Detrusor Strips Research Articles

Changes in the Function and Expression of T-Type and N-Type Calcium Channels in the Rat Bladder after Bladder Outlet Obstruction

We evaluated possible changes in the function and expression of T-type and N-type Ca(2+) channels in the bladder of rats with bladder outlet obstruction. Female Sprague Dawley® rats were divided into a group with bladder outlet obstruction created by partial urethral ligation and a sham operated group. Six weeks postoperatively we determined the mRNA expression of T-type and N-type Ca(2+) channels in the bladder, dorsal root ganglion and spinal cord. We also cystometrically investigated expression by intravenous administration of the T-Ca blocker RQ-00311610 or the N-type Ca(2+) channel blocker ω-conotoxin GVIA. We then performed in vitro functional studies of detrusor strips using these blockers. mRNA expression of T-type Ca(2+) channels in the bladder detrusor and mucosa layers, and the spinal cord dorsal horn, and N-type Ca(2+) channels in the whole bladder and detrusor layer, and the spinal cord dorsal horn was greater in the obstructed group than the sham operated group. In obstructed rats bladder capacity and voided volume increased after RQ-00311610 administration but the number of nonvoiding contractions decreased after ω-conotoxin GVIA administration. Detrusor strips from obstructed rats showed weaker contractile responses to electrical field stimulation, particularly in regard to the purinergic component. ω-Conotoxin GVIA suppressed electrical field stimulation induced contractions only in the detrusor of obstructed rats, especially the cholinergic component. Blocking T-type Ca(2+) channels increased bladder capacity while N-type Ca(2+) channel blockade inhibited nonvoiding contractions in rats with bladder outlet obstruction. Decreased bladder efferent neurotransmission occurred after bladder outlet obstruction, predominantly in its purinergic component and detrusor contractions via cholinergic neurotransmission were activated in a compensatory manner, probably via N-type Ca(2+) channel up-regulation.

Low concentrations of niflumic acid enhance basal spontaneous and carbachol-induced contractions of the detrusor

The urinary bladder expresses Ca(2+)-activated Cl(-) channels (CACC), but its physiological role in governing contractility remains to be defined. The CACC modulator niflumic acid (NFA) is widely used despite the variable results arisen from different drug concentrations used. This study was designed to examine the effects of NFA at low concentrations on detrusor strip contractility. Rat detrusor strips with mucosa-intact (+MU) and mucosa-denuded (-MU) were prepared in transverse (Tr) and longitudinal (Lg) with respect to the bladder orientation. Isometric force measurements were made at baseline (for spontaneous phasic contractile activity) and during drug stimulation (by carbachol, CCh) with and without NFA. NFA (1 and 10 μmol/L) pretreatment enhanced CCh-induced contractions more in +MU than -MU strips with no selectivity on contractile direction. For spontaneous phasic contractions, NFA-treated strips in the Tr direction showed increased phasic amplitude, while phasic frequency was unchanged. The findings suggest low concentrations of NFA having a potentiating effect on detrusor contractions that was sensitive to the MU and contractile direction.

Mucosa of human detrusor impairs contraction and β‐adrenoceptor‐mediated relaxation

To elucidate the impact of the mucosa on detrusor muscle function by investigating force of contraction under various stimulatory conditions and during subsequent relaxation using catecholamines. Detrusor tissue was obtained from patients who had undergone cystectomy for bladder cancer and strips of intact or mucosa-denuded muscle were set up for force measurement. Preparations were precontracted with KCl, carbachol or electric-field stimulation (EFS). Precontracted strips were relaxed using increasing concentrations of catecholamines in the absence and presence of the subtype-selective β-adrenoceptor (AR) blockers CGP 20712A (β1-ARs), ICI 118,551 (β2-ARs), and L-748,337 (β3-ARs). Force development in response to KCl (40 mM), carbachol (1 μM) or EFS was larger in the absence of mucosa than in intact muscle strips. The force of contraction of mucosa-denuded strips with detached urothelium incubated in the same chamber was as low as in intact strips. Noradrenaline relaxed precontracted detrusor strips to a significantly larger extent and at lower concentrations in denuded than in intact strips. CGP 20712A did not affect noradrenaline-induced relaxation of denuded and intact strips, and ICI 118,551 did not influence noradrenaline-induced relaxation in denuded strips, but abolished the difference between denuded and intact strips. The antagonism of the relaxant effects of noradrenaline by L-748,337 was slightly smaller in intact than denuded strips. The mucosa of human detrusor strips impairs force development when stimulated with KCl, carbachol or EFS. The mucosa also blunts the relaxing effects of catecholamines. The latter effect does not involve the activation of β1-ARs but only of β2-ARs, whereas β3-ARs mediate the relaxation of human detrusor.

Crucial role of TRPC1 and TRPC4 in cystitis-induced neuronal sprouting and bladder overactivity.

