Alteration in TRPV1 and Muscarinic (M3) receptor expression and function in idiopathic overactive bladder urothelial cells.

AIM: To examine function of both cholinergic (muscarinic) and TRPV1 receptors in human bladder urothelial (HBUC) from non-neurogenic overactive bladder (OAB) patients as compared to control subjects. METHODS: Primary HBUC cultures were derived from cystoscopic biopsies from OAB and control subjects. Muscarinic and TRPV1 function was assessed by acetylcholine (5 µm) or capsaicin (0.5 µm) evoked ATP release, measured by luciferase assay. Overall, expression of TRPV1 and muscarinic M3 receptors in bladder urothelial cells was accomplished using western immunoblotting. RESULTS: Our findings revealed that the response to acetylcholine in OAB HBUC cultures (which was blocked by the nonselective muscarinic antagonist, atropine methyl nitrate or AMN) was not significantly different than from controls. The acetylcholine M3 receptor was slightly decreased as compared to control. In contrast, OAB HBUC cultures exhibited a capsaicin hypersensitivity and augmented release of ATP (3.2 fold higher), which was blocked by the antagonist capsazepine. The increase in capsaicin sensitivity correlated with increased urothelial TRPV1 expression. CONCLUSION: Though characterized in a small number of subjects, augmented release of urothelial-derived transmitters such as ATP could 'amplify' signalling between and within urothelial cells and nearby afferent nerves.

Randomized multicenter clinical trial of myofascial physical therapy in women with interstitial cystitis/painful bladder syndrome and pelvic floor tenderness.

PURPOSE: We determined the efficacy and safety of pelvic floor myofascial physical therapy compared to global therapeutic massage in women with newly symptomatic interstitial cystitis/painful bladder syndrome. MATERIALS AND METHODS: A randomized controlled trial of 10 scheduled treatments of myofascial physical therapy vs global therapeutic massage was performed at 11 clinical centers in North America. We recruited women with interstitial cystitis/painful bladder syndrome with demonstrable pelvic floor tenderness on physical examination and a limitation of no more than 3 years' symptom duration. The primary outcome was the proportion of responders defined as moderately improved or markedly improved in overall symptoms compared to baseline on a 7-point global response assessment scale. Secondary outcomes included ratings for pain, urgency and frequency, the O'Leary-Sant IC Symptom and Problem Index, and reports of adverse events. We compared response rates between treatment arms using the exact conditional version of the Mantel-Haenszel test to control for clustering by clinical center. For secondary efficacy outcomes cross-sectional descriptive statistics and changes from baseline were calculated. RESULTS: A total of 81 women randomized to the 2 treatment groups had similar symptoms at baseline. The global response assessment response rate was 26% in the global therapeutic massage group and 59% in the myofascial physical therapy group (p=0.0012). Pain, urgency and frequency ratings, and O'Leary-Sant IC Symptom and Problem Index decreased in both groups during followup, and were not significantly different between the groups. Pain was the most common adverse event, occurring at similar rates in both groups. No serious adverse events were reported. CONCLUSIONS: A significantly higher proportion of women with interstitial cystitis/painful bladder syndrome responded to treatment with myofascial physical therapy than to global therapeutic massage. Myofascial physical therapy may be a beneficial therapy in women with this syndrome.

5-year continence rates, satisfaction and adverse events of burch urethropexy and fascial sling surgery for urinary incontinence.

PURPOSE: We characterized continence, satisfaction and adverse events in women at least 5 years after Burch urethropexy or fascial sling with longitudinal followup of randomized clinical trial participants. MATERIALS AND METHODS: Of 655 women who participated in a randomized surgical trial comparing the efficacy of the Burch and sling treatments 482 (73.6%) enrolled in this long-term observational study. Urinary continence status was assessed yearly for a minimum of 5 years postoperatively. Continence was defined as no urinary leakage on a 3-day voiding diary, and no self-reported stress incontinence symptoms and no stress incontinence surgical re-treatment. RESULTS: Incontinent participants were more likely to enroll in the followup study than continent patients (85.5% vs 52.2%) regardless of surgical group (p<0.0001). Overall the continence rates were lower in the Burch urethropexy group than in the fascial sling group (p=0.002). The continence rates at 5 years were 24.1% (95% CI 18.5 to 29.7) vs 30.8% (95% CI 24.7 to 36.9), respectively. Satisfaction at 5 years was related to continence status and was higher in women undergoing sling surgery (83% vs 73%, p=0.04). Satisfaction decreased with time (p=0.001) and remained higher in the sling group (p=0.03). The 2 groups had similar adverse event rates (Burch 10% vs sling 9%) and similar numbers of participants with adverse events (Burch 23 vs sling 22). CONCLUSIONS: Continence rates in both groups decreased substantially during 5 years, yet most women reported satisfaction with their continence status. Satisfaction was higher in continent women and in those who underwent fascial sling surgery, despite the voiding dysfunction associated with this procedure.

