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European Urology
Volume 56, issue 2, pages 237-406, August 2009Voiding Dysfunction
Characteristics of Spontaneous Activity in the Bladder Trigone
Alexander Roosen a d, Changhao Wu b 
, Guiping Sui a, Rasheda A. Chowdhury c, Pravina M. Patel c, Christopher H. Fry a.
Accepted 12 June 2008, Published online 20 June 2008, pages 346 - 354
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Abstract
Background
During bladder filling, the trigone contracts help keep the ureteral orifices open and the bladder neck shut. The trigone generates spontaneous activity as well as responding to neuromuscular transmitters, but the relationship between these phenomena are unclear.
Objectives
To characterise the cellular mechanisms that regulate and modify spontaneous activity in trigone smooth muscle.
Design, setting, and participants
Muscle strips from the superficial trigone of male guinea-pigs were used for tension experiments and immunofluorescent studies.
Measurements
In isolated trigonal cells, intracellular Ca2+ was measured by epifluorescence microscopy using the fluorescent Ca2+ indicator Fura-2.
Results and limitations
Spontaneous intracellular Ca2+ transients and contractions were observed in trigonal single cells and strips and were significantly higher compared to the bladder dome. Ca-free superfusate and verapamil terminated spontaneity. T-type Ca2+ channel block with NiCl2 depressed slightly Ca2+ transients but not spontaneous contractions. Neither the BKCa channel blocker iberiotoxin nor the SKCa channel blocker apamin had any effect on single cell activity. By contrast, the Cl− channel blocker niflumic acid attenuated significantly both Ca2+ transients and muscle contractions. Agonist stimulation (carbachol, phenylephrine) up-regulated activity. Gap junction labelling (Cx43) was approximately 5 times denser in the trigone than in detrusor smooth muscle. The gap junction blocker 18-ß-glycyrrhetinic acid modulated spontaneous contractions in the trigone but not in the bladder dome.
Conclusions
Trigone myocytes employ membrane L-type-Ca2+ channels and Cl− channels to generate spontaneous activity. Intercellular electrical coupling ensures its propagation and, thus, sustains contraction of the whole trigone.
Keywords: Trigone, Spontaneous activity, Connexin43, L-type Ca2+-channels, Cl−-channels, Guinea-pig.
Article Outline
1. Introduction
The trigone has recently attracted urologists’ attention again, initiated by a discussion of whether it should be spared in intradetrusor botulinum toxin injections to treat bladder overactivity [1], and [2]. It is strategically located between the ureteric orifices and bladder outlet; however, little is known about its function during the micturition cycle. The superficial trigone develops, with the ureter, from an outgrowth of the mesonephric duct and provides, as a transverse-orientated interureteric muscle, competent vesico-ureteric anchoring [3], and [4]; it represents an area of dual parasympathetic-muscarinic and sympathetic-adrenergic innervation. There is evidence that micturition is initiated by relaxation of the trigone, which then funnels urine into the proximal urethra; relaxation may be NO/cGMP-mediated through nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase–positive nerves [5], and [6].
However, information about trigone behaviour during filling is sparse, and spontaneous activity might play a crucial role. Various smooth muscles in the lower urinary tract show spontaneous contractile activity during the filling phase. In the bladder dome, this behaviour may help the organ maintain a state of minimal surface area; in the urethra, it contributes significantly to closure pressure [7]. Although spontaneous activity has been intensively investigated in both dome and urethra, studies in the trigone are lacking. Two papers, which primarily investigated agonist effects on detrusor muscle, noted pronounced spontaneous activity of the trigone. Contractile activity was present in 71% and 89% of trigone strips from pigs and humans, respectively, compared to 20% from the dome [8]. Microelectrode recordings showed rhythmic variation of membrane potential in the dome; but in the trigone, there were superimposed additional bursts of spikes [9]. This study also described up-regulation of trigone activity by agonist stimulation.
