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Conference Coverage AUA 2004 - Selected Abstracts AUA 2004 - Non-neurogenic and Neurogenic Voiding Dysfunction Selected Abstracts
Conference Coverage AUA 2004 - Selected Abstracts AUA 2004 - Non-neurogenic and Neurogenic Voiding Dysfunction Selected Abstracts |
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| AUA 2004 - Non-neurogenic and Neurogenic Voiding Dysfunction Selected Abstracts |
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| Sunday, 09 May 2004 | ||
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ATP RELEASE FROM HUMAN BLADDERS WITH SENSORY DISORDER VS NORMAL BLADDERS Sunday, May 09, 2004 Vivekanandan Kumar*, Christopher R Chapple, Russell Chess-Williams, Sheffield, United Kingdom. INTRODUCTION AND OBJECTIVE: Studies have suggested that the purinergic system is important not only in normal bladder function but may also contribute to functional disorders of the bladder. In detrusor from patients with sensory urgency the purinergic activity may be significantly raised. This study aims at establishing the level of ATP release by the urothelium from sensory bladder and compare it with that from the normal human urinary bladder. METHODS: Cold cup biopsies of urothelium, obtained from patients with sensory urgency undergoing cystodistension following full informed consent and approval from local ethics committee, were subjected to stretch (by 50% of the original length). A luciferase assay was used to quantify ATP release. Control bladders were obtained from patients with urodynamically proven stable bladders undergoing surgery for stress urinary incontinence. The neurotoxin tetrodotoxin was used to block the neuronal source of ATP release. RESULTS: There was a significantly greater release of ATP following mechanical stretch of the urothelium from sensory bladders compared with control bladder. The increase in ATP release was statistically significant (P<0.01) whether expressed in absolute values (3791.4±667.9 pmol/gm of tissue compared with 77.6±16.2 pmol/gm of tissue for controls) or as the increase over basal levels (179.8±38.8 % increase above baseline compared with 37.2±6.3 % over basal levels for control tissues). The non-neuronal source of ATP release was 9 % for sensory bladder and 18 % for control bladders. CONCLUSIONS: There is a significantly increased level of ATP release from urothelium of sensory bladders in comparison to normal bladder suggesting an important role for ATP in this condition. This study provides direct evidence for the increased purinergic activity in sensory disorders of bladder. Further studies are under way to determine the distribution of receptors and receptor density in bladder from patients with sensory urgency. CYCLOPENTENONE PROSTAGLANDIN, 15-DEOXY-&DELTA_UPPER;12,14-PGJ2, ATTENUATES THE DEVELOPMENT OF CYCLOPHOSPHAMIDE INDUCED CYSTITIS IN THE RAT Sunday, May 09, 2004 Hitoshi Masuda*, Pittsburgh, PA; Kazunori Kihara, Tokyo, Japan; Michael B Chancellor, Naoki Yoshimura, Pittsburgh, PA. INTRODUCTION AND OBJECTIVE: Nitric oxide (NO) produced by inducible NOS (iNOS) has been reported to play an important role in the pathogenesis of cyclophosphamide (CYP)-induced cyctitis. A cyclopentenone prostaglandin (PG), 15-deoxy-Δ12,14-PGJ2 (15d-PGJ2), has also been reported to have an anti-inflammatory effect and attenuate iNOS expression in colitis in the rat. Therefore, we have investigated whether 15d-PGJ2 can attenuate cyclophosphamide-induced cystitis. METHODS: Adult male rats received a single intraperitoneal injection of CYP (100mg/kg) and cystitis were evaluated 48 hours later by biochemical and histological studies. In a separate group of animals, 15d-PGJ2 (80 μg/kg, i.p. bolus 10min before and 24 hours after CYP injection) or a selective iNOS inhibitor, 1400W (10mg/kg, i.p. bolus before and 12, 24 hours after CYP injection) were administered. Animals were then sacrificed and tissues were removed for evaluation for cystitis 48 hours after CYP injection. RESULTS: 15d-PGJ2 as well as 1400W significantly reduced: (1) plasma protein extravasation (Evans blue dye method), (2) the rise in iNOS enzymatic activity, (3) increase of NOX in urine and (4) increase in the