Decreased Micturition Pressure Research Articles

A Preclinical Study of Human Embryonic Stem Cell-Derived Mesenchymal Stem Cells for Treating Detrusor Underactivity by Chronic Bladder Ischemia

BackgroundThe therapeutic effects of human embryonic stem cell-derived multipotent mesenchymal stem cells (M-MSCs) were evaluated for detrusor underactivity (DUA) in a rat model with atherosclerosis-induced chronic bladder ischemia (CBI) and associated mechanisms.MethodsSixteen-week-old male Sprague–Dawley rats were divided into five groups (n = 10). The DUA groups underwent 30 bilateral repetitions of endothelial injury to the iliac arteries to induce CBI, while the sham control group underwent a sham operation. All rats used in this study received a 1.25% cholesterol diet for 8 weeks. M-MSCs at a density of 2.5, 5.0, or 10.0 × 105 cells (250 K, 500 K, or 1000 K; K = a thousand) were injected directly into the bladder 7 weeks post-injury, while the sham and DUA group were treated only with vehicle (phosphate buffer solution). One week after M-MSC injection, awake cystometry was performed on the rats. Then, the bladders were harvested, studied in an organ bath, and prepared for histological and gene expression analyses.ResultsCBI by iliac artery injury reproduced voiding defects characteristic of DUA with decreased micturition pressure, increased micturition interval, and a larger residual volume. The pathological DUA properties were improved by M-MSC treatment in a dose-dependent manner, with the 1000 K group producing the best efficacy. Histological analysis revealed that M-MSC therapy reduced CBI-induced injuries including bladder fibrosis, muscular loss, and apoptosis. Transplanted M-MSCs mainly engrafted as vimentin and NG2 positive pericytes rather than myocytes, leading to increased angiogenesis in the CBI bladder. Transcriptomes of the CBI-injured bladders were characterized by the complement system, inflammatory, and ion transport-related pathways, which were restored by M-MSC therapy.ConclusionsSingle injection of M-MSCs directly into the bladder of a CBI-induced DUA rat model improved voiding profiles and repaired the bladder muscle atrophy in a dose-dependent manner.Graphical abstract

Therapeutic Efficacy of Human Embryonic Stem Cell-Derived Multipotent Stem/Stromal Cells in Diabetic Detrusor Underactivity: A Preclinical Study.

Mesenchymal stem/stromal cell (MSC) therapy is a promising approach for treatment of as yet incurable detrusor underactivity (DUA), which is characterized by decreased detrusor contraction strength and/or duration, leading to prolonged bladder emptying. In the present study, we demonstrated the therapeutic potential of human embryonic stem cell (ESC)-derived multipotent MSCs (M-MSCs) in a diabetic rat model of DUA. Diabetes mellitus (DM) was induced by intraperitoneal injection of streptozotocin (STZ) (50 mg/kg) into 8-week-old female Sprague-Dawley rats. Three weeks later, various doses of M-MSCs (0.25, 0.5, and 1 × 106 cells) or an equivalent volume of PBS were injected into the outer layer of the bladder. Awake cystometry, organ bath, histological, and gene expression analyses were evaluated 1 week (short-term) or 2 and 4 weeks (long-term) after M-MSC transplantation. STZ-induced diabetic rats developed DUA, including phenotypes with significantly longer micturition intervals, increased residual urine amounts and bladder capacity, decreased micturition pressure on awake cystometry, and contractile responses to various stimuli in organ bath studies. Muscle degeneration, mast cell infiltration, fibrosis, and apoptosis were present in the bladders of DM animals. A single local transplantation of M-MSCs ameliorated DUA bladder pathology, including functional changes and histological evaluation, and caused few adverse outcomes. Immunostaining and gene expression analysis revealed that the transplanted M-MSCs supported myogenic restoration primarily by engrafting into bladder tissue via pericytes, and subsequently exerting paracrine effects to prevent apoptotic cell death in bladder tissue. The therapeutic efficacy of M-MSCs was superior to that of human umbilical cord-derived MSCs at the early time point (1 week). However, the difference in efficacy between M-MSCs and human umbilical cord-derived MSCs was statistically insignificant at the later time points (2 and 4 weeks). Collectively, the present study provides the first evidence for improved therapeutic efficacy of a human ESC derivative in a preclinical model of DM-associated DUA.

Pharmacological Characterization of a Novel Beta 3 Adrenergic Agonist, Vibegron: Evaluation of Antimuscarinic Receptor Selectivity for Combination Therapy for Overactive Bladder.