PurposeDuring cystitis, increased innervation of the bladder by sensory nerves may contribute to bladder overactivity and pain. The mechanisms whereby cystitis leads to hyperinnervation of the bladder are, however, poorly understood. Since TRP channels have been implicated in the guidance of growth cones and survival of neurons, we investigated their involvement in the increases in bladder innervation and bladder activity in rodent models of cystitis.Materials and MethodsTo induce bladder hyperactivity, we chronically injected cyclophosphamide in rats and mice. All experiments were performed a week later. We used quantitative transcriptional analysis and immunohistochemistry to determine TRP channel expression on retrolabelled bladder sensory neurons. To assess bladder function and referred hyperalgesia, urodynamic analysis, detrusor strip contractility and Von Frey filament experiments were done in wild type and knock-out mice.ResultsRepeated cyclophosphamide injections induce a specific increase in the expression of TRPC1 and TRPC4 in bladder-innervating sensory neurons and the sprouting of sensory fibers in the bladder mucosa. Interestingly, cyclophosphamide-treated Trpc1/c4−/− mice no longer exhibited increased bladder innervations, and, concomitantly, the development of bladder overactivity was diminished in these mice. We did not observe a difference neither in bladder contraction features of double knock-out animals nor in cyclophosphamide-induced referred pain behavior.ConclusionsCollectively, our data suggest that TRPC1 and TRPC4 are involved in the sprouting of sensory neurons following bladder cystitis, which leads to overactive bladder disease.

Functional expression of KCNQ (Kv7) channels in guinea pig bladder smooth muscle and their contribution to spontaneous activity

Background and PurposeThe aim of the study was to determine whether KCNQ channels are functionally expressed in bladder smooth muscle cells (SMC) and to investigate their physiological significance in bladder contractility.Experimental ApproachKCNQ channels were examined at the genetic, protein, cellular and tissue level in guinea pig bladder smooth muscle using RT-PCR, immunofluorescence, patch-clamp electrophysiology, calcium imaging, detrusor strip myography, and a panel of KCNQ activators and inhibitors.Key ResultsKCNQ subtypes 1–5 are expressed in bladder detrusor smooth muscle. Detrusor strips typically displayed TTX-insensitive myogenic spontaneous contractions that were increased in amplitude by the KCNQ channel inhibitors XE991, linopirdine or chromanol 293B. Contractility was inhibited by the KCNQ channel activators flupirtine or meclofenamic acid (MFA). The frequency of Ca2+-oscillations in SMC contained within bladder tissue sheets was increased by XE991. Outward currents in dispersed bladder SMC, recorded under conditions where BK and KATP currents were minimal, were significantly reduced by XE991, linopirdine, or chromanol, and enhanced by flupirtine or MFA. XE991 depolarized the cell membrane and could evoke transient depolarizations in quiescent cells. Flupirtine (20 μM) hyperpolarized the cell membrane with a simultaneous cessation of any spontaneous electrical activity.Conclusions and ImplicationsThese novel findings reveal the role of KCNQ currents in the regulation of the resting membrane potential of detrusor SMC and their important physiological function in the control of spontaneous contractility in the guinea pig bladder.

Gap junction expression and the effects of gap junction inhibitors in overactive bladder models: does ovariectomy have a role?

In this study, gap junction expression and the effects of estrogen deficiency and gap junction inhibitors were investigated in overactive bladder models which were created by bladder outlet obstruction. In our study, we created four groups as control, ovariectomy, bladder outlet obstruction (BOO), and ovariectomy + BOO. We investigated the effects of oxybutynin and 18-alpha glycyrrhetinic acid (18-α-GA) which is a gap junction blocker on isolated detrusor strips. Western blot method was used to measure the level of connexin-43 in detrusor. Bladder weights were significantly increased in the BOO and ovariectomy + BOO groups (p < 0.05). There was no statistically significant difference in the maximal contraction responses to carbachol between ovariectomy and control groups. In BOO and ovariectomy + BOO groups, contractile responses were significantly prominent with higher doses of carbachol. Oxybutynin-induced relaxant responses of BOO and ovariectomy + BOO groups were significantly higher than control group (p < 0.05). The relaxation effect of 18-a-GA was more effective in the obstruction groups. Among those two groups, the relaxation observed in BOO group was higher than ovariectomy + BOO group in higher doses of 18-a-GA. Connexin-43 expression was increased in BOO group compared with the control group (p = 0.006). Ovariectomy did not change connexin-43 expression alone; however, when combined with BOO, connexin-43 expression decreased significantly (p = 0.023). Ovariectomy had no effect on the gap junctions in the bladder and bladder overactivity alone. Therefore, obstruction is the main factor that increases the amount of gap junctions, and gap junction blockers are thus more effective in obstruction. However, ovariectomy was shown to decrease the expression of gap junctions and relaxation effect of gap junction blockers, when combined with BOO.

