Reflex Bladder Research Articles

Effects of intrathecal injection of a hyperpolarization‐activated channel (Ih) inhibitor ZD7288 on bladder function in urethane‐anesthetized rats

The role of I(h) channels at the spinal level in the control of bladder function was examined in urethane anesthetized female rats. Bladder activity was recorded via a transurethral catheter under isovolumetric conditions or via an intravesical catheter during continuous infusion cystometry. ZD7288, a selective I(h) channel inhibitor, was administered intrathecally at L6-S1 segmental levels of the spinal cord. Under isovolumetric conditions, intrathecal 0.3 microg of ZD7288 was inactive, whereas 1 and 3 microg abolished bladder activity for 5.6 +/- 1.8 and 20.7 +/- 3.9 min, respectively. Three micrograms also tended to decrease the amplitude of the contractions when they reappeared. During continuous cystometry, an intrathecal injection of ZD7288 at 0.3-3 microg dose-dependently increased the intercontraction intervals by 7.5%, 33.1%, and 57.5% from pre-drug values. A higher dose of ZD7288 (3 microg) tended to increase the pressure threshold for inducing micturition by 16.6% and the maximum voiding pressure by 11.1%. On the other hand, cardiovascular parameters such as heart rate and mean blood pressure were not affected by 0.3-3 microg of intrathecal ZD7288. These results indicate that I(h) channels play an important role in the control of micturition. Because I(h) channel inhibition in the lumbosacral spinal cord reduced the frequency of the micturition reflex without significantly affecting the amplitude of reflex bladder contraction, I(h) channels might preferentially be involved in afferent processing in the spinal cord to control the micturition reflex.

The discovery of the pontine micturition centre by F. J. F. Barrington

The discovery of the pontine micturition centre by F. J. F. Barrington

Bladder activation by selective stimulation of pudendal nerve afferents in the cat

Bladder contractions evoked by pudendal nerve stimulation in both spinal intact and spinal transected cats support the possibility of restoring urinary function in persons with chronic spinal cord injury (SCI). However, electrically evoked bladder responses in persons with SCI were limited to transient contractions at relatively low pressures. This prompted the present study, which presents a detailed quantification of the responses evoked by selective stimulation of individual branches of the pudendal nerve at different stimulation frequencies. In spinal intact cats anesthetized with α-chloralose, selective frequency-dependent electrical activation of the sensory (2 Hz ≤ f ≤ 50 Hz), cranial sensory ( f ≤ 5 Hz), dorsal genital ( f ≥ 20 Hz) and rectal perineal ( f ≤ 10 Hz) branches of the pudendal nerve evoked sustained bladder contractions dependent on the stimulation frequency. Contractions evoked by selective electrical stimulation resulted in significant increases in voiding efficiency compared to bladder emptying by distension-evoked contractions ( p ANOVA < 0.05). Acute spinal transection abolished reflex bladder contractions evoked by low frequency stimulation of the cranial sensory or rectal perineal branches, whereas contractions evoked by high frequency stimulation of the dorsal genital branch remained intact. This study presents evidence for two distinct micturition pathways (spino-bulbo-spinal vs. spinal reflexes) activated by selective afferent pudendal nerve stimulation, the latter of which may be applied to restore bladder function in persons with SCI.

Bladder cooling reflex and external urethral sphincter activity in the anesthetized and awake guinea pig

A spinal bladder cooling reflex, triggered by cold receptors of transient receptor potential melastatin type in the bladder wall, has been identified in several mammals, including man. This reflex and its influence on the external urethral sphincter were further characterized in the urethane anesthetized and awake guinea pigs. A total of 214 bladder infusions were performed in the 12 animals. Compared to controls, cold fluid induced a significant decrease in the threshold volume for reflex bladder contractions (median 82%, p < 0.01). Menthol induced a further decrease (median 50%), signifying a bladder cooling reflex. Detrusor-sphincter activities were dyssynergic during voidings triggered by cold or menthol infusions but were coordinated during control infusions. The bladder cooling reflex was suppressed and the sphincter activity synergic following cold infusions in the awake state. Thus, the bladder cooling reflex is under the active descending inhibitory control in intact, awake animals.