Although the physiologic role of muscarinic receptors in bladder function and the therapeutic efficacy of muscarinic antagonists for the treatment of overactive bladder are well established, the role of β3-adrenergic receptors (β3ARs) and their potential as therapeutics is just emerging. In this manuscript, we characterized the pharmacology of a novel β3AR agonist vibegron (MK-4618, KRP-114V) and explored mechanistic interactions of β3AR agonism and muscarinic antagonism in urinary bladder function. Vibegron is a potent, selective full β3AR agonist across species, and it dose dependently increased bladder capacity, decreased micturition pressure, and increased bladder compliance in rhesus monkeys. The relaxation effect of vibegron was enhanced when combined with muscarinic antagonists, but differentially influenced by muscarinic receptor subtype selectivity. The effect was greater when vibegron was co-administered with tolterodine, a nonselective antagonist, compared with coadministration with darifenacin, a selective M3 antagonist. Furthermore, a synergistic effect for bladder strip relaxation was observed with the combination of a β3AR agonist and tolterodine in contrast to simple additivity with darifenacin. To determine expression in rhesus bladder, we employed a novel β3AR agonist probe, [3H]MRL-037, that selectively labels β3 receptors in both urothelium and detrusor smooth muscle. Vibegron administration caused a dose-dependent increase in circulating glycerol and fatty acid levels in rhesus and rat in vivo, suggesting these circulating lipids can be surrogate biomarkers. The translation of our observation to the clinic has yet to be determined, but the combination of β3AR agonists with M2/M3 antimuscarinics has the potential to redefine the standard of care for the pharmacological treatment of overactive bladder.

Comparative Analysis of the Effects of Antimuscarinic Agents on Bladder Functions in Both Nonhuman Primates and Rodents

Both the physiological role of muscarinic receptors for bladder function and the therapeutic efficacy of antimuscarinic agents for overactive bladder syndrome are well documented. We investigated the effect of antimuscarinic agents with different subtype selectivity on urodynamic parameters in nonhuman primates and rodents and compared plasma levels of these agents between species. Anesthetized rhesus monkeys were transurethrally catheterized, and the bladder was infused with saline. Urodynamic parameters were measured before and after intravenous drug administration. Tolterodine (nonselective) and oxybutynin (moderately M(3)-selective) increased bladder capacity at lower doses than those required to decrease micturition pressure. However, higher doses of darifenacin (M(3)-selective) were needed to increase the bladder capacity than those needed to decrease the micturition pressure. In rats, tolterodine had no effect on the bladder capacity but decreased the micturition pressure at all of the doses administered. Oxybutynin also decreased micturition pressure and increased bladder capacity at the highest dose. Plasma levels of these drugs overlap in both species. These results suggest that, in addition to the M(3) receptor, other muscarinic receptor subtypes contribute to regulate bladder storage function in nonhuman primates, since less subtype-selective tolterodine and oxybutynin showed higher specificity to the bladder capacity effect than the effect on micturition pressure compared with M(3)-selective darifenacin. In addition, the role of muscarinic receptors in bladder storage function varies between primates and rodents. Compared with rodents, muscarinic receptors may play a more active role during the storage phase to regulate the functional bladder capacity in primates.

Impaired Bladder Function in Aging Male Rats

The prevalence of bladder dysfunctions increases with age. In humans it is difficult to separate changes related to exogenous factors from those directly related to the aging process. Some confounding variables can be avoided by studying age related changes in an animal model. We evaluated the impact of age on bladder function in vivo and in vitro, and characterized the corresponding morphological changes. Young (4 to 6 months old) and old (older than 28 to 30 months) male Fischer/Brown Norway rats were used in the study. Cystometric studies were done in conscious, freely moving rats. After cystometry tissue strips from the bladder body were used in in vitro studies of muscarinic receptor activation and electrical field stimulation, and histological examination. Old rats had higher bladder weight than young rats but the bladder-to-body weight ratio did not change. We noted significant age related differences in 8 of 10 cystometric parameters. Old rats had increased bladder capacity, post-void residual volume, micturition volume and frequency, baseline and intermicturition pressure, and spontaneous activity but decreased micturition pressure. Bladder strip responses to carbachol and electrical field stimulation were significantly lower in old than in young rats. Histological examination revealed urothelial thinning, lower muscle mass and higher collagen content in the bladders of old vs young rats. Physiological aging alters bladder function in male rats even when external factors remain constant. Thus, in old rats bladder capacity, post-void residual urine and spontaneous activity are higher, and responses to muscarinic receptor stimulation and electrical field stimulation are lower than in young rats. Such changes correspond to findings in aging human bladders, supporting the view that the Fischer/Brown Norway rat is a useful model in which to study age related bladder function changes.