Expression of Hsp27 correlated with rat detrusor contraction after acute urinary retention

Heat shock protein 27 (Hsp27) can regulate actin cytoskeleton dynamics and contractile protein activation. This study investigates whether Hsp27 expression is related to bladder contractile dysfunction after acute urinary retention (AUR). Female rats were randomized either to AUR by urethral ligation or to normal control group. Bladder and smooth muscle strip contraction at time points from 0 h to 7 days after AUR were estimated by cystometric and organ bath studies. Hsp27 expression in bladder tissue at each time point was detected with immunofluorescence, Western blots, and real-time PCR. Expression of the three phosphorylated forms of Hsp27 was detected by Western blots. Smooth muscle ultrastructure was observed by transmission electron microscopy. Data suggest that maximum detrusor pressure and both carbachol-induced and spontaneous detrusor strip contraction amplitude decreased gradually for the duration from 0 to 6 h, and then increased gradually to near-normal values at 24 h. Treatment of muscle strips with the p38MAK inhibitor, SB203580, inhibited carbachol-induced contractions. Smooth muscle ultrastructure damage was the highest at 6 h after AUR, and then lessened gradually during next 7 days, and ultrastructure was close to normal. Expressions of Hsp27 mRNA and protein and the proteins of the three phosphorylated forms were higher at 0 h, decreased to lower levels up to 6 h, and then gradually increased. Therefore, we conclude that rat bladder contractile function after AUR worsens during 0-6 h, and then gradually recovers. The findings of the current study suggest that Hsp27 modulates bladder smooth muscle contraction after AUR, and that phosphorylation of Hsp27 may be an important pathway modulating actin cytoskeleton dynamics in bladder smooth muscle contraction and reconstruction after injury.

120 CHOLINERGIC MEDIATED DETRUSOR DYSFUNCTION IN A FEMALE RAT MODEL OF PELVIC NEUROPRAXIA

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Basic Research (II)1 Apr 2013120 CHOLINERGIC MEDIATED DETRUSOR DYSFUNCTION IN A FEMALE RAT MODEL OF PELVIC NEUROPRAXIA Johanna L. Hannan, Vinson M. Wang, Xiaopu Liu, Fabio Castiglione, Arthur L. Burnett, Petter Hedlund, and Trinity J. Bivalacqua Johanna L. HannanJohanna L. Hannan Baltimore, MD More articles by this author , Vinson M. WangVinson M. Wang Baltimore, MD More articles by this author , Xiaopu LiuXiaopu Liu Baltimore, MD More articles by this author , Fabio CastiglioneFabio Castiglione Milan, Italy More articles by this author , Arthur L. BurnettArthur L. Burnett Baltimore, MD More articles by this author , Petter HedlundPetter Hedlund Lund, Sweden More articles by this author , and Trinity J. BivalacquaTrinity J. Bivalacqua Baltimore, MD More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1499AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Urogynaecological surgeries for cervical or colorectal cancer often result in damage to the neurovascular bundle and pelvic plexus leading to voiding dysfunction. Although surgical techniques have improved mortality rates in women, the prevalence of voiding dysfunctions severely impacts their quality of life. The objective of this study is to characterize the physiological and molecular changes in the bladder that take place following bilateral pelvic nerve injury (BPNI) in female rats. METHODS Female Sprague-Dawley rats (12 wks, n=40) were divided into sham or BPNI in which nerves from the pelvic plexus innervating bladder were bilaterally crushed. Animals were examined 7, 14, 30 and 60 days following BPNI. The bladder was excised and weighed. Detrusor strips were mounted in a myograph and contractile responses to increasing concentrations of carbachol or frequencies of electrical field stimulation (EFS) were assessed. Gene expression of muscarinic type 2 (MR2) and 3 (MR3) receptors was also assessed by RTPCR. RESULTS There was a significant increase in the bladder weight at 7 and 14 days following nerve injury (Sham: 3.1%; 7d: 3.9%; 14d: 4.3% of body weight, p<0.05). Maximal carbachol-mediated contraction was decreased following nerve injury at all time points (Sham: 17.9mN; 7d: 12.3mN; 14d: 12.8mN; 30d: 13.4mN; 60d: 14.8mN; p<0.05). Furthermore, the EC50 was markedly increased at 14 and 30 days after BPNI (Sham: 7.0 pD2; 14d: 6.0 pD2; 30d: 6.2 pD2, p<0.05) indicating that a higher concentration of carbachol was required to achieve 50% of maximal contraction. By 60 days following BPNI, maximal contraction and EC50 to carbachol had returned to sham levels. EFS mediated contraction was also significantly reduced after nerve injury when stimulate at lower frequencies (<8Hz) but all groups achieved the same maximal contraction at high frequency stimulation (32Hz). Gene expression of MR2 was significantly lower 7 days following nerve injury (Sham: 0.91; 7d: 0.34 MR2 relative expression, p<0.05); however gene expression of MR3 was unchanged 7days following BPNI. CONCLUSIONS These results demonstrate that pelvic nerve injury leads to a temporal increase in bladder weight and decreased cholinergic mediated contractions and MR2 gene expression. These data suggest that this animal model can be used to study the mechanism of neuropraxia-induced voiding dysfunction in female rats. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e48 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Johanna L. Hannan Baltimore, MD More articles by this author Vinson M. Wang Baltimore, MD More articles by this author Xiaopu Liu Baltimore, MD More articles by this author Fabio Castiglione Milan, Italy More articles by this author Arthur L. Burnett Baltimore, MD More articles by this author Petter Hedlund Lund, Sweden More articles by this author Trinity J. Bivalacqua Baltimore, MD More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