myeloperoxidase (MPO) activity in CYP-treated rats. Furthermore, both agents reduced CYP-induced edema and hemorrhage as well as increases in immunohistochemical staining for iNOS in the bladder. CONCLUSIONS: These results indicate that suppression of iNOS induction by 15d-PGJ2 as well as iNOS inhibitor can attenuate the development of CYP-induced cystitis. Thus the treatment with cyclopentenone prostaglandins such as 15d-PGJ2 may be effective for CYP-induced cystitis. CENTRAL MUSCARINIC RECEPTOR SUBTYPES REGULATING VOIDING IN RATS Sunday, May 09, 2004 Masanori Kono*, Yoshiyuki Ishiura, Koichi Kodama, Kazuto Komatsu, Mikio Namiki, Kanazawa, Japan; Yasuo Nakamura, Akita, Japan; Hitoshi Kontani, Kanazawa, Japan. INTRODUCTION AND OBJECTIVE: Muscarinic receptors are classified into five subtypes, M1 to M5. We previously reported that intracerebroventricular (i.c.v.) injection of oxotremorine methiodide (Oxo-M), a non-selective muscarinic agonist increased bladder capacity (BC) and maximal voiding pressure (MVP), and atropine, a non-selective muscarinic antagonist increased BC and decreased MVP in rats. The present study revealed that roles muscarinic receptor subtypes on the central regulation of micturition. METHODS: In female Sprague-Dawley rats (210-280 gr.) anesthetized with halothane, an intravesical catheter was inserted through the bladder dome and an i.c.v. cannula was implanted. After the surgery, cystometry was performed in conscious restrained rats by infusing saline into the bladder at a constant rate (0.1 ml/min.). Pirenzepine, a M1 muscarinic antagonist, methoctramine, a M2 antagonist, p-fluorohexahydro-sila-difenidol hydrochloride (pFHHSiD), a M3 antagonist, and muscarinic toxin-3 (MT-3), a M4 antagonist were injected intracerebroventricularly. RESULTS: Pirenzepine (500 micro gr.) decreased MVP from 44.1 +- 3.90 cmH2O to 30.6 +- 1.71 cmH2O (p<0.01), methoctramine (10 micro gr.) decreased BC from 0.44 +- 0.06 ml to 0.28 +- 0.04 ml (p<0.01) and incresed MVP from 40.1 +- 4.13 cmH2O to 44.1 +- 4.50 cmH2O (p<0.05), pFHHSiD (10 micro gr.) decreased MVP from 50.9 +- 7.14 cmH2O to 42.0 +- 6.24 cmH2O (p<0.05), and MT-3 (1000 micro gr.) decreased BC from 0.58 +- 0.08 ml to 0.43 +- 0.05 ml (p<0.01). CONCLUSIONS: Our results indicate that M1 and M3 muscarinic receptors have a role of excitatory mechanisms, and M2 and M4 muscarinic receptors have inhibitory mechanisms on micturition of rats in the central nerve systems. Y-27362, A SPECIFIC INHIBITOR OF RHO-KINASE, IMPROVES BLADDER FUNCTION IN A RAT MODEL OF HYPERACTIVITY. Sunday, May 09, 2004 Mahadevan Rajasekaran*, Steven Kuntz, Nathan Wilkes, Michael Albo, San Diego, CA. INTRODUCTION AND OBJECTIVE: Molecular mechanisms of hyperactive bladder are not clear. Upregulation of the calcium sensitizing Rho-kinase pathway may be a possible mechanism for increased bladder contractility. In the present study, we measured urodynamic changes in an animal model of overactive bladder (OAB) and evaluated the ability of a specific Rho-kinase inhibitor (Y-27632) to suppress bladder hyperactivity. METHODS: Adult male rats (n = 6) were anesthetized and the carotid artery cannulated for blood pressure monitoring. The bladder was exteriorized and a saline-filled Intracath was fixed into the bladder dome with a purse-string suture. The Intracath was connected to both a pressure transducer and an infusion pump. The bladder was then emptied and allowed to equilibrate at zero volume and pressure. Systemic and bladder pressure data was recorded. Continuous filling cystometrograms (CMGs) were performed by infusing warm saline (37°C) to obtain baseline data on each rat. The threshold pressure (TP) at which micturition began was noted as well as the peak pressure (PP) during micturition. The numbers of voids and of non-voiding contractions (NVCs) per unit time were recorded. In order to create bladder hyperactivity, protamine sulfate (30 mg/ml) followed by KCl (500 mM) was infused intravesically at a rate of 0.04 ml/min. CMG was agin recorded. Finally, 100nmoles of Y-27632 was administered intra-arterially to each rat . The bladder was allowed to equilibrate for 10 min after Y-27632 injection. An additional CMG was performed with KCl (500mM) to observe the