Transplantation of Neuronal and Glial Restricted Precursors into Contused Spinal Cord Improves Bladder and Motor Functions, Decreases Thermal Hypersensitivity, and Modifies Intraspinal Circuitry

Transplanting neuronal and glial restricted precursors (NRP/GRP) into a midthoracic injury 9 d after contusion improved bladder and motor function, diminished thermal hypersensitivity, and modified lumbosacral circuitry compared with operated controls (OP-controls). Histological analysis showed that NRP/GRP survived, filled the lesion site, differentiated into neurons and glia, and migrated selectively. Volume of spinal cord spared was increased in NRP/GRP recipients, suggesting local protection. Bladder areflexia developed in both operated groups, but NRP/GRP recipients exhibited an accelerated recovery, with decreased micturition pressure and fewer episodes of detrusor hyperreflexia. Because noradrenergic receptors proliferate after spinal injury and descending noradrenergic pathways contribute to regulation of bladder control, we examined the effects of administering an alpha-1A-adrenergic antagonist, Tamsulosin, on urodynamics. This improved all cystometric parameters in both operated groups, and micturition pressure in NRP/GRP rats recovered to normal levels. Both operated groups initially showed increased sensitivity to a thermal stimulus applied to the tail; the NRP/GRP rats showed significant improvement over time. NRP/GRP grafts also produced greater recovery of hindlimb function in several tests, although both groups showed persistent and similar deficits in locomotion on a grid. Because bladder, hindlimb, and tail sensory and motor functions are organized through lumbosacral cord, we examined descending and primary afferent projections at L6-S1. The density of serotonergic, noradrenergic, and corticotrophin releasing factor-positive fibers increased in the NRP/GRP group compared with OP-controls, suggesting some sparing and/or sprouting of these modulatory pathways. Immunocytochemical staining density of dorsal root axons in the dorsal horn increased in the OP-controls but appeared normal in the NRP/GRP group. Synaptophysin immunoreactivity in the lumbosacral dorsal horn was similar among groups, consistent with restoration of synaptic density in both groups of operated animals but by different pathways. We suggest that local protection provided by NRP/GRP resulted in increased sparing/sprouting of descending pathways, which prevented sprouting by dorsal root axons, and that this modification in lumbosacral circuitry contributes to the recovery of function.

The effect of tamsulosin on the response of the rabbit bladder to partial outlet obstruction

To determine if tamsulosin treatment prevents or decreases the incidence and severity of outlet obstruction-induced bladder dysfunction in rabbits. Male New Zealand White rabbits were treated with tamsulosin or vehicle for 4 weeks with treatments initiated 1 week prior to sham or obstruction surgery. Cystometry was done on anesthetized rabbits 21 days after surgery. The bladders were then removed, weighed, and prepared for in vitro whole bladder studies. Responses to 32 Hz field stimulation (FS), carbachol, phenylephrine, and KCl were measured. Obstruction resulted in a significant increase in bladder weight, which was unchanged by tamsulosin treatment and a significant increase in micturition pressure in the vehicle-treated group but not in the tamsulosin-treated group. Compliance was significantly decreased in both obstructed groups. The vehicle-treated obstructed rabbits had a very sharp increase in intravesical pressure as the bladder reached capacity; this was not seen in the tamsulosin-treated obstructed rabbits. Tamsulosin did not change the pattern of modifications in contractile responses induced by bladder outlet obstruction. In vitro responses of vehicle and tamsulosin-treated obstructed rabbit groups in this study were similar. A greater micturition pressure was found for the vehicle-treated obstructed group than for the tamsulosin-treated obstructed group, which was probably due to decreased urethral resistance in the latter. On a functional basis, the higher compliance at capacity and decreased micturition pressure in the tamsulosin-treated obstructed group would be considered beneficial for bladder function.