We set out to corroborate previous reports of spontaneous activity in the trigone, to elucidate its origin and physiological characteristics, and to explore possible routes of modulation at an intra- and intercellular level. Data were compared to those from similar detrusor preparations of the bladder dome.
2. Methods
2.1. Tissue preparation and tension experiments
Tissue was obtained from the bladders of 70 male guinea-pigs (400–499 g, Dunkin-Hartley), killed by Schedule-1 cervical dislocation in accordance with the UK Animals (Scientific Procedures) Act, 1986. The bladder dome was resected cranially to the orifices, the base longitudinally opened on the ventral site, and the trigonal area exposed. The superficial layer was distinguished from underlying detrusor by its paler, whitish appearance. After removing the mucosa by blunt dissection, a thin strip (average: 6.4 mg) between the ureteral orifices was cut; only one strip per bladder was prepared. Strips were also dissected from the dome. Preparations (28 trigone, 21 dome) were mounted in a horizontal superfusion trough between a fixed hook and an isometric force transducer and superfused with a 24 mM-NaHCO3/5%CO2-buffered Tyrode's solution (5–10 ml min−1, 37 °C, pH 7.4). All chemicals throughout were from Sigma unless specified in the text. Transducer output was recorded through a bridge amplifier and displayed on a moving-paper chart recorder.
2.2. Cell isolation, measurement of intracellular calcium ([Ca2+]i)
Cells were prepared from muscle strips (37 animals) as might be used for tension measurements by dissociation with a collagenase-based enzyme mixture [10]. Cells were loaded with the fluorescent Ca2+ indicator Fura-2 (Molecular Probes) at 37 °C for 9 min and stored at 4 °C for later use. An aliquot of cell suspension was placed in a glass-bottomed chamber maintained at 37 °C, mounted on the stage of an inverted microscope. After cells had settled to the chamber floor, it was superfused with Tyrode's at 1.5 ml min−1. Cells were illuminated alternately at 340:380 nm (32 Hz) and fluorescent light collected between 410 and 510 nm with a photomultiplier tube. The ratio of fluorescence on excitation at 340 or 380 nm is a function of [Ca2+]i: Calibration of the signal has been detailed elsewhere [10].
2.3. Connexin immunofluorescence
Five de-urothelialised strips were placed immediately in optimal cutting temperature (OCT) embedding medium and snap-frozen in liquid N2. Triplicate sections (10 μm) for each specimen were cut on a cryostat and mounted on 3-aminopropyltriethoxysilane (APES)-coated slides. Sections were post-fixed in methanol (−20 °C, 5 min), blocked in 1% bovine serum albumin (BSA) for 45 min and incubated with primary antibodies against connexin43 and connexin45 (2 h, room temperature: Cx43, mouse mAB, Chemicon 1:1000; Cx45; gift from Professor N Severs, Imperial College London). Binding was visualised using a Cy3-conjugated secondary antibody (goat, anti-mouse, Chemicon, 1:1000), and nuclei counterstained with 4′,6-diamidino-2-phenylindole (DAPI; Invitrogen, 1:50,000) in the same step (45 min, room temperature [RT]). Three representative images per section were taken using a confocal microscope (Zeiss LSM 510 Meta, x40) at fixed pinhole and detector gain settings. The number of punctate connexin fluorescent particles was counted in each image using the ImageJ freeware program (rsb.info.nih.gov/ij/; constant threshold, particles 2-infinity).
2.4. Experimental protocols and data analysis
Experimental variables obtained during interventions were compared to the mean of pre- and postintervention values (controls) and expressed either as absolute values or percentage of control. Control experiments, using only the vehicle for test agents, were always carried out and exerted no significant effects. Values are mean ± SD, n = number of cells or muscle strips. Differences between means of data sets were examined by paired or unpaired student t tests and those between two incidences by Fisher's exact test. The null hypothesis was rejected at p < 0.05.