effects of Rho-kinase inhibition (as affected by the administration of Y-27632) on bladder contractility. RESULTS: Intravesical KCl infusion after protamine exposure resulted in a significantly higher NVC frequency, mean TP, and PP compared to intravesical saline alone (P < 0.05). Y-27362 administration significantly decreased the mean TP (23.61 ± 2.14 cmH2O in Y-27362 treated animals versus 32 ± 5.2 cmH2O in untreated controls), NVCs (40 ± 3.5 NVCs in the untreated controls versus 8 ± 1.6 NVCs per min after Y-27362 treatment), voiding frequency (1.10 ± 1.1 in untreated controls versus 0.60 ± 0.80 voids/min after Y-27362 treatment). CONCLUSIONS: The protamine/KCl-induced bladder hyperactivity produces significant and measurable CMG changes. Inhibition of the calcium sensitizing Rho-kinase pathway significantly suppresses the CMG changes seen in this model and raises the possibility of a novel therapeutic option. RESTORATION OF INNERVATED URINARY SPHINCTER FUNCTION USING MUSCLE PRECURSOR CELLS Sunday, May 09, 2004 Rene Yiou*, James J Yoo, Anthony Atala, Boston, MA. INTRODUCTION AND OBJECTIVE: The use of muscle precursor cells (mpcs) has been proposed for the treatment of urinary sphincter insufficiency. However, the availability of these cells and the status of innervation of the regenerating muscle tissue has not been clearly assessed. In this study we explored the possibility of using autologous mpcs for the functional restoration of irreversibly damaged sphincters in a rat model. METHODS: A model of irreversible damage of the striated urethral sphincter by electrocoagulation was established in rats. Mpcs, isolated from limb muscle fibers, were grown and expanded in culture. Autologous mpcs, infected with an adenovirus carrying the transgene encoding β-galactosidase, were injected in the damaged striated urinary sphincter muscle 37 days after injury creation. RESULTS: Treatment with the autologous mpcs demonstrated the formation of new myotubes during the entire study period. The myotubes possessed acetylcholine receptors, which were interconnected to nerve endings. The injured sphincters that did not receive mpcs resulted in atrophy and fibrosis, and failed to show any evidence of muscle or nerve regeneration. Urodynamic studies of the mpcs treated sphincters resulted in the restoration of sphincter function (41%) by 1 month after injection. In contrast, the sphincters without treatment failed to sustain any bladder pressure. CONCLUSIONS: The precursor cells with myogenic properties, obtained from muscle fibers, are able to form new myotubes within the irreversibly damaged sphincters. The mature myotubes recruit and interconnect with local nerve endings, resulting in the restoration of functional motor units. This study demonstrates the possibility of using mpcs for the functional improvement of irreversible urinary sphincter muscle insufficiency. NITRIC OXIDE SYNTHASE AND CYCLOOXYGENASE INTERACTION IN THE ISCHEMIC BLADDER IN THE RABBIT Sunday, May 09, 2004 Hitoshi Masuda*, Pittsburgh, PA; Keizo Kawano, Masataka Yano, Yasuyuki Sakai, Kazunori Kihara, Tokyo, Japan; Naoki Yoshimura, Pittsburgh, PA; Hiroshi Azuma, Tokyo, Japan. INTRODUCTION AND OBJECTIVE: Interactions between nitric oxide (NO) produced by inducible NOS and prostaglandins (PG) produced by cyclooxygenase-2 (COX-2) have been reported in cerebral and cardiac ischemia. In this study, we investigated the interaction between nitric oxide synthase (NOS) and COX for ischemia-reperfusion injury in the rabbit detrusor. METHODS: Rabbit bladders were exposed to 60 min of ischemia by clamping the major artery entering the bladder in both sides and subsequent 36 hours of reperfusion (I/R) with or without intraperitonal (i.p.) injection of NOS inhibitors. The I/R group was compared with the sham group. The detrusor tissues were processed for isometric tension experiments and measurements of enzymatic NOS activity and production of NOx, cyclic guanosine monophosphate (cGMP) and PGE2. RESULTS: Inducible NOS activity in the detrusor significantly increased following I/R compared to sham. Also, NOx, cGMP and PGE2 productions in the detrusor significantly increased after I/R. Both NG-nitro-L-arginine methylester (L-NAME, 20mg/kg, i.p.) and 