Transplants of fibroblasts expressing BDNF and NT-3 promote recovery of bladder and hindlimb function following spinal contusion injury in rats

We examined whether fibroblasts, genetically modified to express BDNF and NT-3 (Fb-BDNF/NT3) and transplanted into a thoracic spinal injury site, would enhance recovery of bladder function and whether this treatment would be associated with reorganization of lumbosacral spinal circuits implicated in bladder function. Rats received modified-moderate contusion injuries at T8/9, and 9 days later, Fb-BDNF/NT3 or unmodified fibroblasts (OP-controls) were delivered into the cord. Fb-BDNF/NT3 rats recovered from areflexic bladder earlier, showed decreased micturition pressure and fewer episodes of detrusor hyperreflexia, compared to OP-controls. There were also improvements in hindlimb function in the Fb-BDNF/NT3 group although locomotion on a more challenging substrate (grid) and tail withdrawal latency in response to a thermal stimulus showed persisting deficits, little recovery, and no differences between the groups. Immunocytochemistry at L6-S1 revealed changes in density of afferent and descending projections to L6-S1 cord. The density of small dorsal root axons increased in the superficial layers of the dorsal horn in OP-controls but not in Fb-BDNF/NT3, suggesting sprouting of primary afferents following injury that was inhibited by Fb-BDNF/NT-3. In contrast, the trophic factor secreting transplants stimulated sprouting and/or sparing of descending modulatory pathways projecting to the lumbosacral spinal cord. No differences in synaptophysin immunoreactivity were seen in the dorsal horn which suggested that synaptic density was similar but achieved by sprouting of different systems in the two operated groups. Fb-BDNF/NT3 transplanted into injured spinal cord thus improved both bladder and hindlimb function, and this was associated with reorganization of spinal circuitry.

Inhibition of Hyperreflexia by Vaginally Administered Oxybutynin: A Novel Rabbit Model

The rabbit has proved to be an excellent model in which to study lower urinary tract function and pathological mechanisms involved in specific forms of lower urinary tract dysfunctions, including those involved with partial outlet obstruction (benign prostatic hyperplasia), erectile dysfunction, bladder instability and ischemic bladder disease. The current study describes a model of bladder instability induced in female rabbits and its use in developing a novel form of vaginal treatment for this instability. A total of 18 female New Zealand White rabbits were used for this experiment. Each rabbit was anesthetized and the carotid artery was cannulated for blood pressure monitoring. The bladder dome was catheterized for monitoring bladder pressure and for cystometry. A ligature was placed around the urethra just distal to the bladder neck to prevent leakage through the urethra and generate unstable bladder contractions. After initial cystometry the bladder was emptied and then filled to 30 ml. Acetylcholine (Ach) was administered into the vesical artery at 15-minute intervals for 2 hours. Hyperreflexia usually developed within 30 minutes. Vaginal oxybutynin (low and high dose) was applied and the effect on cystometry, the response to Ach, and the amplitude and frequency of hyperreflexia was determined. Each dose of oxybutynin increased bladder capacity and improved compliance. The high oxybutynin dose completely inhibited the response to Ach and significantly decreased micturition pressure. The high dose decreased uninhibited bladder contraction peak pressure in a time dependent manner to a greater extent than it decreased uninhibited contraction frequency. The low dose of oxybutynin had no effect on micturition pressure, significantly inhibited the response to Ach and hyperreflexia amplitude to approximately 40% of control values, and had little effect on hyperreflexia frequency. Vaginally delivered oxybutynin was effective in improving bladder compliance, inhibiting intra-arterial Ach stimulated bladder contractions and significantly decreasing the amplitude of hyperreflexia. Vaginal oxybutynin had a more potent effect on the evoked response to Ach and on hyperreflexia than it did on micturition pressure.

Effects of NAD-299, a new, highly selective 5-HT1A receptor antagonist, on bladder function in rats.

5-HT is involved in micturition control. In the rat, stimulation of the 5-HT(1A) receptor excites, whereas 5-HT(1A) receptor antagonism inhibits micturition. The present study examined the effects of a new, highly selective, 5-HT(1A) receptor antagonist, NAD-299, on micturition in rats. Comparisons were made with WAY100635, a well-characterised antagonist at the 5-HT(1A) receptor. Female Sprague-Dawley rats, conscious or anaesthetised, were used for cystometric studies. Intravenous (i.v.), intraarterial (i.a.), intrathecal (i.t.) or intracerebroventricular (i.c.v.) catheters were implanted for drug administration. In vitro, rat bladder strips were contracted by carbachol or electrical stimulation of nerves. The effects of NAD-299, WAY100635 and 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT, a 5-HT(1A) receptor agonist) on cystometric parameters and contraction of bladder strips were recorded. In conscious rats, i.v. NAD-299 and WAY100635 at 1 micro mol kg(-1) increased bladder capacity (24+/-13% and 27+/-19% respectively) and decreased micturition pressure (16+/-8% and 12+/-10% respectively). Given i.a., 5-HT 0.25 micro mol kg(-1) and 8-OH-DPAT 0.37 micro mol kg(-1) stimulated micturition. The effect of 8-OH-DPAT, but not those of 5-HT, was blocked by i.a. NAD-299. 8-OH-DPAT 0.03 micro mol given i.t. or i.c.v. stimulated micturition, an effect blocked by WAY100635 0.1 micro mol, given i.t or i.c.v. NAD-299 or WAY100635 (0.1 micro mol i.t.) were without significant effects, but given i.c.v. at 0.1 micro mol both drugs increased bladder capacity (34+/-12% and 22+/-13% respectively). Neither NAD-299 nor WAY100635 up to 10(-5) M had effects on electrically- or carbachol-induced contractions of rat bladder strips. NAD-299 1 micro mol kg(-1) i.v. suppressed oxyhaemoglobin-induced detrusor over-activity. It is concluded that NAD-299, acting at a supraspinal site, can inhibit micturition in the rat.

