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INTRODUCTION AND OBJECTIVE:Radiation-induced cystitis (RC) is a progressive debilitating disorder and its most prevalent phase is the chronic phase that occurs months and years after radiation expo...

PURPOSE: Transient receptor potential cation channel subfamily M member 4 (TRPM4) has been shown to play a key role in detrusor contractility under physiological conditions. In this study we investigated the potential role of TRPM4 in detrusor overactivity following spinal cord transection (SCT) in mice. METHODS: TRPM4 expression and function were evaluated in bladder tissue with or without the mucosa from spinal intact (SI) and SCT female mice (T8-T9 vertebra; 1–28 days post SCT) using PCR, western blots, immunohistochemistry and muscle strip contractility techniques. RESULTS: TRPM4 was expressed in the urothelium (UT) and detrusor smooth muscle (DSM) and was upregulated after SCT. Expression levels peaked 3–7 days post SCT in both the UT and DSM. Pharmacological block of TRPM4 with the antagonist, 9-Phenanthrol (30 µM) greatly reduced spontaneous phasic activity that developed after SCT, regardless of the presence or absence of the mucosa. CONCLUSIONS: Detrusor overactivity following spinal cord injury leads to incontinence and/or renal impairment, and represents a major health problem for which current treatments are not satisfactory. Augmented TRPM4 expression in the bladder after chronic SCT supports the hypothesis that TRPM4 channels play a role in DSM overactivity following SCT. Inhibition of TRPM4 may be beneficial for improving detrusor overactivity in SCI.

Loss of bladder control is a challenging outcome facing patients with spinal cord injury (SCI). We report that systemic blocking of pro–nerve growth factor (proNGF) signaling through p75 with a CNS-penetrating small-molecule p75 inhibitor resulted in significant improvement in bladder function after SCI in rodents. The usual hyperreflexia was attenuated with normal bladder pressure, and automatic micturition was acquired weeks earlier than in the controls. The improvement was associated with increased excitatory input to the spinal cord, in particular onto the tyrosine hydroxylase–positive fibers in the dorsal commissure. The drug also had an effect on the bladder itself, as the urothelial hyperplasia and detrusor hypertrophy that accompany SCI were largely prevented. Urothelial cell loss that precedes hyperplasia was dependent on p75 in response to urinary proNGF that is detected after SCI in rodents and humans. Surprisingly, death of urothelial cells and the ensuing hyperplastic response were beneficial to functional recovery. Deleting p75 from the urothelium prevented urothelial death, but resulted in reduction in overall voiding efficiency after SCI. These results unveil a dual role of proNGF/p75 signaling in bladder function under pathological conditions with a CNS effect overriding the peripheral one.

Purpose To determine whether responses to serotonin are altered in bladder strips from cats diagnosed with a naturally occurring form of bladder pain syndrome/interstitial cystitis termed feline interstitial cystitis (FIC). Methods Full thickness bladder strips were isolated from aged matched healthy control cats and cats with clinically verified FIC. Bladder strips were mounted in an organ bath and connected to a tension transducer to record contractile activity. A serotonin dose response (0.01-10μM) was determined for each strip with the mucosa intact or denuded. Results Bladder strips from control and FIC cats contracted in response to serotonin in a dose-dependent manner. The normalized force of serotonin-evoked contractions was significantly greater in bladder strips from cats with FIC (n=7) than from control cats (n=4). Removal of the mucosa significantly decreased serotonin-mediated responses in both control and FIC bladder preparations. Furthermore, the contractions in response to serotonin were abolished by 1μM atropine in both control and FIC bladder strips. Conclusion The effect of serotonin on contractile force, but not sensitivity, was potentiated in bladder strips from cats with FIC, and was dependent upon the presence of the mucosa in control and FIC groups. As atropine inhibited these effects of serotonin, we hypothesize that, serotonin enhances acetylcholine release from the mucosa of FIC cat bladder strips, which could account for the increased force generated. In summary, FIC augments the responsiveness of bladder to serotonin, which may contribute to the symptoms associated with this chronic condition.