1400W (20mg/kg, i.p.) as a selective iNOS inhibitor given 28 hours after ischemia, significantly reduced not only NOx and cGMP but also PGE2 productions in the tissues. In the tension experiment, 3mM L-arginine as a NOS substrate induced detrusor contraction and increased the PGE2 contents in the medium only in the I/R group, which were inhibited by pretreatment with 1μM indomethacin as a COX inhibitor. The selective soluble guanylate cyclase inhibitor, ODQ, did not affect the PGE2 production and L-arginine induced contraction. CONCLUSIONS: These results demonstrate that NO caused by an up-regulation of iNOS increases PG biosynthesis through a cyclic GMP-independent mechanism and suggest that the interaction between NOS and COX pathways may represent an important mechanism for the modulation of the bladder function in the ischemic state. THE EFFECT OF NITRIC OXIDE ON THE RESTING TONE AND THE CONTRACTILE BEHAVIOUR OF THE EXTERNAL URETHRAL SPHINCTER – A FUNCTIONAL URODYNAMIC STUDY IN HEALTHY HUMAN Sunday, May 09, 2004 Andre Reitz*, Sibylle Bretscher, Peter Knapp, Bjoern Wefer, Michael Muentener, Brigitte Schurch, Zurich, Switzerland. INTRODUCTION AND OBJECTIVE: This functional urodynamic study assessed the effect of the nitric oxide donor isosorbide dinitrate on the external urethral sphincter and hypothesised first that the nitric oxide could lower the resting sphincter pressure, second that nitric oxide could influence the sphincter contractility during magnetic stimulation, and third that nitric oxide could induce a faster external urethral sphincter fatigue during continuous magnetic stimulation. METHODS: 8 healthy males gave their written informed consent and were included. A 2-channel microtip pressure transducer catheter was inserted into the urethra measuring the bladder and the external urethral sphincter pressure. Magnetic stimulation of the sacral roots was performed to evoke reproducible contractions of the external urethral sphincter. The baseline protocol included six single pulse stimulations and three stimulations 10 sec in duration each for two frequencies 5 Hz and 50 Hz with intensities above the motor threshold of the pelvic floor. Then the subjects received 10 mg of isosorbide nitrate sublingually and the protocol was repeated 5 min, 20 min, 40 min and 60 min after drug administration. RESULTS: The sublingual administration of isosorbide dinitrate significantly reduced the resting pressure of the external urethral sphincter at least for one hour (p<0.0001). The maximal contractile strength measured as the mean and maximal urethral pressures during single pulse (p<0.001) and continuous (p<0.05) magnetic stimulation of the sacral roots decreased also significantly. Nitric oxide did not induce a significantly faster fatigue of the external urethral sphincter during continuous magnetic stimulation of the sacral roots. CONCLUSIONS: This study shows a functionally relevant effect of nitric oxide on the resting tone and the contractile behaviour of the human external urethral sphincter in vivo. Nitric oxide donors could offer a new pharmacological approach to treat urinary retention due to overactive or non-relaxing external urethral sphincter. EFFECTS OF AGE AND STRETCH ON ATP RELEASE FROM ISOLATED HUMAN BLADDER Sunday, May 09, 2004 Masaki Yoshida*, Akito Inadome, Koichi Masunaga, Masayuki Otani, Koichi Miyamae, Hitoshi Iwashita, Shoichi Ueda, Kumamoto, Japan. INTRODUCTION AND OBJECTIVE: Non-adrenergic and non-cholinergic (NANC) neurotransmitters may contribute to contractions in human bladder with pathological conditions or aged bladder. It has been reported that adenosine triphosphate (ATP) is the most important NANC neurotransmitter in bladder contraction. Furthermore, several reports suggest that urothelium releases ATP, which may be one of the stimulating factors for afferent nerves resulting to increase in micturition reflex. The present study was designed to evaluate the effects of age and bladder stretch on ATP release in isolated human bladder strips with or without urothelium. METHODS: Human bladders were obtained from 20 patients (average 65.8 years old). Bladder strips with or without urothelium were suspended in organ bath filled with