Re-established micturition reflexes show differential activation patterns after lumbosacral ventral root avulsion injury and repair in rats

Previous studies have demonstrated that an acute implantation of lesioned lumbosacral ventral roots into the rat conus medullaris (CM) results in functional reinnervation of the lower urinary tract (LUT). Although the root implantation procedure results in a return of reflexive micturition, voiding efficiency (VE) remains incompletely recovered. Here, we performed a detailed urodynamic analysis of cystometry and external urethral sphincter (EUS) electromyography (EMG) recordings to determine underlying mechanisms for the incompletely recovered VE. For this purpose, adult female rats were studied at 12 weeks after a bilateral L5–S2 ventral root avulsion injury followed by an acute surgical implantation of the avulsed L6 and S1 ventral roots into the CM ( n = 6). Age-matched sham-operated rats ( n = 6) were included for control purposes. Compared to sham-operated controls, rats of the implanted series showed 1) reflex bladder contractions with a significantly shortened urine expulsion phase, 2) markedly decreased phasic EUS EMG activity during micturition, and 3) a pronounced bladder–sphincter dys-coordination, as demonstrated by a significantly delayed onset of the switch from low-amplitude tonic EUS EMG activity to either phasic EUS EMG activity or a large-amplitude tonic EUS EMG activity during the urine expulsion phase. Our findings provide a mechanistic explanation for the incomplete recovery of the VE following implantation of avulsed ventral roots into the spinal cord. Our future studies will aim to increase successful axonal regeneration in attempts to augment the recovery of the LUT after cauda equina injury and repair.

Variable Patterned Pudendal Nerve Stimuli Improves Reflex Bladder Activation

We evaluated variable patterns of pudendal nerve (PN) stimuli for reflex bladder excitation. Reflex activation of the bladder has been demonstrated previously with 20-33 Hz continuous stimulation of PN afferents. Neuronal circuits accessed by afferent mediated pathways may respond better to physiological patterned stimuli than continuous stimulation. Unilateral PN nerve cuffs were placed in neurologically intact male cats. PN stimulation (0.5-100 Hz) was performed under isovolumetric conditions at bladder volumes up to the occurrence of distension evoked reflex contractions. Stimulus evoked reflex bladder contractions were elicited in eight cats. Across all experiments, bursting of 2-10 pulses at 100-200 Hz repeated at continuous stimulation frequencies evoked significantly larger bladder responses than continuous (single pulse) stimulation (52.0+/-44.5%). Bladder excitation was also effective at 1 Hz continuous stimuli, which is lower than typically reported. Variable patterned pulse bursting resulted in greater evoked reflex bladder pressures and increased the potential stimulation parameter space for effective bladder excitation. Improved bladder excitation should increase the efficacy of neuroprostheses for bladder control.

Case Study: Rating Syringomelia by Fifth Edition

Case Study: Rating Syringomelia by Fifth Edition

Effective Treatment of Neurogenic Detrusor Dysfunction by Combined High-Dosed Antimuscarinics without Increased Side-Effects