3. Results
3.1. Characteristics of spontaneous activity
Spontaneous activity was recorded in trigonal single cells and strips as Ca2+ transients and contractions, respectively. Myocytes were about half the size of those from the bladder dome (Fig. 1A). The resting [Ca2+]i was 45.5 ± 21.1 nM (n = 107 cells), and cells always showed either discrete Ca2+ transients (Fig. 1B) or, less often, more sustained and fused increases of [Ca2+]i (Fig. 1C). Their mean amplitude was 78 ± 34.1 nM. Cells regularly responded with visible contractions to Ca2+ transients (Fig. 1A, B and C: c), demonstrating their functional relevance. Furthermore, the majority of strips generated regular contractions after 45-min equilibration (Fig. 1D)—mean amplitude 5.8 ± 1.5 mN mm−2 (75.3% and 207.1% of force generated by 10 μM phenylephrine [PE] and 1 μM carbachol, respectively). Preparations not generating phasic contractions always displayed uncoordinated baseline changes (Fig. 1E).
00741-0/assets/gr1.jpg)
Fig. 1
(A) Image of a trigonal myocyte before (upper left) and after (lower left) spontaneous contraction compared to a myocyte from the bladder dome (right) (all images on the same scale); (B and C) representative [Ca2+]i recordings of unstimulated trigonal single cells; (D and E) tension recordings from unstimulated strips.
c, contraction parts (A), (B), and (C).
The incidence of activity in the trigone was significantly higher than in the dome. Only eight of 21 strips (38%) from the dome showed spontaneous contractions after 45 min, whereas most trigone strips (19 of 28, 68%) developed them. The mean frequency of Ca2+ transients and contractions was greater in the trigone compared to detrusor: Ca2+ transients 3.2 ± 1.2 vs 0.6 ± 0.5 min−1 (n = 40,6; amplitude 78 ± 34.1 vs 31.4 ± 18.9 nM); contractions 1.8 ± 0.4 vs 1.4 ± 0.2 min−1 (n = 14,6; no significant differences in amplitude).
3.2. Cellular mechanisms for spontaneous activity
Intracellular stores were investigated as a possible source of spontaneous activity. Thapsigargin (1 μM, 20 min, n = 10) and carbonylcyanide-4-(trifluoromethoxy)-phenylhydrazone (FCCP; 4 μM, 20 min, n = 4) were used to block Ca2+ release from the sarcoplasmic reticulum or mitochondria, respectively: however, no alteration to activity was seen (Fig. 2A). By contrast, the L-type-Ca2+ channel blocker, verapamil (20 μM, Knoll) or Ca-free solution (zero-added Ca plus 0.5 mM ethylene glycol tetraacetic acid [EGTA], n = 20) abolished activity completely (Fig. 2A, B). Corresponding spontaneous muscle contractions and baseline tension were reduced by verapamil (20 μM, n = 6; Fig. 2C) and Ca-free solution (n = 4, Fig. 2D).
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Fig. 2
(A) Representative trace showing the effect of thapsigargin (blocking Ca2+ release from the SR), FCCP (blocking mitochondrial Ca2+ release), verapamil (blocking L-type Ca2+ channels) on spontaneous [Ca2+]i transients; (B) effect of Ca2+-free solution on spontaneous [Ca2+]i transients; (C) effect of verapamil on spontaneous contractions; (D) effect of Ca2+-free solution on spontaneous contractions; (E) summary of the effects of NiCl2 and MgCl2 on the amplitude and frequency of spontaneous [Ca2+]i transients—data as % of control, mean ± SD, *p < 0.05.
T-type-Ca2+ channels may facilitate the opening of L-type channels, as the former are activated at more negative potentials. Superfusion with 100 μM NiCl2, to block selectively T-type channels attenuated significantly the frequency and amplitude of Ca2+ transients: frequency 4.2 ± 1.7 to 3.3 ± 1.5 min−1 (77.2 ± 16.7% control); amplitude 70 ± 29.3 to 53.2 ± 18.8 nM (78.2 ± 14% control, n = 5)—Fig. 2E. However, there was no corresponding effect on muscle strips (n = 4).