Effects Of Tiagabine, A γ-Aminobutyric Acid Re-Uptake Inhibitor, on Normal Rat Bladder Function

Previous reports have demonstrated the inhibitory effect of exogenous gamma-aminobutyric acid (GABA) on micturition. In the current study we tested whether tiagabine (Sanofi Synthelab., Newcastle-upon Tyne, United Kingdom), a GABA re-uptake inhibitor increasing endogenous GABA concentrations, would affect micturition in awake rats or influence rat detrusor contraction in vitro. Nonanesthetized female Sprague-Dawley rats underwent cystometric investigation in a metabolic cage. Micturition was stimulated by infusing saline intravesically. Micturition parameters were recorded and compared before and after drug administration. In vitro the effects of tiagabine on electrical and carbachol induced contractions in bladder strips were investigated. Furthermore, it was studied whether tiagabine interfered with electrically induced release of acetylcholine. Intravenous administration of 5 and 20 mg. kg.-1 tiagabine in 7 and 9 rats decreased micturition pressure a mean plus or minus standard error of mean of 21% +/- 11% and 42% +/- 9%, and decreased voided volume a mean of 31% +/- 9% and 33% +/- 9%, respectively. At 20 mg. kg.-1 tiagabine intravenously increased post-void residual volume a mean of 300% +/- 120% and decreased bladder capacity a mean of 14% +/- 3%. Tiagabine (100 microg.) intrathecally in 7 rats reduced micturition pressure a mean of 34% +/- 10% and increased bladder capacity a mean of 30% +/- 9% and post-void residual volume a mean of 250% +/- 75%. However, voided volume was not changed. In vitro studies demonstrated that tiagabine attenuated bladder contractions induced by electrical field stimulation to a mean of 69% +/- 6% of controls at 100 microM. but did not affect contractions induced by carbachol. Release studies revealed that tiagabine inhibited electrical induced acetylcholine release to a mean of 82% +/- 5% of controls at 100 microM. The current results show that tiagabine has an inhibitory action on rat micturition. The site of action may be central and peripheral.

Role of supraspinal tachykinins for micturition in conscious rats with and without bladder outlet obstruction.

In order to clarify the role of supraspinal tachykinins in volume-induced micturition and in bladder hyperactivity secondary to bladder outlet obstruction, conscious, normal, female Sprague-Dawley rats were investigated cystometrically before and after intracerebroventricular administration of RP 67,580, a selective antagonist of neurokinin (NK)-1 receptors and/or SR 48,968, a selective antagonist of NK-2 receptors. In normal rats, RP 67,580 or SR 48,968, at a dose of 2 nmol, caused no marked changes in cystometric parameters. Higher doses (up to 20 nmol) caused dose-dependent decreases in micturition pressure and increased bladder capacity, micturition volume and residual urine. A combination of the two drugs, each at a dose of 2 nmol, significantly decreased micturition pressure and increased bladder capacity. In rats with bladder outlet obstruction, the antagonists suppressed micturition dose-dependently, producing urinary retention in two out of eight rats already at a dose of 2 nmol. At a dose of 20 nmol, dribbling incontinence, due to urinary retention, was seen in five out of ten rats. A combination of the two drugs (2 nmol of each drug) caused urinary retention in three out of nine animals and significantly increased bladder capacity, micturition volume and residual volume. The results suggest that outflow obstruction in rats increases the effects of tachykinins in supraspinal structures involved in micturition, and that antagonism of supraspinal NK-receptors may depress the micturition reflex. Whether or not this implies that supraspinal NK-receptors can be targets for drugs aimed for inhibiting bladder hyperactivity in humans should be explored.