31 THE ROLE OF TRPA1 IN THE BLADDER COOLING REFLEX; A POSSIBLE NEW THERAPEUTIC TARGET

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Basic Research (1)1 Apr 201331 THE ROLE OF TRPA1 IN THE BLADDER COOLING REFLEX; A POSSIBLE NEW THERAPEUTIC TARGET Pieter Uvin, Mathieu Boudes, Jan Franken, Silvia Pinto, Wouter Everaerts, Anne-Cathérine Boeve, Roma Rietjens, Yeranddy A. Alpizar, Grzegorz Owsianik, Karel Talavera, Dirk De Ridder, and Thomas Voets Pieter UvinPieter Uvin Leuven, Belgium More articles by this author , Mathieu BoudesMathieu Boudes Leuven, Belgium More articles by this author , Jan FrankenJan Franken Leuven, Belgium More articles by this author , Silvia PintoSilvia Pinto Leuven, Belgium More articles by this author , Wouter EveraertsWouter Everaerts Leuven, Belgium More articles by this author , Anne-Cathérine BoeveAnne-Cathérine Boeve Leuven, Belgium More articles by this author , Roma RietjensRoma Rietjens Leuven, Belgium More articles by this author , Yeranddy A. AlpizarYeranddy A. Alpizar Leuven, Belgium More articles by this author , Grzegorz OwsianikGrzegorz Owsianik Leuven, Belgium More articles by this author , Karel TalaveraKarel Talavera Leuven, Belgium More articles by this author , Dirk De RidderDirk De Ridder Leuven, Belgium More articles by this author , and Thomas VoetsThomas Voets Leuven, Belgium More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1406AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES For decades, bladder contraction upon instillation of ice-cold saline into the bladder has been used as a diagnostic test for neurogenic detrusor overactivity (NDO). TRPA1 is a member of the Transient Receptor Potential (TRP) ion channel superfamily that is activated by irritant chemicals, as well as cold temperatures. TRPA1 is expressed in sensory afferents in the bladder and chemical activation of this channel induces urinary frequency in mice and rats. We hypothesized that TRPA1 plays a crucial role in the bladder cooling reflex, leading to ice-water induced detrusor overactivity. METHODS Female 10-12 weeks old Wistar rats were used for experiments. Expression of TRPA1 was assessed by quantitative RT-PCR on L6-S1 dorsal root ganglion neurons (DRG), urothelial cells and detrusor smooth muscle cells. Functional responses to cold stimuli were evaluated in ex vivo isolated detrusor strip contractility measurements. In vivo cystometry recordings were performed in urethane-anaesthetized rats (1g/kg, intraperitoneal (i.p.) injection) and in conscious rats 3-4 weeks after spinal cord injury (SCI). The TRPA1 blocker HC030031 (100mg/kg, i.p.), the ganglionic blocker hexamethonium (20mg/kg, i.p.) and the vehicle were administered 20 minutes before the cold stimulus. RESULTS TRPA1 is highly expressed in DRG neurons, while the expression in urothelium and detrusor is very low (n = 3). In urethane-anaesthetized (n = 8) and SCI rats (n = 7), the intravesical pressure increases were observed upon instillation of ice-cold saline. These effects were significantly smaller in HC030031 pretreated animals, compared to vehicle. Administration of hexamethonium had no effect on the bladder cooling reflex (n = 6). In detrusor strip experiments, we observed a clear cold-induced contraction (n = 8), which could be largely abolished by the TRPA1 blocker HC030031 (100μM) (n = 8). This cold-induced bladder contraction was significantly diminished in a 0 Ca2+ BAPTA solution (n = 8) and in indomethacin 1μM (n = 8) but not in atropine 1μM (n = 8) and suramine 300μM (n = 8). CONCLUSIONS The bladder cooling reflex in rats is largely mediated by TRPA1. Surprisingly, it doesn't involve a spinar reflex arch, but seems to be a “local” phenomenon via local release of neuropeptides. Ultimately, TRPA1 could become an important therapeutic target to relieve symptoms in patients with NDO and OAB. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e12 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Pieter Uvin Leuven, Belgium More articles by this author Mathieu Boudes Leuven, Belgium More articles by this author Jan Franken Leuven, Belgium More articles by this author Silvia Pinto Leuven, Belgium More articles by this author Wouter Everaerts Leuven, Belgium More articles by this author Anne-Cathérine Boeve Leuven, Belgium More articles by this author Roma Rietjens Leuven, Belgium More articles by this author Yeranddy A. Alpizar Leuven, Belgium More articles by this author Grzegorz Owsianik Leuven, Belgium More articles by this author Karel Talavera Leuven, Belgium More articles by this author Dirk De Ridder Leuven, Belgium More articles by this author Thomas Voets Leuven, Belgium More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