Aims To present a synopsis of the presentations and discussions from Think Tank I, “Implications for afferent–urothelial bidirectional communication” of the 2014 International Consultation on Incontinence-Research Society (ICI-RS) meeting in Bristol, UK.

Aims: To determine the role of p75 neurotrophin receptor (p75NTR) and the therapeutic effect of the selective small molecule p75NTR modulator, LM11A-31, in spinal cord injury (SCI) induced lower urinary tract dysfunction (LTUD) using a mouse model. Methods: Adult female T8-T9 transected mice were gavaged daily with LM11A-31 (100mg/kg) for up to 6 weeks, starting 1 day before, or 7 days following injury. Mice were evaluated in vivo using urine spot analysis, cystometrograms (CMGs), and external urethral sphincter (EUS) electromyograms (EMGs); and in vitro using histology, immunohistochemistry, and Western blot. Results: Our studies confirm highest expression of p75NTRs in the detrusor layer of the mouse bladder and lamina II region of the dorsal horn of the lumbar-sacral (L6-S1) spinal cord which significantly decreased following SCI. LM11A-31 prevented or ameliorated the detrusor sphincter dyssynergia (DSD) and detrusor overactivity (DO) in SCI mice, significantly improving bladder compliance. Furthermore, LM11A-31 treatment blocked the SCI-related urothelial damage and bladder wall remodeling. Conclusion: Drugs targeting p75NTRs can moderate DSD and DO in SCI mice, may identify pathophysiological mechanisms, and have therapeutic potential in SCI patients.

The isolated bladder shows autonomous micromotions, which increase with bladder distension, generate sensory nerve activity, and are altered in models of urinary dysfunction. Intravesical pressure resulting from autonomous activity putatively reflects three key variables; the extent of micromotion initiation, distances over which micromotions propagate, and overall bladder tone. In vivo, these variables are subordinate to the efferent drive of the central nervous system. In the micturition cycle storage phase, efferent inhibition keeps autonomous activity generally at a low level, where it may signal 'state of fullness', whilst maintaining compliance. In the voiding phase, mass efferent excitation elicits generalised contraction (global motility initiation). In lower urinary tract dysfunction, efferent control of the bladder can be impaired, for example due to peripheral 'patchy' denervation. In this case, loss of efferent inhibition may enable unregulated micromotility, and afferent stimulation, predisposing to urinary urgency. If denervation is relatively slight, the detrimental impact on voiding may be low, as the adjacent innervated areas may be able to initiate micromotility synchronous with the efferent nerve drive, so that even denervated areas can contribute to the voiding contraction. This would become increasingly inefficient the more severe the denervation, such that ability of triggered micromotility to propagate sufficiently to engage the denervated areas in voiding declines, so the voiding contraction increasingly develops the characteristics of underactivity. In summary, reduced peripheral coverage by the dual efferent innervation (inhibitory and excitatory) impairs regulation of micromotility initiation and propagation, potentially allowing emergence of overactive bladder and, with progression, detrusor underactivity.

Introduction Neural stimulation has become an established minimally invasive treatment for various lower urinary tract symptoms. The results both short- and long-term are encouraging, however, there is still a lack of knowledge of obvious risk factors, which may affect the outcome of treatment. Although neural stimulation has been embraced by healthcare professionals and patients, the exact mechanism by which neural stimulation works is still unclear. Discussion A condense review of knowledge available on this topic is presented. Several research questions are raised. Outlines of research studies, both clinical and basic science, are suggested. Conclusions Further studies are necessary to understand mechanism of action of neural stimulation and its implications on treatment outcomes. Neurourol. Urodynam. 33:618–621, 2014. © 2014 Wiley Periodicals, Inc.