ObjectivesPatients with neurogenic bladder dysfunction demonstrate an insufficient treatment outcome under dosage-escalated monotherapy. With the objectives of continence and normalised bladder pressure, safe and tolerable non-invasive treatment alternatives were evaluated by using combined antimuscarinics. MethodsTwenty-seven patients who were previously registered in a doubled antimuscarinics study were enrolled in this study. The patients demonstrated urodynamic-proven neurogenic bladder dysfunction with incontinence, reduced bladder capacity, and increased intravesical pressure, resulting from spinal cord injury (n=21); spinal cord dysplasia (myelomeningocele; n=3); multiple sclerosis (n=2), and viral encephalomyelitis (n=1). On the basis of the initial study treatment, they were allocated into three groups and treated with two antimuscarinics. Before enrolment, at 4 wk, and at 6 mo, patients underwent urodynamics and recorded bladder diaries, including side-effects. ResultsIn all three groups, significant changes were noted at the 4-wk follow-up. Incontinence events decreased from an average of 7 to 1 event per day. The average median bladder capacity (180–393ml) and reflex volume (125–335ml) increased; detrusor compliance also improved (average, 15–33ml/cm H2O). Seven patients reported side-effects; two discontinued the successful treatment. Two other patients did not reach satisfactory amelioration of the detrusor dysfunction. ConclusionWith combined high-dosage antimuscarinic medications, 85% of the patients who previously demonstrated unsatisfactory outcome with dosage-escalated monotherapy were treated successfully. The appearance of side-effects was comparable to that of normal-dosed antimuscarinics. Further studies are required to investigate the long-term pharmacological and physiological background of our findings.

Inhibitory and excitatory perigenital-to-bladder spinal reflexes in the cat

This study revealed that in awake chronic spinal cord-injured (SCI) cats reflexes from perigenital skin area to the bladder can be either inhibitory or excitatory. Electrical perigenital stimulation at frequencies between 5 and 7 Hz significantly inhibited large-amplitude rhythmic reflex bladder activity, whereas frequencies between 20 and 40 Hz induced large-amplitude bladder contractions even at low bladder volumes when reflex bladder activity was absent. Both inhibitory and excitatory effects were enhanced as the stimulation intensity increased (5-30 V, 0.2-ms pulse width). During cystometrograms, the inhibitory stimulation (7 Hz) significantly increased the micturition volume threshold 35 +/- 13% above the control volume, while the excitatory stimulation (30 Hz) significantly reduced the threshold 21 +/- 3%. Mechanical perigenital stimulation applied by repeated light stroking of the perigenital skin with a cotton swab only induced an excitatory effect on the bladder. Both electrical and mechanical perigenital stimuli induced large-amplitude (>30 cm H(2)O) bladder contractions that were relatively consistent over a range of bladder volumes (10-90% of the capacity). However, the excitatory electrical stimulation only induced bladder contractions lasting on average 42.2 +/- 3.9 s, but the mechanical stimulation induced bladder contractions that lasted as long as the stimulation continued (2-3 min). Excitatory electrical or mechanical perigenital stimulation also induced poststimulus voiding. The ability to either inhibit or excite the bladder by noninvasive methods could significantly transform the current clinical management of bladder function after SCI.

Comparison of the effects of complete and incomplete spinal cord injury on lower urinary tract function as evaluated in unanesthetized rats

In rats, phasic external urethral sphincter (EUS) activity (bursting) is postulated to be crucial for efficient voiding. This has been reported to be lost after spinal cord transection (txSCI), contributing to impaired function. However, anesthesia may confound evaluating EUS activity. We therefore evaluated urodynamic parameters in unanesthetized, restrained rats and compared the effects of txSCI to that of a clinically relevant, incomplete, contusive injury (iSCI) on lower urinary tract function. Adult female rats were subjected to txSCI or standardized iSCI at the T8 vertebral level. As expected, all injured rats were initially unable to void but developed a reflex bladder with time, with iSCI rats recovering more rapidly than txSCI rats. LUT function was evaluated urodynamically at 2 and 6 weeks after injury. In response to infusion of saline into the bladder, controls consistently exhibited coordinated contraction of the bladder and activation of the EUS in a phasic pattern and had a high voiding efficiency (86.4 ± 2.5%). Voiding efficiency of iSCI rats was reduced to approximately 57% and txSCI rats to approximately 32%. However, two different patterns of EUS activity during voiding were present in both txSCI and iSCI groups at both time points: (1) rats with phasic EUS activity, similar to controls and (2) those that only exhibited tonic EUS activity during voiding. The former had more normal voiding efficiencies. Thus, phasic EUS activity and the improved voiding efficiency associated with it can occur and can be detected in unanesthetized rats after both incomplete and complete SCI.