Effects of dimethyl sulphoxide and heparin on stretch-activated ATP release by bladder urothelial cells from patients with interstitial cystitis.

OBJECTIVE: To determine whether dimethyl sulphoxide (DMSO) and heparin reduce the greater stretch-activated ATP release in interstitial cystitis (IC), as ATP serves as a nocio-neurotransmitter in the bladder, and thus explain their beneficial effects in patients with IC (a disease characterized by hypersensory bladder symptoms). MATERIALS AND METHODS: Bladder epithelia in IC release more ATP in response to stretch than do control samples. Both DMSO and heparin are used intravesically to treat IC; such agents can modulate urothelial function because they directly contact bladder urothelium. Biopsies taken from patients with IC and from control subjects were grown in primary cultures using established cell-culture techniques. Cultured urothelial cells were stretched with the Flexcell device (Flexcell International Corp., McKeesport, PA, USA) and supernatant ATP was measured, using a luciferin-luciferase assay. DMSO (0.1%, 0.5% and 1%) or heparin (50, 200, 800 and 1600 U) was added to the cells at the start of the stretch experiments and the ATP released into the supernatant measured. Cell viability was also determined using Trypan Blue staining. RESULTS: IC cells released significantly more ATP in response to stretch than did control cells. This increased release of ATP by stretched IC cells was significantly blocked by adding DMSO or heparin at all concentrations used. Heparin appeared to have a greater dose-dependent effect on ATP release than did DMSO. CONCLUSIONS: These findings are consistent with the hypothesis that the urothelium provides sensory input via ATP release and that this process is increased in IC. Furthermore, stretch-activated ATP release was blocked by adding DMSO and heparin, both intravesical agents commonly used to treat the symptoms of IC. This study supports the notion that purinergic-targeted therapy is warranted in treating IC. Further studies are needed to determine the mechanisms of increased ATP release by IC urothelial cells.

Modified techniques of S3 foramen localization and lead implantation in S3 neuromodulation.

INTRODUCTION: We describe a reproducible and less invasive surgical approach to sacral neuromodulation (InterStim Therapy) in the treatment of voiding dysfunction. Twenty patients underwent modified lead implantation (mean operative time 45 minutes) without any difficulties or complications, with a mean follow-up of 8 months (range 1 to 14).Technical Considerations. The highlights of these modifications include (a) fluoroscopy to localize the S3 foramen; (b) paramedian incision; (c) use of a cutoff S3 finder needle and a 14-gauge Angiocath to direct permanent lead into the S3 foramen without dissection; (d) use of lateral fluoroscopy to determine the depth of the Angiocath insertion; and (e) anchoring the lead to the lumbodorsal fascia (superficial to the sacral periosteum) using a moveable lead anchor system. These modifications simplify and minimize the invasiveness of this therapy without compromising the efficacy. CONCLUSIONS: Because of the simplicity of these modifications, we are currently using an implanted lead, rather than the temporary percutaneous lead, to assess patients' clinical response before implanting a pulse generator.

Augmented stretch activated adenosine triphosphate release from bladder uroepithelial cells in patients with interstitial cystitis.