Magnesium stabilises various smooth muscle tissues by attenuating L-type Ca2+ current [11]. MgCl2 (2 mM, 5 min, n = 5) reduced Ca2+ transient frequency (4.1 ± 1.1 to 2.7 ± 0.9 min−1 [69 ± 21% control]) and amplitude (59.1 ± 32.2 to 41.1 ± 31 nM, [65.7 ± 14.8% control])—Fig. 2E—without a significant effect on muscle strips.
In detrusor myocytes, large conductance Ca2+-activated K+-channels (BKCa) are mainly responsible for repolarisation of spontaneous electrical activity, and small conductance K+-channels (SKCa) have a modulatory role. Neither the BKCa-channel blocker iberiotoxin (50 nM) nor the SKCa-channel blocker apamin (100 nM) had any effect on isolated trigonal cells when applied for up to 20 min (n = 8 and 9, respectively; data not shown).
Cl− channels can generate spontaneous Ca2+ transients in distinct cell populations of the lower urinary tract (eg, urethral and bladder interstitial cells) [12], and [13]. With isolated trigonal cells, the Cl− channel blocker niflumic acid (100 μM) attenuated the frequency (3.2 ± 1.2 to 8 ± 1.2 min−1 [55.9 ± 44.2% control, n = 7]) and amplitude (79.6 ± 32.9 to 31.9 ± 21.1 nM [50.5 ± 14.23% control, n = 7], Fig. 3A and E) of Ca2+ transients. Muscle contractions were similarly reduced in frequency and amplitude (1.7 ± 0.2 to 1.1 ± 0.3 min−1 [66 ± 24.1% control] and 6.9 ± 1.2 to 3.7 ± 1.5 mN mm−2 [52.7 ± 19% control], n = 5, Fig. 3C and F).
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Fig. 3
(A) Representative trace showing the effect of the Cl− channel blocker niflumic acid on spontaneous [Ca2+]i transients; (B) effect of low Cl− solution on spontaneous [Ca2+]i transients—note the interruption of the trace during the low Cl− intervention; (C) effect of niflumic acid on spontaneous contractions; (D) effect of low Cl− solution on spontaneous contractions; (E and F) summary of the effects of niflumic acid and low Cl− solution on the amplitude and frequency of spontaneous [Ca2+]i transients (E) and spontaneous contractions (F)—data as % of control, mean ± SD, *p < 0.05.
Reducing extracellular Cl− shifts the reversal potential of a Cl− current to more positive potentials, depolarises cells, and should increase spontaneous activity. Extracellular Cl− was reduced from 127.6 to 10.7 mM by equimolar replacement of NaCl with Na isethionate, whilst maintaining constant Ca2+ activity by raising extracellular CaCl2 from 1.8 to 2.34 mM [14]. Ca2+ transient frequency and amplitude in isolated cells were increased (4.0 ± 1.4 to 7.4 ± 2.3 min−1 [188 ± 26.8% control] and 35.6 ± 15.5 to 98.4 ± 51.5 nM [218.6 ± 71.6% control, n = 5], Fig. 3B and F). In strips, contraction amplitude was similarly augmented (6.7 ± 1.1 to 9.6 ± 1.6 mN mm−2 [146.6 ± 35.4% control, n = 5], Fig. 3D and F), although frequency was not significantly altered (1.6 ± 0.3 min−1 vs 1.9 ± 0.5 min−1).
3.3. Modulation of trigonal spontaneity
Adrenergic and muscarinic receptor stimulation increases spontaneous activity in detrusor myocytes [9], and [15]. Trigone myocytes were exposed to 10 μM PE and 1 μM carbachol for 1 min (concentrations exerting intermediate inotropic effects on muscle strips; see also Roosen et al [16]). Both interventions significantly increased the frequency and amplitude of Ca2+ transients (PE: frequency 1.9 ± 0.6 to 3.9 ± 1.4 min−1 [210.4 ± 49% control], amplitude 105.7 ± 56.2 to 131.8 ± 53.8 nM [141.1 ± 50%, n = 9]; carbachol: frequency 1.5 ± 0.5 to 3.3 ± 0.8 min−1 [221.9 ± 57.2% control], amplitude 55 ± 30.9 to 73.4 ± 32.9 nM [149.1 ± 61.9% control, n = 9]), as shown in Fig. 4A and B. In muscle strips, spontaneous activity normally ceased with the development of agonist-induced contractions.