Bladder inhibition or voiding induced by pudendal nerve stimulation in chronic spinal cord injured cats

To investigate pudendal-to-bladder spinal reflexes in chronic spinal cord injured (SCI) cats induced by electrical stimulation of the pudendal nerve. Bladder inhibition or voiding induced by pudendal nerve stimulation at different frequencies (3 or 20 Hz) was studied in three female, chronic SCI cats under alpha-chloralose anesthesia. Voiding induced by a slow infusion (2-4 ml/min) of saline into the bladder was very inefficient (voiding efficiency=7.3%+/-0.9%). Pudendal nerve stimulation at 3 Hz applied during the slow infusion inhibited reflex bladder activity, and significantly increased bladder capacity to 147.2+/-6.1% of its control capacity. When the 3-Hz stimulation was terminated, voiding rapidly occurred and the voiding efficiency was increased to 25.4+/-6.1%, but residual bladder volume was not reduced. Pudendal nerve stimulation at 20 Hz induced large bladder contractions, but failed to induce voiding during the stimulation due to the direct activation of the motor pathway to the external urethral sphincter. However, intermittent pudendal nerve stimulation at 20 Hz induced post-stimulus voiding with 78.3+/-12.1% voiding efficiency. The voiding pressures (39.3+/-6.2 cmH2O) induced by the intermittent pudendal nerve stimulation were higher than the voiding pressures (23.1+/-1.7 cmH2O) induced by bladder distension. The flow rate during post-stimulus voiding induced by the intermittent pudendal nerve stimulation was significantly higher (0.93+/-0.04 ml/sec) than during voiding induced by bladder distension (0.23+/-0.07 ml/sec). This study indicates that a neural prosthetic device based on pudendal nerve stimulation might be developed to restore micturition function for people with SCI.

Minimally‐invasive electrical stimulation of the pudendal nerve: A pre‐clinical study for neural control of the lower urinary tract

Electrical stimulation of afferent pudendal nerve fibers can evoke sustained bladder contractions (SBC) in cats, yet evidence of therapeutic efficacy in human subjects is lacking. This pre-clinical study was undertaken to test the hypothesis that robust bladder contractions can be generated with a minimally-invasive needle electrode. In seven adult cats, triggered electromyographic (EMG) signals from the external anal sphincter (EAS) were used to minimize the needle-to-nerve distance; while reflex bladder contractions were recorded as 20-sec trains of current pulses of varying amplitude (threshold to 10 mA) and frequency (1-100 Hz) were applied to the nerve. This stimulation paradigm was repeated at successively greater needle-to-nerve distances (0.5 cm intervals) and also at different electrode positions along the nerve. Electrophysiological access to the pudendal nerve was consistently achieved, as indicated by the average threshold for EAS activation (0.31+/-0.19 mA). Using different combinations of stimulus amplitude and frequency, robust SBCs were evoked in every experiment. More rostral electrode positions exhibited stimulation amplitudes and corresponding maximum bladder pressures (0.68+/-0.36 mA and 25.3+/-3.5 cmH2O, respectively) that were comparable to those of more invasive stimulation methods. The needle electrode provides a minimally-invasive approach that will enable the study of reflexes mediated by pudendal afferents in humans, and allow pre-operative testing before implanting a permanent device.