Role of supraspinal tachykinins for volume- and L-dopa-induced bladder activity in normal conscious rats.

To clarify the roles of tachykinins in volume-induced micturition and in bladder hyperactivity, presumed to originate from supraspinal structures, normal, female Sprague-Dawley rats were investigated cystometrically before and after intracerebroventricular (i.c.v) administration of RP 67,580, a selective antagonist of NK-1 receptors, and/or SR 48,968, a selective antagonist of NK-2 receptors. The effects of RP 67,580 and SR 48,968 on intra-peritoneal (i.p.) L-dopa-induced bladder hyperactivity were also investigated. I.c.v. administration of RP 67,580 (20 nmol) SR 48,968 (20 nmol) suppressed micturition. Combination of i.c.v. RP 67, 580 (2 nmol) and SR 48,968 (2 nmol) significantly decreased micturition pressure (18%), and increased bladder capacity (26%), micturition volume (18%), and residual volume (223%). In rats pretreated with i.p. carbidopa 50 mg/kg, i.p. L-dopa 50 mg/kg caused bladder hyperactivity that was attenuated by the combination of i.c. v. RP 67,580 (2 nmol) and SR 48,968 (2 nmol). The results suggest that tachykinins, via stimulation of NK receptors in supraspinal structures, are involved in both volume and L-dopa-induced stimulation of bladder activity. This may imply that tachykinins can influence both the supraspinal and spinal control of the urinary bladder. It also implies that supraspinal NK receptors are a possible target for drugs aimed for elimination of bladder hyperactivity mediated via these pathways. Neurourol. Urodynam. 19:101-109, 2000.

EFFECTS OF INTRAVESICAL ADMINISTRATION OF THE K + CHANNEL OPENER, ZD6169, IN CONSCIOUS RATS WITH AND WITHOUT BLADDER OUTFLOW OBSTRUCTION

Purpose To investigate the urodynamic effects of the new K ATP channel opener, ZD6169, given intravesically, in rats with and without bladder outflow obstruction. Materials and Methods Female, conscious Sprague-Dawley rats, normal or with bladder hypertrophy and overactivity secondary to bladder outflow obstruction, were given ZD6169 intravesically (10 or 100 ng./ml.), and intra-arterially (1 mg./kg.). Continuous cystometry was performed. Results In normal and obstructed rats, intravesical ZD6169 had similar, dose-dependent effects on bladder function. In obstructed rats, ZD6169 (100 ng./ml.) significantly decreased micturition pressure (17%), and increased bladder capacity (32%), micturition volume (18%), residual volume (145%), and inter-contraction interval (71%). There was a marked decrease in both frequency (40%) and amplitude (43%) of the spontaneous bladder activity. When given intra-arterially in obstructed rats ZD6169 increased bladder capacity (19%) and residual volume (47%) and decreased amplitude (51%), but not frequency, of the spontaneous bladder activity. Conclusions In both normal and obstructed rats, intravesical ZD6169, at the investigated doses, significantly affected bladder function. In obstructed rats, the drug markedly reduced bladder overactivity. If the results have clinical validity, intravesical ZD6169 may offer an alternative way of treating bladder overactivity in selected patients.

Effect of NS-21, an anticholinergic drug with calcium antagonistic activity, on lower urinary tract function in a rat model of urinary frequency.

NS-21 is under development for the treatment of urinary frequency and urinary incontinence. The purpose of this study was to investigate the effects of NS-21 and its active metabolite, RCC-36, on lower urinary tract function in an experimental rat model of urinary frequency. Cystometrograms were recorded in anesthetized rats with bilaterally transected hypogastric nerves. All drugs were administered intraduodenally. In sham-operated rats, NS-21 (> or = 50 mg/kg) significantly increased the bladder capacity without significantly decreasing micturition pressure, while RCC-36 (100 mg/kg) significantly increased bladder capacity, and at a dose of > or = 30 mg/kg, also caused a decrease in micturition pressure. This increase in bladder capacity appeared at lower doses of both NS-21 and RCC-36 in the hypogastric nerve-transected rats. Propiverine (100 mg/kg) increased bladder capacity and at > or = 30 mg/kg, decreased micturition pressure in both sham-operated and nerve-transected rats. Oxybutynin (100 mg/kg) and atropine (30 mg/kg) decreased the micturition pressure in both sham-operated and nerve-transected rats without increasing the bladder capacity, while a similar anticholinergic calcium antagonist, terodiline (100 mg/kg) had no effect on bladder capacity in either sham-operated or nerve-transected rats. Flavoxate (500 mg/kg) significantly increased bladder capacity without significantly decreasing micturition pressure in both sham-operated and nerve-transected rats, while 50 mg/kg of verapamil significantly increased bladder capacity without significantly decreasing the micturition pressure in nerve-transected rats. NS-21 and RCC-36 increased bladder capacity at lower doses in hypogastric nerve-transected rats than in sham-operated rats. Furthermore, NS-21 increased the bladder capacity without suppressing micturition pressure, suggesting that NS-21 may be a more effective therapeutic drug than propiverine, oxybutynin or flavoxate for the treatment of urinary frequency and urinary incontinence.