105 BLADDER DYSFUNCTION IN MICE WITH EXPERIMENTAL AUTOIMMUNE ENCEPHALITIS IS A MODEL FOR NEUROGENIC OAB IN MULTIPLE SCLEROSIS

You have accessJournal of UrologyUrodynamics/Incontinence/Female Urology: Basic Research (II)1 Apr 2013105 BLADDER DYSFUNCTION IN MICE WITH EXPERIMENTAL AUTOIMMUNE ENCEPHALITIS IS A MODEL FOR NEUROGENIC OAB IN MULTIPLE SCLEROSIS Jan Franken, Mathieu Boudes, Pieter Uvin, Anne-Cathérine Boeve, Thomas Voets, Jerome Hendriks, Niels Hellings, and Dirk De Ridder Jan FrankenJan Franken Leuven, Belgium More articles by this author , Mathieu BoudesMathieu Boudes Leuven, Belgium More articles by this author , Pieter UvinPieter Uvin Leuven, Belgium More articles by this author , Anne-Cathérine BoeveAnne-Cathérine Boeve Leuven, Belgium More articles by this author , Thomas VoetsThomas Voets Leuven, Belgium More articles by this author , Jerome HendriksJerome Hendriks Hasselt, Belgium More articles by this author , Niels HellingsNiels Hellings Hasselt, Belgium More articles by this author , and Dirk De RidderDirk De Ridder Leuven, Belgium More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1484AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Experimental autoimmune encephalitis (EAE) in mice has widely been used as a model for multiple sclerosis (MS). Most MS patients suffer from bladder dysfunction, often refractory to antimuscarinics. It was shown that EAE mice void more frequently and that their bladder walls undergo morphological changes. We investigated whether bladder dysfunction in EAE mice mimics the clinical phenotype in MS patients. METHODS To induce EAE, we injected 12-weeks-old C57/Bl-6 mice with Myelin Oligodendrocyte Glycoprotein (MOG35-55). Controls were injected with Complete Freund's Adjuvant (CFA). We monitored all mice daily for clinical signs of EA and divided them in groups according to severity, with at least 5 animals per group. Scoring went as follows: Score 1, flaccid tail; Score 2, hindlimb weakness; Score 3, hindlimb paralysis; Score 4, paralysis up to diaphragm. First, we assessed muscarinic receptor function and contractile strength in vitro. Therefore, we exposed detrusor strips to increasing concentrations of carbachol (0.1-10μM). Afterwards, maximal contractile strength was measured with a high-potassium solution. Second, in in vivo experiments, bladder capacity (BC) and post-void residue (PVR) were measured. Physiological bladder contractions were assessed with urodynamic investigations in anesthetized mice, by placing a catheter in the dome of the bladder while continuously perfusing it with saline. RESULTS In vitro, maximal contractile strength decreased 30% in response to high potassium, but relative contribution of muscarinic receptors to contractile function was not altered. In vivo, BC increased in EAE mice with score 2 or higher, up to a 2.5 fold increase in score 4. Unlike controls, all animals with EAE score 3 and 4 had a PVR larger than 20μl. In mice with EAE score 4 voiding frequency increased 3.6 fold. In score 3 we observed a 62% and in score 2 a 36% increase, but the latter was not significant. The amount of NVC was significantly higher in score 1, 2 and 3. In score 4 we did not observe NVC, because there was dribbling or voiding in every contraction. Basal pressure increased 2.6 fold in all animals with score 2 or more, whereas maximal pressure was comparable with controls. CONCLUSIONS Bladder dysfunction in EAE mice closely mimics its clinical counterpart in MS. Severity of symptoms on urodynamic investigation always correlated with disease score. Interestingly, in vitro contractile properties were also altered, suggesting that at least part of the urinary symptoms arise from local changes in the bladder. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e42 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Jan Franken Leuven, Belgium More articles by this author Mathieu Boudes Leuven, Belgium More articles by this author Pieter Uvin Leuven, Belgium More articles by this author Anne-Cathérine Boeve Leuven, Belgium More articles by this author Thomas Voets Leuven, Belgium More articles by this author Jerome Hendriks Hasselt, Belgium More articles by this author Niels Hellings Hasselt, Belgium More articles by this author Dirk De Ridder Leuven, Belgium More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

38 MOUSE VOIDING PATTERNS ARE STRAIN-SPECIFIC: A NOVEL ASSAY FOR AGE-RELATED DEVELOPMENT OF LOWER URINARY TRACT SYMPTOMS (LUTS)