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Fig. 4
(A) Representative trace showing the effect of 10 μM phenylephrine (PE, 1 min) and 1 μM carbachol (carb, 1 min) on spontaneous [Ca2+]i transients; (B) summary of the effects of 10 μM PE and 1 μM Carb on the amplitude and frequency of spontaneous [Ca2+]i transients (measured 3 min after intervention); data as % of control, mean ± SD; *p < 0.05.
Gap junctions electrically couple adjacent cells and are formed from different isoforms of the connexin (Cx) protein family. In the trigone and detrusor, we found labelling for Cx43 (Fig. 5A and B). However, labelling was more intense in the trigone (146.8 ± 64.8 vs 34.7 ± 14.8 punctate regions per 230 × 230-μm section, n = 4, Fig. 5E). Moreover, in the trigone, labelling was more evenly distributed, whereas in the detrusor it was strictly confined to a chain-like distribution associated with cells that had smaller, long nuclei between muscle bundles (arrows, Fig. 5A). No labelling for Cx45 was observed in the trigone, as has been recorded in the detrusor between adjacent smooth muscle cells.
00741-0/assets/gr5.jpg)
Fig. 5
(A and B) Representative sections (10 μm) through a muscle preparation from the bladder dome (A) and the trigone (B), labelling for connexin 43 (red) and nuclei (blue)—note the sparse, chain-like distribution of Cx43 in the detrusor along a band of spindle-shaped nuclei, presumably interstitial cells (yellow arrows); (C and D) representative tension recordings showing the effect of gap junction-blocker 18-ß-glycyrrhetinic acid (18-ß-GA, 10 μM) on spontaneous contractions of strips from the bladder dome (C) and the trigone (D); (E) summary of gap junction counts per 230 × 230-μm field in the detrusor and trigone; (F) summary of the effect of 18-ß-GA on the amplitude and frequency of spontaneous contractions in strips from the bladder dome and trigone; data as % of control, mean ± SD; *p < 0.05.
To test the significance of gap junction proteins for spontaneous contractions in trigone and detrusor strips, we perfused them with the gap junction blocker 18-ß-glycyrrhetinic acid (10 μM, 18-ß-GA) for 5 min. Detrusor strips were not significantly affected (frequency: 1.3 ± 0.3 vs 1.4 ± 0.2 min−1; amplitude: 5.6 ± 0.1 vs 5.6 ± 0.8 mN mm−2, n = 5, Fig. 5C and F), However, trigonal contractions became significantly more frequent (2.1 ± 0.3 vs 4.8 ± 1.1 min−1 [224.9 ± 60% control]) and smaller (5.7 ± 1.1 vs 3.6 ± 0.6 mN mm−2 [64.3 ± 9.5%, n = 5], Fig. 5D and F). In addition, even smaller contractions appeared between larger ones, generating a less-organised contractile pattern. 18-ß-GA had no effect on Ca2+ transients in trigonal single cells (n = 4).