Aims To present a brief review on discussions from “Do we understand any more about lower urinary tract interstitial cells?” session at the 2013 International Consultation on Incontinence-Research Society (ICI-RS) meeting in Bristol, UK. Methods Discussion focused on bladder interstitial cell (IC) subtypes, their localization and characterization, and communication between themselves, the urothelium, and detrusor smooth muscle. The role of ICs in bladder pathologies and new methods for studying ICs were also addressed. Results ICs have been studied extensively in the lower urinary tract and have been characterized based on comparisons with ICs of Cajal in the gastro-intestinal tract. In fetal bladders it is believed that ICs drive intrinsic contractions to expel urine through the urachus. These contractions diminish postpartum as bladder innervation develops. Voiding in human neonates occurs when filling triggers a spinal cord reflex that contracts the detrusor; in rodents, maternal stimulation of the perineum triggers voiding. Following spinal cord injury, intrinsic contractions, and spinal micturition reflexes develop, similar to those seen during neonatal development. These enhanced contractions may stimulate nociceptive and mechanosensitive afferents contributing to neurogenic detrusor overactivity and incontinence. The IC-mediated activity is believed to be initiated in the lamina propria by responding to urothelial factors. These IC may act syncytially through gap junction coupling and modulate detrusor activity through unknown mechanisms. Conclusion There has been a great deal of information discovered regarding bladder ICs, however, many of their (patho)physiological functions and mechanisms are still unclear and necessitates further research. Neurourol. Urodynam. 33:573–576, 2014. © 2014 Wiley Periodicals, Inc.

Background Botulinum neurotoxin A (BoNTA), which alleviates overactive bladder symptoms, is thought to act predominantly via the inhibition of transmitter release from parasympathetic nerves. However, actions at other sites such as afferent nerve terminals are possible. Objective To evaluate the effects of BoNTA on bladder afferent neuropeptide release and firing. Design, setting, and participants One side of the bladder of control and chronic (1–2 wk) spinal cord transected (SCT; T 8 -T 9 ) adult female mice was injected with BoNTA (0.5 U/5μl saline). After 48h, bladders with L 6 -S 2 spinal nerves were prepared for in vitro recordings. Outcome measurements and statistical analysis In bladder preparations, tension and optical mapping of Ca 2+ transients were used to measure intrinsic contractions, those evoked by capsaicin or the electrical stimulation of spinal nerves. Afferent firing was evoked by stretch or intrinsic bladder contractions. The numbers of responding units and firing rates were measured. Animal numbers were used to detect moderate to large between-group differences based on Cohen's criteria. Two-way analysis of variance was used to test spatial/temporal differences in Ca 2+ signals as mean plus or minus standard deviation. Differences between data sets were tested with the student t test and skewed data sets with a Mann-Whitney U test (significant when p Results and limitations In control and SCT bladders, BoNTA treatment decreased the contractions evoked by electrical stimulation of spinal nerves without altering intrinsic contractions. Afferent firing on untreated sides in response to stretch/intrinsic contractions was increased in SCTs versus controls. On BoNTA-treated sides, afferent firing rates were greatly attenuated in response to mechanical stimulation as were the capsaicin-evoked optical signals mediated by neuropeptide release. Conclusions SCT caused an increased sensitivity of afferent nerves to mechanical stimulation that was reduced by BoNTA treatment. Increased intrinsic activity after SCT was unaffected by the toxin. Thus BoNTA suppresses neurogenic detrusor overactivity by targeting afferent as well as efferent pathways in the bladder.