They were divided into two groups; BOO and sham operated (SHAM) groups. BOO was created by a partial ligation of the urethra with a nylon suture, whereas SHAM group of rats underwent only exposure of the urethra without ligation. Six weeks after surgery, three experiments were performed: 1) mRNA expression of N-Ca in the bladder, 2) in vitro functional studies of detrusor strips focusing on the effects of -conotoxin GVIA (CTX; a selevctive N-Ca inhibitor, 10 6 M) on carbachol (CCh)-induced and electrical field stimulation (EFS)-induced contractions, and 3) cystometric investiogations (CMG) with continuous salineinstillation in a conscious state with intravenous administration of CTX (3 g/kg). RESULTS: N-Ca mRNA expression in the detrusor was about 50 times higher in BOO than SHAM group. CTX did not suppress CCh-induced contractions in either group. The EFS-induced contractions were weaker in the detrusor of BOO than SHAM group. CTX significantly suppressed the EFS-induced contractions in BOO group, but not in SHAM group (Figure 1 A, B). The suppressive effect of CTX on the EFS-induced contractions was more pronounced in the cholinergic component at 10 Hz. In CMG, the number of non-voiding contractions (NVCs) was significantly greater in BOO group than SHAM group. The number of NVCs was significantly decreased after CTXadministration in BOO rats (Figure 1 C). CONCLUSIONS: The results of the current study suggest that N-Ca may have an important role in facilitating release of excitatory transmitters including ACh from the efferent nerve terminals innervating the detrusor of the BOO rats, and this may link to the development of detrusor overactivity associated with BOO.

Involvement of Connexins 43 and 45 in Functional Mechanism of Human Detrusor Overactivity in Neurogenic Bladder

To assess the involvement of connexins (Cxs) 43 and 45 in the selective inhibition of detrusor contractions in patients with neurogenic detrusor overactivity (NDO). Detrusor strips with and without mucosa were obtained from patients undergoing cystectomy for either neurogenic bladder with NDO refractory to pharmacologic treatment or bladder cancer with no overactive bladder symptoms (ie, control group). The strips were isometrically mounted in organ baths. The effects of the selective inhibitors Cxs 43 and 45 on carbachol-induced contractions of bladder strips were assessed. Overall, 47 patients with a median age of 61 years (range 19-83) were included. The female-to-male ratio was 0.42. Selective inhibitors of Cxs 43 and 45 significantly inhibited carbachol-induced contractions of bladder strips from patient with NDO (P <.01) but had no effect in the control group. When tested without mucosa, the effect of the Cx 43 inhibitor in the NDO patient strips was abolished, but it remained unchanged with the Cx 45 inhibitor (P <.05). The pharmacologic modulation of Cx-mediated intercellular communication, targeting Cxs 43 and 45, is directly involved in the functional mechanism of NDO and might represent a future target for the treatment of NDO.

The effects of Glivec on the urinary bladder excitation of rats with suprasacral or sacral spinal cord transection

BackgroundTo investigate the effects of the c-kit blocker imatinib mesylate (Glivec) on the bladders of animals with suprasacral cord injury (SSCI) and sacral cord injury (SCI). Materials and methodsWe randomized 60 female Sprague-Dawley rats into control, sham, SSCI (T8/9 transection), and SCI (S1-3 transection) groups. Six weeks later, we evaluated the effects of stepwise Glivec administrations on urinary bladder contraction using cystometry and the detrusor strip stretch-test. We investigated spontaneous calcium transients of kit-positive interstitial cells of Cajal (ICCs) with the preloaded Ca2+ indicator fluo-3AM. The expression levels of c-kit and the number of ICCs in those bladders were determined using Western blot and fluorescence staining analyses, respectively. ResultsBladder capacity and compliance were decreased in SSCI bladders and increased in SCI bladders (P<0.05). The amplitude and frequency of spontaneous contractions of detrusor strips, the frequency and relative fluorescence intensity of the spontaneous Ca2+ waves, and c-kit expression in the bladder were significantly increased in the SSCI group and decreased in the SCI group compared with the control and sham groups (P<0.05). The dose-dependent effects of Glivec also confirmed consistent functional variations in bladder activity. ConclusionsThe expressions and effects of Glivec were enhanced in SSCI bladders and inhibited in SCI bladders, which may indicate potential roles of ICCs for the c-kit signaling pathway in the pathogenesis of SSCI and SCI bladder.

Fourier Transform Analysis of Rabbit Detrusor Autonomous Contractions Reveals Length Dependent Increases in Tone and Slow Wave Development at Long Lengths

Bladder wall muscle (detrusor) develops low amplitude rhythmic contractions. Low amplitude rhythmic contraction activity is increased in detrusor from patients with overactive bladder. In this in vitro study we used fast Fourier transforms to assess the length dependence of low amplitude rhythmic contraction components. Rabbit detrusor strips were placed in a muscle bath between 2 clips to adjust length and record isometric tension. Tissues stretched from 70% to 130% of a reference muscle length at 10% increments were allowed to develop low amplitude rhythmic contractions at each length for 20 minutes. Low amplitude rhythmic contraction data were analyzed using fast Fourier transforms and represented by a frequency rather than a time spectrum. Based on fast Fourier transform analysis summarized by signal peaks within specific frequency ranges, rabbit low amplitude rhythmic contraction waveforms were divided into 1 tonic and 2 phasic components, defined as A0 + A1F1 + A2F2, where A0 is a length dependent basal tonic component that increases linearly, A1F1 is a slow wave with a length dependent specific amplitude (A1) and a length independent constant frequency (F1) of approximately 11.2 Hz, and A2F2 is a fast wave with a length dependent amplitude (A2) and frequency (F2) of approximately 0.03 Hz. Fast Fourier transform analysis revealed that rabbit low amplitude rhythmic contractions consist of a basal tonic component plus 2 phasic components. The amplitude of all 3 components was length dependent. The frequency of the fast component was not length dependent and the slow component was absent at short muscle lengths, developing only at muscle lengths beyond that producing a maximum active contraction.