4. Discussion
This is the first study to investigate the nature and modulation of spontaneous activity in the trigone. The Ca2+ transients were comparable to those generated by receptor agonists of the major contractile neurotransmitters PE and carbachol, and thus represent significant events that could support cellular contraction. Cells always responded to both PE and carbachol which differentiated them from detrusor myocytes (respond to carbachol alone) or urethral myocytes (respond to PE alone). The general harmony of observations with isolated cells and muscle strips suggests that Ca2+ transients in trigone myocytes are the precursor of spontaneous contractions. The source of Ca2+ for intracellular transients is probably through L-type Ca2+ channels, as reducing extracellular Ca, the addition of verapamil, or raising extracellular Mg all attenuated activity. However, blocking Ca2+ release from internal sources had no effect. T-type channels might also contribute, as they are activated at more negative potentials [17] and so facilitate L-type channel opening. Of interest, Cl− channels also modulated spontaneous activity, as evidenced by their attenuation with niflumic acid or enhancement by reducing extracellular Cl−. However, blockade of BKCa or SKCa channels had no effect, which implies that they do not have predominant effects on membrane activity. Furthermore, as trigone myocytes respond to both adrenergic and muscarinic receptor agonists by increasing force [16] and intracellular Ca2+, both agonists also increased significantly spontaneous activity, although the cellular pathways remain to be elucidated. Practically, monitoring changes to Ca2+ transients in isolated cells was a more sensitive index than changes to spontaneous contractions. Most likely, this is because the latter are a summation of asynchronous behaviour in many individual cells so that relatively uncoordinated changes in myocytes would manifest themselves less distinctly in multicellular preparations.
The cellular origin of spontaneous activity in the trigone shows differences from other lower urinary tract cells that demonstrate similar activity. In the detrusor, spontaneous contractile activity is associated with action potentials whose upstroke is carried by L-type Ca2+ current and repolarised largely by BKCa channel activity [7], [15], and [18]. Transmembrane Ca2+ fluxes may be supplemented by Ca2+ release from internal stores, which are replenished through a feedback mechanism employing BKCa and L-type Ca2+ channels [10]. In other regions of the urinary tract, such as the renal pelvis and urethra, electrically active atypical smooth muscle cells [19] or interstitial cells [12] may drive or modulate spontaneous activity in true muscle cells. Where studied, they operate through a combination of Ca2+ release from internal stores and opening of depolarising Ca2+-activated Cl− channels [20], and Cl− channel blockers have a marked depressant effect [12], and [13]. However, K+ channels have not been reported to be of particular importance, and Ca2+ channels are of lower density than in true muscle cells. Both spontaneous [Ca2+]i rises and Ca2+-activated Cl− channels are characteristic of suburothelial myofibroblasts that also have little L-type Ca2+ current [21]. Therefore, trigonal myocytes seem to have physiological characteristics intermediate between detrusor myocytes and other cells of the urinary tract, including interstitial cells or atypical renal pelvis cells.
The difference between the trigone and the bladder dome was also reflected in gap junction make-up and distribution. The trigone throughout was characterised by Cx43 labelling, whereas this was confined to labelling between muscle bundles in the detrusor (see also Ikeda et al [22]). Cx45 was not detected in the trigone but comprises intermuscular connections in the detrusor [23]. The dense distribution of trigonal Cx43 is in line with an electron-microscopic (EM) study in humans suggesting marked electrical coupling in the trigone [24]. The relevance to spontaneous muscle contractions was demonstrated by the effects of the gap junction blocker 18-ß-GA, which reduced the amplitude but increased the frequency of spontaneous contractions. Such a pattern might be expected from reduced intercellular coupling so that spontaneous activity originating in individual cells would be less well co-ordinated throughout the tissue mass. The lack of effect of 18-ß-GA on detrusor strips is consistent with a lower gap junction density, so that spontaneous activity must already be poorly co-ordinated, and is consistent with 18-ß-GA being ineffective on spontaneous activity of normal rat bladder [22]. It can be hypothesised, therefore, that electrical coupling in the trigone is better than in the detrusor, although still less than in extensively coupled tissues such as myocardium.
The relative difference of electrical coupling will have physiological consequences. Poorer coupling in the bladder dome prevents extensive propagation of electrical signals and allows individual muscle bundles to adjust their length in response, say, to a volume change without synchronous activation of the muscle mass that would elevate intravesical pressure (“micromotions,” [7]). The trigone is believed to contract during the filling phase, thus contributing to opening the ureteric orifices and closing the bladder outlet. The high spontaneous activity and extensive electrical coupling may help the trigone adopt a state of sustained contracture during urine storage, augmented by neurotransmitter stimulation.