PURPOSE: Extracellular adenosine triphosphate (ATP) has been shown to mediate inflammation and nociception and, therefore, it may have a role in symptoms associated with interstitial cystitis. We theorized that the bladder uroepithelium releases ATP in response to stretch and, furthermore, this process is augmented in interstitial cystitis. MATERIALS AND METHODS: We quantitated ATP using the luciferin-luciferase assay. Urinary ATP levels were compared in 35 patients with interstitial cystitis and in 33 normal controls after pH correction. Cultured interstitial cystitis and normal urothelial cells from the bladder biopsies of 5 patients each were stretched with the Flexcell 2000 machine (Flexcell International Corp., McKeesport, Pennsylvania) and supernatant ATP concentrations were measured. RESULTS: Mean urinary ATP plus or minus standard error of mean was significantly higher in patients with interstitial cystitis than in controls (L value 985 +/- 161 versus 377 +/- 27, p = 0.0007). Supernatant ATP released by stretched interstitial cystitis cells was stretch intensity dependent when comparing 0%, 10% and 20% elongation, and was also significantly higher in stretched interstitial cystitis than in stretched normal cells. CONCLUSIONS: Adenosine triphosphate was significantly elevated in the urine of individuals with interstitial cystitis and the stretch activated release of ATP was augmented in interstitial cystitis urothelium. Increased extracellular ATP may have a role in mechanosensory transduction and to our knowledge it represents a novel hypothesis.

Altered neural control of micturition in the aged F344 rat.

The purpose of this study was to determine whether micturition reflexes are altered in aged rats. Voiding frequencies and awake cystometrograms (CMGs) were measured in young (3-5 months old) and aged (24 months) F344 male rats. Bladder contractions induced by subcutaneous apomorphine and intravesical capsaicin stimulation were measured using awake CMGs. Urodynamic parameters were compared. Aged rats voided less frequently (4.1 vs 6.9 times/18 h, P = 0.006), with a higher volume per void (1.1 vs 0.7 ml, P = 0.02) and had a higher micturitional threshold pressure (8.7 vs 4.6 mmHg, P = 0.0001) than the young rats. Apomorphine induced a higher frequency of bladder contractions in aged animals compared to young animals (5.5 vs 3.1 contractions/min, P = 0.03). Intravesical capsaicin caused a lower pressure bladder response in the aged rats (38.5 vs 70.6 mmHg, P = 0.01) compared to the young rats. Bladder afferents and central micturition pathways may be altered in aged rats. Impaired bladder contractility in the elderly may be exacerbated by reduced sensory input, whereas the propensity for detrusor instability could result from altered central processing. This study demonstrated the utility of the F344 animal model to study micturitional changes resulting from aging.

Percutaneous sacral third nerve root neurostimulation improves symptoms and normalizes urinary HB-EGF levels and antiproliferative activity in patients with interstitial cystitis.

OBJECTIVES: A highly effective treatment for interstitial cystitis (IC) remains elusive. We determined whether sacral third nerve root (S3) percutaneous neurostimulation (PNS) might be effective in relieving symptoms of IC, as well as in normalizing urinary factors that are specifically altered in IC. METHODS: Six consecutive patients with symptoms and cystoscopic findings compatible with IC underwent 5 days of continuous S3 neurostimulation by way of leads placed percutaneously into the S3 foramen. Patients filled out voiding frequency diaries and pain and urgency questionnaires before PNS and at the end of PNS when the leads were removed. Urine specimens were collected at these two time points and measured for heparin-binding epidermal growth factor-like growth factor (HB-EGF) by enzyme-linked immunosorbent assay and for antiproliferative factor (APF) activity by (3)H-thymidine uptake by normal human bladder urothelial cells. RESULTS: S3 PNS significantly improved all measured parameters toward normal values. Voiding frequency decreased twofold from 23.1 +/- 4.6 to 10.6 +/- 4.0 voids daily during PNS (P = 0.0001). Pelvic pain on a scale of 1 to 10 decreased from 7.0 +/- 1.6 to 2.3 +/- 3.2 (P = 0.05). Urinary urgency on a scale of 1 to 10 decreased from 6.0 +/- 2.2 to 1.8 +/- 1.7 (P = 0. 02). Urinary HB-EGF concentration increased sevenfold from 1.5 +/- 2. 1 to 11.0 +/- 1.7 ng/mL (P <0.0001), and urinary APF activity decreased from -76.1% +/- 31% to -4.5% +/- 8.8% (P = 0.005). CONCLUSIONS: S3 PNS significantly decreased symptoms and normalized urinary HB-EGF and APF activity in patients with IC. These results suggest that permanent S3 PNS may be beneficial in treating IC.