Stimulation of bladder activity by volume, L-dopa and capsaicin in normal conscious rats--effects of spinal alpha 1-adrenoceptor blockade.

To study possible differences in alpha 1-adrenoceptor involvement in the spinal mechanisms mediating bladder activity induced by volume (bladder filling), central (L-dopa), and peripheral (capsaicin) stimulation, we investigated if these types of bladder activity were modified by intrathecal (i.t.) or intra-arterial (i.a.) administration of the alpha 1-adrenoceptor antagonist, indoramin. Indoramin is selective for the alpha 1A-adrenoceptor subtype, whereas most clinically used alpha 1-adrenoceptor antagonists, including doxazosin, have no subtype selectivity. The drug effects were studied by continuous cystometry in normal, conscious rats and rats with bladder activity evoked by intraperitoneal L-dopa (50 mg/kg after carbidopa pretreatment), or by intravesical capsaicin (30 microM). I.t. indoramin (50 nmol) significantly decreased micturition pressure, and increased bladder capacity and micturition volume. Dribbling incontinence due to urinary retention was observed in one of ten rats. L-dopa-stimulated bladder overactivity was significantly attenuated by i.t. or i.a. indoramin (50 nmol). Similar effects of i.t. and i.a. doxazosin (50 nmol) have been reported previously. Intravesical capsaicin (30 microM) caused bladder activity, which was attenuated by i.t. indoramin (50 nmol), but not by i.t. doxazosin (50 nmol). I.a. indoramin did not reduce capsaicin-induced bladder activity; doxazosin was moderately effective. The results suggest that the bulbospinal micturition reflex evoked by bladder filling and L-dopa involves a descending pathway where transmission is partly mediated by spinal alpha 1-adrenoceptors. Bladder overactivity evoked by intravesical capsaicin, which elicits a vesicospinal-vesical reflex, was not affected by i.t. doxazosin in a dose that attenuates activity mediated through the bulbo-spinal pathway. This suggests less involvement of spinal alpha 1-adrenoceptors in the vesico-spinal-vesical than in the bulbo-spinal voiding reflex.

Role of Spinal and Peripheral Alpha sub 2 Adrenoceptors in Micturition in Normal Conscious Rats

Purpose To explore the potential of alpha 2 -adrenoceptors in modulating vesicourethral function at the spinal and bladder levels. Material and Methods Continuous cystometry was performed in normal, conscious rats, and the urodynamic effects of alpha 2 -adrenoceptor stimulation and blockade were recorded. Results Given intrathecally, the selective alpha 2 -adrenoceptor agonist, dexmedetomidine (dex) 3 nmol., decreased micturition pressure, bladder capacity, micturition volume and residual volume. These effects were counteracted by the selective alpha 2 -adrenoceptor antagonist, atipamezole 30 nmol. Intrathecally dex 10 nmol. caused total incontinence in 9 of 9 rats. Given intra-arterially near the bladder, dex 6 nmol. decreased micturition pressure, bladder capacity, micturition volume, residual urine and basal pressure. Atipamezole 30 nmol. intrathecally increased micturition pressure, bladder capacity and residual urine, and decreased micturition volume. Similar effects were obtained when atipamezole 60 nmol. was given intra-arterially. The selective alpha 2 -adrenoceptor antagonist, rauwolscine 200 nmol., had effects similar to those of atipamezole. Conclusions The results show that alpha 2 -adrenoceptors can modulate micturition in normal conscious rats both at the spinal and the peripheral levels.

Acute effects of unilateral pelvic ganglionectomy on urinary bladder function in vivo in the male rat.