Background Botulinum neurotoxins type A (BoNT/A), β3-adrenergic receptor agonists, and phosphodiesterase type 5 (PDE5) inhibitors are promising agents that mitigate lower urinary tract symptoms by attenuating the sensory system. However, whether they act directly on afferent nerves or indirectly through the other cell types is unclear. Methods Spinal cord transected female mice were used as a model for neurogenic bladder overactivity. In vivo methods utilized decerebrate mouse cystometry. In vitro approaches included optical mapping of Ca2+ transient, single unit afferent nerve recordings and tension measurements from bladder sheets and wall cross-sections. Immunohistochemistry was used to measure the expression of β3-adrenergic receptors on dorsal root ganglion (DRG) neurons. Results Our unique approaches revealed the direct effects of BoNT/A in inhibiting neuropeptide release and firing rates in afferents following bladder injections. β3-adrenergic receptor agonists are demonstrated to directly inhibit afferent nerve firing independent of the relaxing effects on bladder smooth muscle. Moreover, data suggest the expression of these receptors on DRG neurons that send projections to the bladder. The mechanism of action of PDE5 inhibitors on bladder overactivity is discussed. Discussion The questions raised during the plenary session of the 2011 International Consultation on Incontinence-Research Society meeting regarding the benefits of BoNT/A, β3-adrenergic receptor agonist and PDE5 inhibitor treatments of overactive bladder are addressed. Conclusion Our findings suggest that the abovementioned agents, in low enough concentrations, can directly inhibit afferent excitability without decreasing detrusor contractility. Accordingly, they have considerable potential for treating the sensory component of lower urinary tract dysfunctions. Neurourol. Urodynam. 31:300–308, 2012. © 2012 Wiley Periodicals, Inc.

Common methods for inducing neurogenic bladder overactivity include: (1) spinal cord transection or contusion injury, (2) supraspinal injury (decerebration, local lesions, middle cerebral artery occlusion and (3) systemic (e.g., cyclophosphamide) or intravesical administration of irritant or inflammatory agents (e.g., acrolein, acid, lipopolysaccharide). These insults can cause peripheral, spinal and supraspinal effects which are difficult to resolve using traditional cystometry alone. To address this problem, we have developed new methods as well as new highly reproducible models to study neurogenic bladder dysfunction in the mouse. These models include direct sensitization of the bladder using ionizing radiation or indirect sensitization through colonic irradiation (figure 1). The classic model for neurogenic bladder overactivity is spinal cord injury which results in non-voiding contractions during the filling phase. However, there is also an intrinsic myogenic component that is present when studied in vitro. Our radiation models are purely neurogenic and do exhibit overactivity in isolated tissues. Figure: 1 New in vitro studies utilize bladder-urethra sheets, spinal cord slices and in-line cultured cells. Bladder-urethra sheets can be isolated with the hypogastric (T11-L2) and pelvic (L6-S2) nerves and imaged for the propagation of optical activity along the bladder wall simultaneously with afferent nerve and tension recordings. Alternatively, bladder sheets can be mounted vertically as cross-sections for measuring the spread of activity across the wall from mucosa to serosa. Spinal cord slices are sectioned 0.5 to 1 mm thick to include the dorsal and ventral roots which can be stimulated to optically map pathologically-induced changes in spinal cord circuitry. In-line cultured cells, on the other hand, allow for different linear arrangements of urothelial, interstitial, smooth muscle and dorsal root ganglion cells to map alterations in the flow of information in cell-to-cell communication. In vivo studies utilize cystometry which can be combined with cerebro-cortical imaging of optical activity during bladder filling and emptying. The new models of neurogenic overactivity are induced by irradiation of the bladder (direct sensitization) or colon (indirect cross-sensitization) both of which result in inflammation and urothelial damage leading to bladder overactivity. The changes induced by these models have been compared to those induced by the well established models of chronic spinal cord transection at the thoracic (T8-T9) and lumbar (L4-L5) levels.

A substantial portion of the current research on lower urinary tract dysfunction is focused on afferent mechanisms. The main goals are to define and modulate the signaling pathways by which afferent information is generated, enhanced and conveyed to the central nervous system. Alterations in bladder afferent mechanisms are a potential source of voiding dysfunction and an emerging source for drug targets. Established drug therapies such as muscarinic receptor antagonists, and two emerging therapies, beta(3)-adrenergic receptor agonists and botulinum toxin type-A, may act partly through afferent mechanisms. This review focuses on these two new principles and new and established methods for determining their sites of action. It also provides brief information on the innervation of the bladder, afferent receptors and transmitters and how these may communicate with the urothelium, interstitial cells and detrusor smooth muscle to regulate micturition. Peripheral and central mechanisms of afferent sensitization and myogenic mechanisms that lead to detrusor overactivity, overactive bladder symptoms and urgency sensations are also covered. This work is the result from 'Think Tank' presentations, and the lengthy discussions that followed, at the 2010 International Consultation on Incontinence Research Society meeting in Bristol, UK. Neurourol. Urodynam. 30:684-691, 2011.