Pilot study of sequential oral antibiotics for the treatment of interstitial cystitis.

PURPOSE: Interstitial cystitis is a chronic disease of unknown etiology characterized by bladder pain, urgency and frequency. Although a single microbe has not been implicated as a cause of interstitial cystitis, several groups noted various organisms in the urine of some women with interstitial cystitis and some patients reported that antibiotics decrease symptoms. Consequently we performed a prospective, randomized, double-blinded, placebo controlled pilot study of sequential oral antibiotics. MATERIALS AND METHODS: We randomized 50 patients with interstitial cystitis to receive 18 weeks of placebo or antibiotics, including rifampin plus a sequence of doxycycline, erythromycin, metronidazole, clindamycin, amoxicillin and ciprofloxacin for 3 weeks each. RESULTS: Intent to treat analysis demonstrated that 12 of 25 patients (48%) in the antibiotic and 6 of 25 (24%) in the placebo group reported overall improvement (p = 0.14), while 10 and 5, respectively, noticed improvement in pain and urgency (p = 0.22). In the antibiotic group 20 participants (80%) had adverse effects compared with 10 (40%) in the placebo group (p = 0.009). CONCLUSIONS: Our findings suggest that these antibiotics alone or in combination may sometimes be associated with decreased symptoms in some patients but they do not represent a major advance in therapy for interstitial cystitis.

Bladder stretch alters urinary heparin-binding epidermal growth factor and antiproliferative factor in patients with interstitial cystitis.

PURPOSE: The etiology of interstitial cystitis is unknown. Urine from patients with interstitial cystitis has been shown to inhibit urothelial proliferation through a putative antiproliferative factor and to contain decreased levels of heparin-binding epidermal growth factor-like growth factor (HB-EGF) compared to controls. Stretch of detrusor smooth muscle cells is known to stimulate HB-EGF production. Because bladder hydrodistention sometimes alleviates the symptoms of interstitial cystitis, we determined whether the stretch stimulus of hydrodistention alters antiproliferative factor activity and/or HB-EGF in interstitial cystitis urine specimens. MATERIALS AND METHODS: Urine was collected immediately before, and 2 to 4 hours and 2 weeks after hydrodistention from 15 patients with symptoms and cystoscopic findings compatible with interstitial cystitis and 13 controls. Hydrodistention was performed with the subject under general or regional anesthesia and bladders were distended to 80 cm. water 3 times. Urinary HB-EGF was measured by enzyme-linked immunosorbent assay and urinary antiproliferative factor activity was determined by measuring 3H-thymidine uptake by normal human bladder urothelial cells. RESULTS: Hydrodistention significantly increased urinary HB-EGF in patients with interstitial cystitis toward normal control values (before distention p = 0.003, 2 weeks after distention p = 0.67). Urine antiproliferative factor activity decreased significantly after hydrodistention in patients with interstitial cystitis. However, antiproliferative factor activity in interstitial cystitis and control specimens was still statistically different 2 weeks after distention (before distention p = 0.0000004, 2 weeks after distention p = 0.04). CONCLUSIONS: Bladder stretch increased HB-EGF and conversely reduced antiproliferative factor activity in urine from patients with interstitial cystitis but not controls up to 2 weeks after distention. These results provide additional evidence for the possible role of antiproliferative factor and decreased HB-EGF in the pathophysiology of interstitial cystitis. To our knowledge this is also the first human study to show that in vivo bladder stretch can alter urinary factors that regulate cell growth.

Persistently increased voiding frequency despite relief of bladder outlet obstruction.