Further structural and functional studies should reveal whether trigonal muscle cells display a phenotype more reminiscent of bladder smooth muscle or interstitial cells or even ureter. Homogenous positive labelling for alpha smooth muscle actin (α-SMA) has been shown [4], and [25], but labelling for c-kit and vimentin—which have both been proposed as markers for interstitial cells—in the lower urinary tract [26], and [27] would be valuable. Furthermore, the ability of verapamil to abolish spontaneous activity in the ureter [28] was mirrored here, and further comparison between these tissues would be useful. Finally, the present study focused on intrinsic tension generation in the trigone, but an investigation of relaxing mechanisms would also be valuable. NO has been demonstrated as a key factor [6]. Whether it suppresses spontaneous depolarisations of the membrane through the NO/cGMP/PKG pathway as in functionally related interstitial cells of the urethra [29] or lowers Ca2+ sensitivity of the contractile machinery is unknown.
5. Conclusion
Trigonal myocytes show marked spontaneous activity in the form of [Ca2+]i transients resulting from transmembrane Ca2+ influx through L-type Ca2+ channels. As with lower urinary tract interstitial cells, Cl− channels rather than K+ channels also contribute to the generation of spontaneity. Extensive gap junction coupling ensures electrical propagation and sustained spontaneous contraction of the whole trigone, thus contributing to its proposed physiological functions.
Author contributions: Alexander Roosen, Changhao Wu and Christopher H. Fry had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy take of the data analysis.
Study concept and design: Roosen, Sui, Wu, Fry.
Acquisition of data: Roosen.
Analysis and interpretation of data: Roosen, Wu, Fry.
Drafting of the manuscript: Roosen.
Critical revision of the manuscript for important intellectual content: Wu, Fry.
Statistical analysis: Roosen, Wu.
Obtaining funding: Roosen, Wu, Fry.
Administrative, technical, or material support: Chowdhury, Patel.
Supervision: Sui, Wu, Fry.
Other (specify): none.
Financial disclosures: I certify that all conflicts of interest, including specific financial interests and relationships and affiliations relevant to the subject matter or materials discussed in the manuscript (eg, employment/affiliation, grants or funding, consultancies, honoraria, stock ownership or options, expert testimony, royalties, or patents filed, received, or pending), are the following: none.
Funding/Support and role of the sponsor: A. Roosen is recipient of a grant of the German Research Foundation. Wellcome Trust and British Heart Foundation provided financial support.
Acknowledgement statement: We acknowledge the support of the Deutsche Forschungsgemeinschaft, the Wellcome Trust, and the British Heart Foundation who have no commercial interest in the data.
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Footnotes
a Department of Surgery, University College London, London, United Kingdom
b Department of Medicine, University College London, London, United Kingdom
c Department of Cardiac Electrophysiology, National Heart and Lung Institute, Imperial College, London, United Kingdom
d Department of Urology, Ludwig-Maximilians-University, Muenchen, Germany
Corresponding author. Postgraduate Medical School, Faculty of Health and Medical Sciences, University of Surrey, Guildford GU2 7WG, UK. Tel. +44 1483 682517.
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Comments
Thank you for this valid study. 1-Findings enclosed are contraductorial to Tanagho findings. He noted that antireflux mechanism of ureteral orifice depends mainly on spasm of trigone. Its relaxation leads to reflux. 2-Filling of bladder depends mainly on the overcome force exerted on ureteral orifice by propagation of peristalsis waves of the ureter. That leads to a jet-like effect of urine passage to bladder. 3-Evidence : Cystoscopic observation of ureteral orifice during bladder filling. Rythmic (Open and Close) of ureteral oifice were found, that coinciding with peristalsis waves. Best Regards. Dr.Mohamed Essam Abdalla Ahmed Urology Consultant-Riyadh-Saudi Arabia Member of EAU
2009-08-05 03:27:08 | Mohamed Essam Abdalla Ahmed