OBJECTIVE: To assess the safety and efficacy of anticholinergics in men with lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) by a systematic review of published reports and a meta-analysis of the reported outcomes. METHODS: We searched Medline, Embase and Cochrane databases (1966-2006), and hand-searched relevant reference lists and conference proceedings, for studies on the use of anticholinergics in men with BPH or bladder outlet obstruction. Eligible studies were assessed for quality and foreign language studies were translated. We collected data on all reported outcomes, conducted meta- analyses on the maximum urinary flow rate (Q(max)), postvoid residual urine volume (PVR) and volume at first contraction, and calculated the acute urinary retention (AUR) rate. We used sensitivity analysis to confirm the findings. RESULTS: We identified five randomized controlled trials (RCTs) and 15 observational studies. Four RCTs incorporating 633 patients were included in the meta-analyses. Anticholinergics did not significantly alter Q(max) (0.1 mL/s, 95% confidence interval, CI, 0.6-0.7). The PVR was increased by 11.6 mL (95% CI 4.5-18.6) although there was no significant difference between AUR rates. The total International Prostate Symptom Scores (IPSS) were not significantly different, but there were improvements for IPSS storage subscores in one RCT. The AUR rate was 0.3% at the 12-week follow-up in 365 men in the RCTs and observational studies. CONCLUSION: Anticholinergic use in men with LUTS suggestive of BPH appears to be safe. Further studies are required to establish efficacy with a suitable precision.

INTRODUCTION AND OBJECTIVES: Many urologic conditions are associated with dysfunction of autonomic bladder innervation. A precise understanding of the anatomical relationship of the bladder neck and its autonomic nervous system could substantially improve treatment modalities. Currently there is no comprehensive model that provides a detailed three-dimensional (3D) view of bladder neck innervation. Therefore, we sought to create a 3D reconstruction of autonomic nervous tissue innervating the bladder neck using male and female cadaver histopathology. METHODS: We obtained intact pelvic tissues from a male and female cadaver. Axial cross-sections of the bladder neck were generated at 3e5mm intervals and stained for S100 protein. Distances between autonomic nerves and bladder mucosa were recorded. Nerve tracings were manually demarcated and imported into SolidWorks (Waltham, MA, USA) and Blender (Amsterdam, Netherlands) software programs to generate 3D reconstructions of autonomic nerve anatomy. RESULTS: Longitudinal and circular nerve tracings were successfully demarcated and converted into a 3D image reconstruction. We successfully precisely characterized anatomic nerve distributions. Axial cross-sections and 3D images showed that autonomic innervation was highly concentrated in the posterior aspect of the bladder neck in both male and female bladder specimens (Figures 1 and 2). The mean distances between autonomic nerve tissue and the bladder mucosa was 1.15mm posteriorly and 4.0mm anteriorly in the male bladder (0.27-2.87 vs. 2.03-6.20, p