Voiding reflex in chronic spinal cord injured cats induced by stimulating and blocking pudendal nerves

To induce efficient voiding in chronic spinal cord injured (SCI) cats. Voiding reflexes induced by bladder distension or by electrical stimulation and block of pudendal nerves were investigated in chronic SCI cats under alpha-chloralose anesthesia. The voiding efficiency in chronic SCI cats induced by bladder distension was very poor compared to that in spinal intact cats (7.3 +/- 0.9% vs. 93.6 +/- 2.0%, P < 0.05). In chronic SCI cats continuous stimulation of the pudendal nerve on one side at 20 Hz induced large amplitude bladder contractions, but failed to induce voiding. However, continuous pudendal nerve stimulation (20 Hz) combined with high-frequency (10 kHz) distal blockade of the ipsilateral pudendal nerve elicited efficient (73.2 +/- 10.7%) voiding. Blocking the pudendal nerves bilaterally produced voiding efficiency (82.5 +/- 4.8%) comparable to the efficiency during voidings induced by bladder distension in spinal intact cats, indicating that the external urethral sphincter (EUS) contraction was caused not only by direct activation of the pudendal efferent fibers, but also by spinal reflex activation of the EUS through the contralateral pudendal nerve. The maximal bladder pressure and average flow rate induced by stimulation and bilateral pudendal nerve block in chronic SCI cats were also comparable to those in spinal intact cats. This study shows that after the spinal cord is chronically isolated from the pontine micturition center, bladder distension evokes a transient, inefficient voiding reflex, whereas stimulation of somatic afferent fibers evokes a strong, long duration, spinal bladder reflex that elicits efficient voiding when combined with blockade of somatic efferent fibers in the pudendal nerves.

Effect of 5-Hydroxytryptamine 1 Serotonin Receptor Agonists on Noxiously Stimulated Micturition in Cats With Chronic Spinal Cord Injury

The serotonin 5-hydroxytryptamine(1A/7) receptor agonist (R)-8-OH-DPAT (8-hydroxy-2-(di-n-propylamino)tetralin) (Sigma) and the 5-hydroxytryptamine(1A/1B/1D) agonist GR-46611 (3-[3-(2-dimethylaminoethyl)-1H-indol-5-yl]-N-(4-methoxybenzyl)acrylamide) (Tocris Cookson, Ellisville, Missouri) inhibit bladder activity during saline infusion into the bladder of cats with chronic spinal cord injury (saline infused, spinal cord injured cats), suggesting an effect on mechanosensitive bladder afferent C fibers or their targets. We investigated the effects of (R)-8-OH-DPAT and GR-46611 on bladder activity in chronic spinal cord injured cats during infusion of dilute acetic acid into the bladder to stimulate chemosensitive bladder afferent C fibers (acid infused, spinal cord injured cats). Chloralose anesthetized, spinal cord injured cats were catheterized through the bladder dome for filling cystometry during 0.5% acetic acid infusion. Dose-response curves for (R)-8-OH-DPAT (0.3 to 30 microg/kg intravenously) or GR-46611 (0.03 to 300 microg/kg intravenously) were followed by the 5-hydroxytryptamine(1A) antagonist WAY-100635 (N-tert-butyl-3-(4-(2-methoxyphenyl)-piperazin-1-yl)-2-phenylpropanamide) (Sigma) (300 microg/kg). Threshold volume, bladder capacity, residual volume, micturition volume and arterial pressure were measured and external urethral sphincter electromyogram was recorded. Acid infused, spinal cord injured cats responded to (R)-8-OH-DPAT but not to GR-46611 with dose dependent increases in threshold volume, capacity and residual volume (significant above 3 microg/kg). Effects of (R)-8-OH-DPAT were largely reversed by WAY-100635. Neither (R)-8-OH-DPAT nor GR-46611 augmented external urethral sphincter electromyogram activity. Based on differences in the response to GR-46611 in saline vs acid infused, spinal cord injured animals it is tempting to speculate that 2 distinct populations of bladder afferent C fibers (1 chemosensitive and 1 mechanosensitive) can initiate spinal bladder reflexes. Because 5-hydroxytryptamine(1A) receptor agonists increased bladder capacity under saline or acid infused conditions, they are promising candidates for decreasing bladder hyperactivity and increasing bladder capacity in patients with chronic spinal cord injury.