PURPOSE: Bladder outlet obstruction (BOO) can increase urinary frequency. Even after surgical relief of obstruction, up to 30% of patient are still bothered by irritative voiding symptoms. We tested the hypothesis that deligation of a partial bladder outlet obstruction model mimics this clinical observation. MATERIALS AND METHODS: Female Wistar rats were obstructed for 3 weeks by partial urethral ligation and then were relieved of obstruction by urethral deligation. Measurements of voiding frequency and voided volumes were measured preoperatively, after ligation, and after deligation. Relief of obstruction was confirmed by measuring flow rates through ex vivo perfusion of deligated urethras. Urine osmolality and bladder weights were determined. Awake cystometrograms (CMGs) were performed 3 weeks after deligation to measure bladder function. RESULTS: Neither sham ligation nor sham deligation altered voiding frequency. Ligation doubled mean voiding frequency (in cc) from 2.01 +/- 0.32 to 3.96 +/- 0.22 per 4 hours (p = 0.0002). Three weeks after deligation, voiding behavior of the animals segregated into 2 groups: 20% had persistent hyperactive voiding frequency (6.67 +/- 1.23 per 4 hours) while 80% normalized voiding frequency (1.53 +/- 0.20 per 4 hours). The difference in voiding frequency in these 2 groups could not be attributed to alterations in urine osmolality, persistence of urethral obstruction, difference in bladder weights or severity of initial obstruction created. Awake CMGs revealed a higher peak micturition pressure and lower voided volume in the hyperactive voiders. CONCLUSIONS: 20% of the animals after urethral deligation had persistent hyperactive voiding which parallels clinical observations. Because the CMG data suggested persistent obstruction, yet urethral perfusion and bladder weights indicated no obstruction, we propose that these 20% of animals have a "functional" bladder outlet obstruction and can be used to study mechanisms underlying hyperactive voiding.

Use of the flexible cystoscope as a vaginoscope to aid in the diagnosis of artificial sling erosion.

Two patients with bladder neck suspension by artificial slings presented with complaints of vaginal pain and drainage as well as irritative voiding. Pelvic examination and flexible cystoscopy were negative. Flexible vaginoscopy detected sling erosion in both. Vaginoscopy is a valuable adjunct procedure in detecting this problem.

The incidence of a positive ice water test in bladder outlet obstructed patients: evidence for bladder neural plasticity.

PURPOSE: The ice water test triggers a C fiber, capsaicin sensitive spinal micturition reflex. We postulated that the ice water test is positive in a high proportion of patients with compared to those without bladder outlet obstruction. MATERIALS AND METHODS: Prospective evaluation of 111 consecutive patients was undertaken. Symptoms of urgency, urge incontinence, nocturia and daytime frequency as well as the presence of neurological disease were obtained from history and physical examination. Fluorourodynamics, including ice water cystometry, and pressure-flow studies were done for all 111 subjects. Obstruction was defined using the Abrams-Griffith nomogram and urethral resistive index. A positive ice water test was defined as presence of uninhibited bladder contraction with instillation of 0C saline at 50 cc per minute up to a maximum of 250 cc. Detrusor instability was defined according to the International Continence Society criteria using room temperature saline instillation. RESULTS: When patients with neurological disease were excluded, a positive ice water test was found in 71% of subjects with bladder outlet obstruction (12 of 17), which was significantly higher (p <0.0005, Yates corrected chi-square test) than the 7% positive ice water test rate in nonobstructed subjects (3 of 44). Conversely, the incidence of positive detrusor instability was not statistically different between the patients with or without bladder outlet obstruction. Of the subjects with neurological disease 85% (42 of 50) had a positive ice water test. The incidence of a positive ice water test was only 5 to 9% in patients with storage lower urinary tract symptoms. CONCLUSIONS: A positive ice water test has been previously described in infants and individuals with neurogenic bladders. However, subjects with bladder outlet obstruction had a significantly higher incidence of a positive ice water test compared to those without it, supporting the hypothesis of an enhanced spinal micturition reflex possibly due to plasticity of bladder afferents after bladder outlet obstruction. The ice water test may be useful in prognosticating bladder outlet obstruction treatment outcomes and determining the etiology of treatment failure.

Neurophysiology of micturition and continence in women.

Micturition and continence involve the coordination of complex neural events between the central and peripheral nervous systems. An understanding of these events provides a foundation for the treatment of voiding disorders in women such as stress urinary incontinence, urge incontinence and interstitial cystitis. The purpose of this paper is to comprehensively review the neuroanatomy, neurophysiology and neuropharmacology of micturition and continence. However, a brief section discussing clinical correlations will follow each of these topics to help integrate the basic science with clinical observations.

Neurophysiology of micturition and continence.

This article reviews the neuroanatomy, neurophysiology, and neuropharmacology involved in micturition and continence. Knowledge of these topics helps the clinician diagnose and treat voiding disorders that are caused by disease, trauma, drugs, and aging.