The micturition reflex involves afferent nerve activation when the bladder is sufficiently full and subsequent controlled firing of parasympathetic efferent nerves to contract the detrusor muscle as part of the voiding mechanism. Alteration of the sensitivity of afferent activation or loss of control over transmitter release could lead to sensory- or motor-activated incontinence, respectively. The control mechanisms that regulate these 2 activities remain poorly understood. Current opinion is that the sensation of bladder fullness is relayed by afferent nerves in the mucosal layer, which are activated by the release of chemical mediators, such as adenosine triphosphate (ATP), from the urothelium when it is stretched as the bladder fills. This hypothesis supports the concept that other chemical signals that affect bladder sensation (eg, changes in urine composition and agents such as capsaicin) can modulate the sensitivity of the basic system. It has also been proposed that a layer of myofibroblasts immediately below the basal lamina of the urothelium acts as a variable gain regulator of the sensory process between ATP release and afferent excitation. These myofibroblasts are functionally connected to form an electrical syncytium, make close contact with nerves, and respond by generating electrical responses and transient increases in intracellular Ca2+ when exposed to ATP. On the efferent side, using a guinea pig detrusor model, possible modulators of transmitter release have been investigated, including adenosine (the breakdown product of the neurotransmitter ATP). Adenosine reduces the force of nerve-mediated contractions by acting predominantly at presynaptic sites at the nerve-muscle junction via a subtype of an adenosine receptor-the A1 receptor. An additional effect, possibly via A2 receptors, is also present on the detrusor muscle itself. These actions of adenosine are less evident in human detrusor muscle but remain a potential modulatory target. In summary, the cellular and molecular regulation of bladder fullness sensation and efferent transmitter release are becoming better understood and represent potential drug targets for the management of detrusor overactivity.

Contractile activation of detrusor smooth muscle is initiated by the release of transmitters from motor nerves. Acetylcholine is a ubiquitous transmitter, as also is adenosine triphosphate (ATP) in many animal bladders and in people from several patient groups with pathological bladder function. In recent years there has been progress in explaining several cellular mechanisms that link transmitter release to contraction and these will be considered. The lifetime of ATP in the neuromuscular junction is finite and broken down ultimately to adenosine, which can exert modulatory control of contractile activation. Adenosine depresses nerve-mediated contractions and two sites of action have been proposed: an action on the motor nerves via A receptors to depress further transmitter release and a less well-defined depressant effect on the detrusor muscle. The Ca2+ ions that activate the contractile proteins are derived from intracellular stores, which releases their content via IP receptor activation and Ca2+-induced Ca2+ release. Filling of the stores in the rest interval is mediated via transmembrane flux of Ca2+through Ca2+ channels. Activation of the channels is regulated by the level of the intracellular [Ca2+], via activation and inactivation of Ca2+-sensitive K channels. Thus, Ca2+ store filling is regulated by intracellular [Ca2+] via a negative feedback process. The presence and physiological function of spontaneous contractions in detrusor remain contentious and little is known about their origin. One possibility is that they originate from random Ca2+ sparks, i.e. localized transient increases of [Ca2+] that may eventually progress to generate a cellular Ca2+ transient. Observations by confocal microscopy have revealed the presence of such sparks, especially near the cell membrane, and thus provide a cellular basis for spontaneous contractions. Finally, the questions arises as to whether detrusor smooth muscle is a functional syncitium. The demonstration of small gap junctions by electron microscopy and the demonstration of the gap junction protein connexin45 indicate that the muscle mass may indeed be functionally connected. The implications regarding the spread of excitation are discussed.

Interstitial cystitis is a chronic pelvic pain syndrome of which the origin and mechanisms involved remain unclear. In this study Ca(2+) transients in the bladder wall of domestic cats diagnosed with naturally occurring feline interstitial cystitis were examined.Cross-sections of full-thickness bladder strips from normal cats and cats with feline interstitial cystitis were examined by optically mapping Ca(2+) transients and recording tension. Responses of Ca(2+) activity and detrusor contractions to pharmacological interventions were compared. In addition, pharmacological responses were compared in mucosa denuded preparations.Optical mapping showed that feline interstitial cystitis bladders had significantly more spontaneous Ca(2+) transients in the mucosal layer than control bladders. Optical mapping also demonstrated that feline interstitial cystitis bladders were hypersensitive to a low dose (50 nM) of the muscarinic receptor agonist arecaidine when the mucosal layer was intact. This hypersensitivity was markedly decreased in mucosa denuded bladder strips.In feline interstitial cystitis cat bladders there is increased Ca(2+) activity and sensitivity of muscarinic receptors in the mucosal layer, which can enhance smooth muscle spontaneous contractions.