response to bladder wall stretching, as a neuron‐like property. Recently, it has been reported that intercellular communication through gap junctions might participate in the pathophysiology of detrusor overactivity associated with BOO, and these gap junction proteins are differentially regulated during BOO. In addition, the changes in several receptors, such as the muscarinic and purinergic receptors, could have a role in mediating the afferent sensory responses and development of detrusor overactivity following BOO.

Social stress in mice induces voiding dysfunction and bladder wall remodeling

Several studies have anecdotally reported the occurrence of altered urinary voiding patterns in rodents exposed to social stress. A recent study characterized the urodynamic and central changes in a rat model of social defeat. Here, we describe a similar voiding phenotype induced in mice by social stress and in addition we describe potential molecular mechanisms underlying the resulting bladder wall remodeling. The mechanism leading to the altered voiding habits and underlying bladder phenotype may be relevant to the human syndrome of dysfunctional voiding which is thought to have a psychological component. To better characterize and investigate social stress-induced bladder wall hypertrophy, FVB mice (6 wk old) were randomized to either social stress or control manipulation. The stress involved repeated cycles of a 1-h direct exposure to a larger aggressive C57Bl6 breeder mouse followed by a 23-h period of barrier separation over 4 wk. Social stress resulted in altered urinary voiding patterns suggestive of urinary retention and increased bladder mass. In vivo cystometry revealed an increased volume at micturition with no change in the voiding pressure. Examination of these bladders revealed increased nuclear expression of the transcription factors MEF-2 and NFAT, as well as increased expression of the myosin heavy chain B isoform mRNA. BrdU uptake was increased within the urothelium and lamina propria layers in the social stress group. We conclude that social stress induces urinary retention that ultimately leads to shifts in transcription factors, alterations in myosin heavy chain isoform expression, and increases in DNA synthesis that mediate bladder wall remodeling. Social stress-induced bladder dysfunction in rodents may provide insight into the underlying mechanisms and potential treatment of dysfunctional voiding in humans.

Urinary Bladder Function and Somatic Sensitivity in Vasoactive Intestinal Polypeptide (VIP)−/− Mice

Vasoactive intestinal polypeptide (VIP) is an immunomodulatory neuropeptide widely distributed in neural pathways that regulate micturition. VIP is also an endogenous anti-inflammatory agent that has been suggested for the development of therapies for inflammatory disorders. In the present study, we examined urinary bladder function and hindpaw and pelvic sensitivity in VIP(-/-) and littermate wildtype (WT) controls. We demonstrated increased bladder mass and fewer but larger urine spots on filter paper in VIP(-/-) mice. Using cystometry in conscious, unrestrained mice, VIP(-/-) mice exhibited increased void volumes and shorter intercontraction intervals with continuous intravesical infusion of saline. No differences in transepithelial resistance or water permeability were demonstrated between VIP(-/-) and WT mice; however, an increase in urea permeability was demonstrated in VIP(-/-) mice. With the induction of bladder inflammation by acute administration of cyclophosphamide, an exaggerated or prolonged bladder hyperreflexia and hindpaw and pelvic sensitivity were demonstrated in VIP(-/-) mice. The changes in bladder hyperreflexia and somatic sensitivity in VIP(-/-) mice may reflect increased expression of neurotrophins and/or proinflammatory cytokines in the urinary bladder. Thus, these changes may further regulate the neural control of micturition.

A porcine model of bladder outlet obstruction incorporating radio-telemetered cystometry

To present a novel porcine model of bladder outlet obstruction (BOO) with a standardized bladder outlet resistance and real-time ambulatory radio-telemetered cystometry, as BOO is a common condition with many causes in both adults and children, with significant morbidity and occasional mortality, but attempts to model this condition in many animal models have the fundamental problem of standardising the degree of outlet resistance. BOO was created in nine castrated male pigs by dividing the mid-urethra; outflow was allowed through an implanted bladder drainage catheter containing a resistance valve, allowing urine to flow across the valve only when a set pressure differential was generated across the valve. An implantable radio-telemetered pressure sensor monitored the pressure within the bladder and abdominal cavity, and relayed this information to a remote computer. Four control pigs had an occluded bladder drainage catheter and pressure sensor placed, but were allowed to void normally through the native urethra. Intra-vesical pressure was monitored by telemetry, while the resistance valve was increased weekly, beginning with 2 cmH2O and ultimately reaching 10 cmH2O. The pigs were assessed using conventional cystometry under anaesthesia before death, and samples conserved in formalin for haematoxylin and eosin staining. The pigs had radio-telemetered cystometry for a median of 26 days. All telemetry implants functioned well for the duration of the experiment, but one pig developed a urethral fistula and was excluded from the study. With BOO the bladder mass index (bladder mass/body mass x 10 000) increased from 9.7 to 20 (P = 0.004), with a significant degree of hypertrophy of the detrusor smooth muscle bundles. Obstructed bladders were significantly less compliant than control bladders (8.3 vs 22.1 mL/cmH2O, P = 0.03). Telemetric cystometry showed that there was no statistically significance difference in mean bladder pressure between obstructed and control pigs (4.8 vs 6.7 cmH2O, P = 0.7), but that each void was longer in the pigs with BOO. This new model of BOO provides a method of reliably and precisely defining the bladder outlet resistance; it induces the changes classically seen with BOO, including increased bladder mass, increased smooth muscle bundle size and decreased compliance.