Pudendal nerve stimulation evokes reflex bladder contractions in persons with chronic spinal cord injury

Although electrical stimulation of the pudendal nerve has been shown to evoke reflex micturition-like bladder contractions in both intact and spinalized cats, there is little evidence to suggest that an analogous excitatory reflex exists in humans, particularly those with spinal cord injury (SCI). We present two cases where electrical activation of pudendal nerve afferents was used to evoke excitatory bladder responses. A percutaneously placed catheter electrode was used to electrically stimulate the pudendal nerve trunk in two males with SCI. The response was quantified with recorded changes in detrusor pressure and EMG activity of the external anal sphincter. In both individuals, frequency specific (f = 20-50 Hz) activation of the pudendal nerve trunk evoked excitatory bladder contractions that also depended on the stimulus amplitude and bladder volume. The results suggest that selective activation of the perineal branches of the pudendal nerve may further augment the excitatory reflex evoked by electrical stimulation.

Neural Upregulation in Interstitial Cystitis

Interstitial cystitis (IC) is a syndrome of bladder hypersensitivity with symptoms of urgency, frequency, and chronic pelvic pain. Although no consensus has been reached on the underlying cause of IC, several pathophysiologic mechanisms, including epithelial dysfunction, mast cell activation, and neurogenic inflammation, have been proposed. Despite multiple different causes of urinary cystitis, the bladder's response to cystitis is limited and typical. Animal experiments have shown upregulation of proteinase-activated receptors, tryptase, beta-nerve growth factor, inducible nitric oxide synthase, nuclear transcription factor-kappaB, c-Fos, phosphodiesterase 1C, cyclic adenosine monophosphate (cAMP)-dependent protein kinase, and proenkephalin B. After the noxious stimulus has abated, downregulation of genes appears to follow. Distention of the bladder results in the release of adenosine triphosphate (ATP) from urothelial cells, which activates purinergic P2X3 receptors. Activation by ATP of P2X3-expressing afferents is a fundamental signaling factor in bladder sensation and appears to play a role in bladder reflexes. Fos proteins present in spinal cord neurons have been shown to be upregulated in animals that have undergone cyclophosphamide-induced chemical cystitis. These and other findings suggest that neural upregulation occurs both peripherally and centrally in subjects with chronic cystitis. It is unclear whether neural mechanisms and inflammation are the cause of IC or the result of other initiating events. Neural upregulation is known to play a role in the chronicity of pain, urgency, and frequency and represents an exciting area of research that may lead to additional treatments and a better understanding of IC.

Transient Receptor Potential Vanilloid Subfamily 1 is Essential for the Generation of Noxious Bladder Input and Bladder Overactivity in Cystitis

We evaluated the role of transient receptor potential vanilloid subfamily 1 for the generation of noxious bladder input and bladder overactivity associated with cystitis. Spinal c-fos expression triggered by innocuous bladder distention (10 cm water) was studied in sham and lipopolysaccharide inflamed transient receptor potential vanilloid subfamily 1 +/+ and -/- mice. Bladder reflex activity was studied using urethane anesthesia in sham and lipopolysaccharide inflamed transient receptor potential vanilloid subfamily 1 +/+ and -/- mice. Inflammatory changes in the bladder of transient receptor potential vanilloid subfamily 1 +/+ and -/- mice were identical. Bladder distention in sham inflamed +/+ mice induced a mean +/- SD of 4 +/- 2 Fos cells per section. Bladder distention after lipopolysaccharide inflammation increased Fos cells to 34 +/- 5 (p <0.001). The number of Fos cells after bladder distention in sham and lipopolysaccharide inflamed -/- mice was similar (2 +/- 1 and 2 +/- 1, respectively, p >0.05). During saline infusion of sham inflamed bladders in +/+ mice 0.46 +/- 0.14 contractions per minute were documented. In lipopolysaccharide inflamed +/+ mice that frequency was increased to 1.13 +/- 0.12 contractions per minute (p <0.001). In sham and lipopolysaccharide inflamed -/- mice bladder frequency was similar (0.47 +/- 0.08 and 0.61 +/- 0.10, respectively, p >0.05). Our data demonstrate that transient receptor potential vanilloid subfamily 1 is essential for the generation of noxious bladder input and bladder overactivity associated with cystitis.