Mean and maximal micturition volumes following a standardized water intake were determined before and up to three days after unilateral pelvic ganglionectomy or sham operation in adult male rats. Sham operation did not change the volumes. Unilateral ganglionectomy on the other hand decreased significantly both mean and maximal micturition volumes (and thus increased micturition frequency). The effect was most pronounced 1 day after ganglionectomy, but was still significant after 3 days. Cystometrograms were recorded without and with atropine (1 mg/kg) before operation and 1, 2 or 3 days after sham-operation or ganglionectomy. Micturition pressure decreased to about 50% 1 day after ganglionectomy and remained at this level. Atropine decreased micturition pressure in the controls to about 55% of the initial. The atropine resistant pressure response in the ganglionectomized rats amounted to 90% after 1 day, and was still above 70% after 3 days. The sham-operated controls had no residual urine without or with atropine. The unilaterally ganglionectomized animals had no residual urine in the absence of atropine, but after administration of the drug the animals rapidly developed a significant residual urine.

Effects of Neurokinin Receptor Antagonists on L-DOPA Induced Bladder Hyperactivity in Normal Conscious Rats

Purpose The urodynamic effects of intrathecal neurokinin (NK) receptor blockade on L-dopa-induced bladder hyperactivity were investigated in a rat model. Materials and Methods Continuous cystometry was performed in normal, conscious, female Sprague-Dawley rats. Results In rats pretreated with intraperitoneal carbidopa 50 mg./kg., intraperitoneal L-dopa 50 mg./kg. caused bladder hyperactivity that could be attenuated by intrathecal administration of the NK 1 receptor selective antagonist SR 140,333 (2 nmol.), whereas the NK 2 receptor selective antagonist SR 48,968 (2 nmol.) failed to do so. Combination of SR 140,333 (2 nmol.) and SR 48,968 (2 nmol.), which by itself decreased micturition pressure, practically abolished the L-dopa-induced hyperactivity. Conclusions The present results suggest that tachykinins, via stimulation of NK 1 (and/or NK 2) receptors, are involved in L-dopa-induced bladder hyperactivity, most probably at the spinal level. This implies tachykinin involvement in the supraspinal pathways that control the sacral parasympathetic center innervating the urinary bladder. It also implies that spinal NK receptors are a possible target for drugs aimed for elimination of bladder hyperactivity mediated via these pathways.

Role of intrathecal tachykinins for micturition in unanaesthetized rats with and without bladder outlet obstruction.

1. The effects on micturition of RP 67,580, a selective NK1 receptor antagonist, and SR 48,968, a highly, potent antagonist at NK2 receptor sites, given intrathecally (i.t.) or intra-arterially (i.a.) near the bladder, were investigated in unanaesthetized rats with and without bladder outlet obstruction. 2. In normal rats, RP 67,580, given i.t. in doses of 2 and 20 nmol per rat, decreased micturition pressure, but did not change other cystometric parameters. After 20 nmol of RP 67,580, dribbling incontinence due to retention was observed in 1 out of 7 animals. This effect was reversible. I.t. RP 67,580 in a dose of 2 nmol, had no effect on hyperactivity induced by intravesically instilled capsaicin. 3. In animals with bladder hypertrophy secondary to outflow obstruction, RP 67,580, given i.t. in a dose of 2 nmol per rat, decreased the micturition pressure, but had no effect on other cystometric parameters. After 20 nmol, dribbling incontinence due to retention was observed in 5 out of 7 animals. 4. RP 67,580, given i.a. in a dose of 4 nmol, had little effect on the cystometric parameters investigated, both in normal animals and rats with bladder hypertrophy. 5. SR 48,968, given i.t. in doses of 2 and 20 nmol per rat, had no clear-cut effects on the micturition pattern in normal rats, or rats with bladder hypertrophy. However, the drug reduced capsaicin-induced bladder hyperactivity. When given i.a. in a dose of 4 nmol, SR 48,968 had no effect on cystometric parameters in normal rats or rats with bladder hypertrophy. 6. The effects of both RP 67,580 and SR 48,968 were stereoselective, their enantiomers (RP 68,651 and SR 48,965) being inactive.7. These results thus suggest that at the spinal level there is a tachykinin involvement (via NK,receptors) in the micturition reflex induced by bladder filling, both in normal rats, and, more clearly, in animals with bladder hypertrophy secondary to outflow obstruction. The bladder response to filling was not influenced by blockade of vesical NKI and NK2 receptors. On the other hand, the bladder hyperactivity evoked by intravesical capsaicin seems to involve NK2 receptors both at the bladder and spinal levels.