A MALE MURINE MODEL OF PARTIAL BLADDER OUTLET OBSTRUCTION REVEALS CHANGES IN DETRUSOR MORPHOLOGY, CONTRACTILITY AND MYOSIN ISOFORM EXPRESSION

Mice with gene deletion or targeted over expression are important for understanding the remodeling that follows partial bladder outlet obstruction (PBOO). This condition predominates in males. We produced PBOO in male mice and now report the physiological, histological and molecular consequences. Male C57bl/6 mice were surgically obstructed or subjected to sham surgery and unoperated mice served as controls. Four weeks later the bladders were excised and their function was assessed with in vitro whole bladder cystometry. The optimum volume for pressure generation was determined and isometric pressures were measured for field stimulation and depolarization with KCl. Bladder hypertrophy was classified as severe-bladder mass greater than 50 mg or mild-bladder mass less than 50 mg. The percent muscle fraction was determined by histological analyses. The expression of C-terminal (SM1 and SM2) and N-terminal (SM-B and SM-A) isoforms of myosin heavy chain was analyzed by reverse transcriptase-polymerase chain reaction. Severely hypertrophied bladders had larger optimum volume (p >0.001) and generated less pressure in response to field stimulation (p >0.001) and KCl (p >0.01) with a slower rate of pressure generation than controls or sham operated mice. Increased SM1-to-SM2 and SM-A-to-SM-B ratios were noted in severely obstructed bladders relative to controls or sham operated mice (p <0.05). The muscle fraction decreased slightly in the severely hypertrophied group (p not significant). Our male mouse model of PBOO demonstrates an increase in bladder mass, larger capacity and significantly decreased pressure generation in the in vitro whole bladder model. Obstruction induced increases in the expression of C-terminal (SM1) and N-terminal (SM-A) myosin heavy chain isoforms.

Effect of timing of diuresis on response of rat bladder to outlet obstruction

Objectives To determine whether results similar to a previous study, in which sucrose-induced diuresis started before creating partial outlet obstruction resulted in protection of bladder contractile function in obstructed animals, would be obtained if diuresis were begun after creation of partial outlet obstruction. Methods Thirty-six rats were separated into six groups of 6 rats each: group 1, control; group 2, diuresis; group 3, obstructed; group 4, obstructed immediately after diuresis; group 5, obstructed with diuresis beginning 10 days after obstruction; and group 6, obstructed before diuresis. Diuresis was induced by feeding 5% sucrose instead of water. After 3 weeks of obstruction, all bladders were excised, cut into longitudinal strips, and mounted in isolated muscle baths for physiologic study. Results Sucrose-induced diuresis resulted in a statistically significant increase in bladder mass and a mild increase in smooth muscle contractility compared with the control water-fed group. Obstruction resulted in a fourfold increase in bladder weight and substantial decreases in the responses to all forms of stimulation. The bladder weights of the obstruction plus diuresed groups were all similar to the diuresis-alone group and significantly lower than those of the obstructed group. All diuresed groups showed protection of the contractile responses compared with the obstruction-alone group. Unexpectedly, the group that began diuresis 1 day after obstruction showed the strongest protection. Conclusions Diuresis significantly reduced the increase in bladder mass induced by obstruction and also reduced the level of contractile dysfunction. Diuresis begun 1 day after obstruction resulted in the greatest level of protection.

An essential role of Cav1.2 L-type calcium channel for urinary bladder function.

Mice deficient in the smooth muscle Cav1.2 calcium channel (SMACKO, smooth muscle alpha1c-subunit calcium channel knockout) have a severely reduced micturition and an increased bladder mass. L-type calcium current, protein, and spontaneous contractile activity were absent in the bladder of SMACKO mice. K+ and carbachol (CCh)-induced contractions were reduced to 10-fold in detrusor muscles from SMACKO mice. The dihydropyridine isradipine inhibited K+- and CCh-induced contractions of muscles from CTR but had no effect in muscles from SMACKO mice. CCh-induced contraction was blocked by removing extracellular Ca2+ but was unaffected by the PLC inhibitor U73122 or depletion of intracellular Ca2+ stores by thapsigargin. In muscles from CTR and SMACKO mice, CCh-induced contraction was partially inhibited by the Rho-kinase inhibitor Y27632. These results show that the Cav1.2 Ca2+ channel is essential for normal bladder function. The Rho-kinase and Ca2+-release pathways cannot compensate the lack of the L-type Ca2+ channel.