Krebs-Henseleit solution, and tension developments were recorded. Microdialysis probe was inserted into the strip, and Ringer solution was perfused into the probe. The effects of age and the elevation of the resting tension (0 to 4 g) induced by strip stretch on non-neuronal ATP releases were evaluated. Furthermore, the dialysate during electrical field stimulation (EFS: 0.5 msec duration, 60 Hz and 3 sec train at 1 min interval for 10 min) was collected, and the effect of age on EFS-induced ATP releases (neuronal ATP) is also evaluated. The amount of ATP released in the dialysate was measured by luciferine-lusiferase assay. RESULTS: In human bladder trips before EFS, there were non-neuronal ATP releases, which were not inhibited by treatment with tetrodotoxin. The non-neuronal ACh releases from bladder strips with urothelium were significantly higher than that from strips without urothelium. The non-neuronal ATP releases from bladder strips with urothelium were increased with age. The non-neuronal ATP in older (over 65 years) patients was significantly higher than that in younger patients (under 65 years). In the strips of older patients, stretch of the strips caused significant increase in non-neuronal ATP release. EFS caused significant increases in neuronal ATP releases in both bladder strips with and without urothelium. Neuronal ATP release was increased with age, and there was a significant positive correlation between age and the release. CONCLUSIONS: The data demonstrate that there are age-related increases in neuronal and non-neuronal ATP releases in human bladder. It is suggested that bladder urothelium may play an important role on non-neuronal ATP release, and that bladder stretch may contribute to increase in ATP release from urothelium in the elderly. PROTECTIVE EFFECTS OF ARGINASE INHIBITOR ON THE ENDOTHELIN-1 INDUCED DETRUSOR OVERACTIVITY IN RATS Sunday, May 09, 2004 Hitoshi Masuda*, Jang H Kim, Teruyuki Ogawa, Pittsburgh, PA; Kazunori Kihara, Tokyo, Japan; William C De Groat, Michael B Chancellor, Sidney Morris, Naoki Yoshimura, Pittsburgh, PA. INTRODUCTION AND OBJECTIVE: It has been reported that nitric oxide (NO) in the bladder can suppress bladder afferent activity to suppress bladder overactivity and nociceptive responses. NO production can be modulated by inhibition of an enzyme, arginase, that shares a common substrate, L-arginine. Therefore, we investigated the effects of arginase inhibition on endothelin-1 (ET-1) induced detrusor overactivity in rats. METHODS: Continuous cystometrogram (CMG) was performed in adult female Sprague-Dawley rats under urethane anesthesia. The effects of intraarterial injection of an arginase inhibitor, N-ω-Hydroxy-nor-L-arginine (nor-NOHA, 3~12mg/kg), above the aortic bifurcation were examined on detrusor overactivity induced by intravesical injection of endothelin-1 (10μM). Pretreatments with intraarterial injection of NG-nitro-L-arginine methylester (L-NAME, 10mg/kg) as a NOS inhibitor, and L-arginine (100mg/kg) as a NOS substrate were also performed to investigate whether the effects of nor-NOHA were mediated via the NO system. RESULTS: Intravesical injection of ET-1(0.1-10μM) induced detrusor overactivity evidenced by a reduction in the intercontraction interval (ICI) in a dose-dependent manner (10μM ET-1, P<0.01, n=7, 35.6±4.7% of the control ICI), which was suppressed by capsaicin pretreatment (125mg/kg, subcutaneously, 4days before). Intraarterial injection of nor-NOHA suppressed 10μM ET-1 induced detrusor overractivity in a dose-dependent manner (P<0.05, 12mg/kg nor-NOHA with 10μM ET-1; 68.2±4.2% of the control ICI vs ICI at 10μM ET-1 only). Moreover, the inhibitory effect of nor-NOHA was abolished by intraarterial injection of L-NAME pretreatment.Intraarterial injection of L-arginine also significantly inhibited the ET-1 induced bladder overactivity. CONCLUSIONS: These results indicate that the blockade of arginase activity can enhance NO production by increasing L-arginine available to NOS, and thereby suppress C-fiber mediated bladder overactivity induced by ET-1. Thus, arginase inhibition may be effective for treating bladder overactivity and/or painful conditions.
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