Bladder dysfunction in a transgenic mouse model of multiple system atrophy

Multiple system atrophy (MSA) is an adult-onset neurodegenerative disorder presenting with motor impairment and autonomic dysfunction. Urological function is altered in the majority of MSA patients, and urological symptoms often precede the motor syndrome. To date, bladder function and structure have never been investigated in MSA models. We aimed to test bladder function in a transgenic MSA mouse featuring oligodendroglial α-synucleinopathy and define its applicability as a preclinical model to study urological failure in MSA. Experiments were performed in proteolipid protein (PLP)-human α-synuclein (hαSyn) transgenic and control wild-type mice. Diuresis, urodynamics, and detrusor strip contractility were assessed to characterize the urological phenotype. Bladder morphology and neuropathology of the lumbosacral intermediolateral column and the pontine micturition center (PMC) were analyzed in young and aged mice. Urodynamic analysis revealed a less efficient and unstable bladder in MSA mice with increased voiding contraction amplitude, higher frequency of nonvoiding contractions, and increased postvoid residual volume. MSA mice bladder walls showed early detrusor hypertrophy and age-related urothelium hypertrophy. Transgenic hαSyn expression was detected in Schwann cells ensheathing the local nerve fibers in the lamina propria and muscularis of MSA bladders. Early loss of parasympathetic outflow neurons and delayed degeneration of the PMC accompanied the urological deficits in MSA mice. PLP-hαSyn mice recapitulate major urological symptoms of human MSA that may be linked to αSyn-related central and peripheral neuropathology and can be further used as a preclinical model to decipher pathomechanisms of MSA.

The impact of vitamin E supplementation on urinary bladder contractility in streptozotocin-induced diabetic rats

Background: To determine whether vitamin E protects streptozotocin-induced diabetic rats from diabetic urinary bladder dysfunction and discover its possible mechanism. Materials and Methods: A total of 40 rats were randomly divided into four groups: a control group (A), a diabetic group (B), a group given vitamin Eonly (C), and a diabetic group given vitamin E therapy for 8 weeks (D). Diabetes was induced in the rats by 65 mg/kg streptozosin (STZ) via an intraperitoneal (i.p.) injection. Vitamin E was given in a dose of 50 mg/kg/day i.p. Under urethane anaesthesia (1.2 g/kg) subcutaneously and decapitation, contractile responses to carbachol of detrusor strips in all groups were studied in vitro. The levels of nitrite, nitrate, malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) were detected in bladder tissues homogenates. Apoptosis studies were performed by detection of the levels of caspase 3 and cell death detection. Results: The bladder weights were significantly increased (p&lt;0.001) in diabetic groups compared to the other studied groups. Contractile responses to carbachol increased in the diabetic group more than in the other groups (p&lt;0.001). Vitamin E improved the contractile responses of group D and improved them but still significantly higher than those of control group (p &lt; 0.05). Vitamin E treatment decreased the tissue MDA, nitrite, nitrate and GSH levels of group D which were significantly higher in group B than A and C groups (p&lt;0.001). All enzyme activities of group B were significantly lower than those of the other groups, although they increased significantly in group D but still lower than those of A and C groups. However, no significant differences were detected between the levels of GPx and SOD of group D and those of A and C groups. Conclusions: These data suggest that vitamin E supplementation may be beneficial in delaying the progression of diabetic dysfunction in experimental animal model.

The effect of hypercholesterolemia on the contractions of detrusor strips in response to neural stimulation in rats

Aim: To investigate the effect of hypercholesterolemia on in vitro contractions of the detrusor smooth muscle stimulated by an electrical field in rats and the relative contributions of cholinergic and noncholinergic neurotransmission. Materials and methods: Sprague-Dawley rats were fed with standard diet (NC group) or with 4% cholesterol diet (HC group). After 4 weeks, the urinary bladder was excised under anesthesia and 4 detrusor strips were prepared. Blood samples were collected for serum lipid profile, and aortic arches were dissected for histopathological examination. In vitro contractile function of the detrusor smooth muscle was evaluated with: 1) electrical field stimulation (EFS) only, 2) EFS in the presence of muscarinic antagonist atropine (10–6 M), and 3) EFS in the presence of atropine and P2X antagonist phosphate-6-azo (benzene-2,4-disulfonic acid) tetrasodium salt hydrate (10–4 M) or P2X agonist α,β-methylene-adenosine-5’-triphosphate (10–5 M). Results: Plasma cholesterol was elevated in the HC group (P < 0.05); there was no sign of atherosclerosis. Although EFS-induced contractions were higher in the HC group, the difference was not significant. Cholinergic or purinergic antagonists decreased contractile response by desensitization with the purinergic agonist. The contribution of cholinergic and purinergic components were also similar in both groups. Conclusion: The contribution of cholinergic and noncholinergic mechanisms of neurotransmission seems to be independent of the high cholesterol content of the diet and the plasma cholesterol level in rat detrusor muscle.