Selection of the sacral nerve posterior roots to establish skin–CNS–bladder reflex pathway: An experimental study in rats

The purpose of this study was to explore the innervations of different sacral nerve posterior roots to bladder, and to provide evidence for further study of skin-CNS-bladder reflex pathway in the spinal cord injury patient. Spinal cord injury was produced in 10 rats. The bilateral spinal posterior roots of S1-S4 were electrically stimulated, and the bladder plexus action, bladder smooth muscle complex action potential, and intravesical pressure were examined and measured. The results showed that all the sacral nerve posterior roots were involved in innervations of bladder in rats. Among them, the S2 sacral nerve is the dominant nerve in innervations of bladder, followed by S1, S3, and S4 sacral nerve posterior roots. This study has provided valuable information for selection of sacral nerve posterior root for further study of the artificial bladder reflex arc for improving the micturition function in spinal cord injury patients.

Persistence of therapeutic effect after repeated injections of botulinum toxin type A to treat incontinence due to neurogenic detrusor overactivity

Objectives To study the effect of repeated botulinum toxin type A injections into the detrusor in patients with neurogenic detrusor overactivity and incontinence to determine the safety of repeated injections, the persistence of the clinical and urodynamic treatment efficacy, and potential changes in bladder compliance. Methods Seventeen patients with neurogenic detrusor overactivity who had received three or more botulinum toxin type A injections into the detrusor were studied. The clinical and urodynamic data were analyzed at baseline (before the first injection), after the first injection, and after the last repeated injection. Results No systemic side effects were observed for the total of 91 injections. The mean number of injections per patient was 5.4 (range 3 to 9). The mean number of incontinence episodes per day decreased from 2.6 at baseline to 0 after the first injection, and remained at 0 after the last injection. The maximal cystometric bladder capacity and reflex volume increased significantly after the first and last injection compared with at baseline. The maximal detrusor pressure decreased significantly after the first and last injection compared with at baseline. No difference in compliance was found from baseline to the first or last injection. Conclusions Repeated injections of botulinum toxin A into the detrusor muscle are a safe and valuable treatment option for neurogenic detrusor overactivity. After repeated injections, the effect on the clinical and urodynamic parameters remained constant. Also, repeated injections did not decrease bladder compliance.

Effects of Substitution on 9-(3-Bromo-4-fluorophenyl)-5,9-dihydro-3H,4H-2,6-dioxa-4- azacyclopenta[b]naphthalene-1,8-dione, a Dihydropyridine ATP-Sensitive Potassium Channel Opener

Structure-activity relationships were investigated on the tricyclic dihydropyridine (DHP) KATP openers 9-(3-bromo-4-fluorophenyl)-5,9-dihydro-3H,4H-2,6-dioxa-4-azacyclopenta[b]naphthalene-1,8-dione (6) and 10-(3-bromo-4-fluorophenyl)-9,10-dihydro-1H,8H-2,7-dioxa-9-azaanthracene-4,5-dione (65). Substitution off the core of the DHP, absolute stereochemistry, and aromatic substitution were evaluated for KATP channel activity using Ltk- cells stably transfected with the Kir6.2/SUR2B exon 17- splice variant and in an electrically stimulated pig bladder strip assay. A select group of compounds was evaluated for in vitro inhibition of spontaneous bladder contractions. Several compounds were found to have the unique characteristic of partial efficacy in both the cell-based and electrically stimulated bladder strip assays but full efficacy in inhibiting spontaneous bladder strip contractions. For compound 23b, this profile was mirrored in vivo where it was fully efficacious in inhibiting spontaneous myogenic bladder contractions but only partially able to reduce neurogenically mediated reflex bladder contractions.