Involvement of stretch‐induced Rho‐kinase activation in the generation of bladder tone

The present study investigated the role of the Rho-kinase (ROK) pathway in the maintenance of bladder tone during the storage phase, and its effects on bladder compliance. Muscle strips from isolated rat bladder (detrusor strips) were used to evaluate the effects of the ROK inhibitors Y-27632 and HA-1077 on resting tension, which is independent of G-protein coupled pathways. Stretch-induced ROK activation was assessed by measuring phosphorylation of MYPT1 (myosin phosphatase targeting subunit) using Western blotting. The effect of ROK inhibitors on bladder compliance during bladder filling was assessed in an in vitro whole bladder model. Y-27632 and HA-1077 caused concentration-dependent relaxation of detrusor strips. Stretch increased MYPT1-p[Thr853] levels by approximately 1.5-fold in normal Krebs buffer. The ROK inhibitor Y-27632 abolished the stretch-induced increase and reduced the level of MYPT1-p[Thr853] to <50% of the basal values in normal Krebs buffer. Stretch-induced phosphorylation of MYPT1 was independent of Ca(2+) originating from either extracellular or intracellular stores. When tested in the isolated whole rat bladder model, HA-1077 significantly increased bladder compliance at both 3 and 10 µM. This study demonstrates that the ROK pathway is constitutively active and stretch further activates the ROK pathway, which contributes to the generation of bladder tone, thereby affecting bladder compliance.

The novel β3-adrenoceptor agonist mirabegron reduces carbachol-induced contractile activity in detrusor tissue from patients with bladder outflow obstruction with or without detrusor overactivity

β3-Adrenoceptors are major players in detrusor relaxation and have been suggested as a new putative target for the treatment of overactive bladder syndrome. We determined the effects of mirabegron (YM178), a novel β3-adrenoceptor agonist, on carbachol-induced tone in isolated human detrusor preparations from patients with bladder outflow obstruction (BOO) with and without detrusor overactivity (DO), and from patients with normal bladder function. We compared the effects to those of isoprenaline, a non-selective β-adrenoceptor agonist. Detrusor specimens were obtained from patients with benign prostatic hyperplasia undergoing cystoscopy and from patients undergoing radical prostatectomy/cystectomy (in total 33 donors). Detrusor contractility was evaluated by organ bath studies and strips were incubated with carbachol (1μM) to induce and enhance tension. Both mirabegron and isoprenaline reduced carbachol-induced tone in tissues from all groups. Isoprenaline decreased tension with higher potency than mirabegron in normal, BOO and BOO+DO detrusor strips with pIC50 values of 7.49±0.16 vs. 6.23±0.26 (P=0.0002), 6.89±0.34 vs. 6.04±0.31 (P=0.01), and 6.57±0.20 vs. 5.41±0.08 (P<0.0001, n=4), respectively. The maximal relaxant effect of isoprenaline and mirabegron in the normal, BOO and BOO+DO detrusor was 37.7±14.4% and 36.1±23.3%, 14.4±12.2% vs. 33.4±21.0% and 18.3±10.0% vs. 28.3±12.2% (n=4, P>0.05), respectively. Mirabegron and isoprenaline reduced carbachol-induced tone in both normal bladders and obstructed bladder with and without DO. Isoprenaline had higher potency than mirabegron, but the efficacy of mirabegron effect was the same as that of isoprenaline.

Variations in carbachol- and ATP-induced contractions of the rat detrusor: effects of gender, mucosa and contractile direction

Contractile characteristics of the bladder may depend on variables such as gender, mucosa (MU) and direction of the contractions. However, definitive information is not yet available despite earlier studies on the effects of one variable or another. Here, we explored the differences in the rat detrusor attributable to gender, mucosa and contractile direction. K+, carbachol (CCh) and ATP were used as contractile stimuli on rat detrusor strips with and without MU. Contractility was monitored using a myograph system. Both tonic and phasic contractile activities were analyzed. MU-independent contractions induced by CCh were more potent in females, an effect specific to the longitudinal direction only. The maximal CCh response was larger also in females when MU was removed, suggesting a stronger MU-independent component in the contraction. The larger area under curves of the females under ATP stimulation showed dependence on MU and contractile direction as well. ATP-induced contractions in the males were affected more by MU in the transverse direction than in the females. Direction- and MU-dependent variability of ATP responses was also observed in the males but not in females. Findings here added new information to the understanding of bladder contractile physiology, providing insights into the quest for better drugs in managing bladder disorders.