Does Reduction of Number of Intradetrusor Injection Sites of aboBoNTA (Dysport®) Impact Efficacy and Safety in a Rat Model of Neurogenic Detrusor Overactivity?

Intradetrusor injections of Botulinum toxin A-currently onabotulinumtoxinA-is registered as a second-line treatment to treat neurogenic detrusor overactivity (NDO). The common clinical practice is 30 × 1 mL injections in the detrusor; however, protocols remain variable and standardization is warranted. The effect of reducing the number of injection sites of Dysport(®) abobotulinumtoxinA (aboBoNTA) was assessed in the spinal cord-injured rat (SCI). Nineteen days post-spinalization, female rats received intradetrusor injections of saline or aboBoNTA 22.5 U distributed among four or eight sites. Two days after injection, continuous cystometry was performed in conscious rats. Efficacy of aboBoNTA 22.5 U was assessed versus aggregated saline groups on clinically-relevant parameters: maximal pressure, bladder capacity, compliance, voiding efficiency, as well as amplitude, frequency, and volume threshold for nonvoiding contractions (NVC). AboBoNTA 22.5 U significantly decreased maximal pressure, without affecting voiding efficiency. Injected in four sites, aboBoNTA significantly increased bladder capacity and compliance while only the latter when in eight sites. AboBoNTA significantly reduced NVC frequency and amplitude. This preclinical investigation showed similar inhibiting effects of aboBoNTA despite the number of sites reduction. Further studies are warranted to optimize dosing schemes to improve the risk-benefit ratio of BoNTA-based treatment modalities for NDO and further idiopathic overactive bladder.

Bladder and erectile dysfunctions in the Type 2 diabetic Goto-Kakizaki rat.

Despite the fact that urogenito-sexual complications significantly impact the quality of life of diabetic patients, a robust in vivo experimental model is lacking. Bladder and erectile function in the Type 2 diabetic Goto-Kakizaki (GK) rat and responses to standard-of-care treatments for each disorder have been assessed. GK rats (n = 25, 18-wk-old, GK/Par colony) and age-matched Wistar rats (n = 23), characterized for their metabolic parameters, were used. Bladder function was assessed by cystometry in conscious rats treated by intravenous solifenacin (1 mg/kg). Subsequently, erectile function was assessed under anesthesia following electrical stimulation of the cavernous nerve in presence of intravenous sildenafil (0.3 mg/kg). GK rats displayed detrusor overactivity with a significant increase in frequency/amplitude of nonvoiding contractions during the filling phase, together with an increase in bladder capacity, intercontraction interval, voided volume, and maximal pressure of voiding contraction. Solifenacin significantly decreased parameters characterizing voiding contractions without modifying voiding efficiency. Erectile function in GK rats was markedly impaired and remained so after sildenafil treatment despite a significant improvement. GK rats display both bladder and erectile dysfunctions and respond at least partially to standard-of-care treatments for each disorder, thus representing a suitable model to investigate the pathophysiology and assess the efficacy of new therapeutic agents for Type 2 diabetes-associated bladder and erectile complications.

A new experimental rat model of erectile dysfunction and lower urinary tract symptoms associated with benign prostatic hyperplasia: the testosterone-supplemented spontaneously hypertensive rat.

UNLABELLED: What's known on the subject? and What does the study add? Lower urinary tract symptoms (LUTS) resulting from benign prostatic hyperplasia (BPH) and erectile dysfunction (ED) are common problems in the aging male population. Moreover, several recent studies have shown that ED is closely associated with the presence and severity of LUTS independently of co-morbidities. However, the pathophysiological mechanisms linking LUTS/BPH and ED remain largely unexplored. The major difficulty in studying such relationships between ED and LUTS/BPH, and of exploring the impact of new therapeutic approaches for both LUTS/BPH and ED, is the lack of experimental model combining ED, prostate enlargement and bladder dysfunction all at once. The present study describes a new model of BPH, the SHR supplemented with testosterone which is the first animal model which displays all at once the key features of BPH: prostate enlargement and an increased sympathetic tone of bladder outlet mimicking the static and the dynamic components of voiding symptoms of BPH, a significant impairment of bladder function which reflects the storage symptoms of BPH and finally, ED. This model could be very relevant to better characterize the close relationship that exists between BPH/LUTS and ED, and to evaluate new therapeutic strategies for BPH together with their side effect profile on sexual function on the same animal, thus allowing a reduction of the number of animals to be used in such studies. Study Type - Aetiology (case control) Level of Evidence 3b. OBJECTIVE: • To design a new experimental model combining erectile dysfunction, prostate enlargement and urodynamic impairment characteristic of lower urinary tract symptoms (LUTS) associated with benign prostate hyperplasia (BPH). MATERIALS AND METHODS: • Three groups of animals (12-week-old; n= 7/group) were considered: Wistar Kyoto (control) rats (WKY), untreated spontaneously hypertensive rats (SHR) and SHR treated with testosterone (SHR-T, 3 mg/kg/day) for 3 weeks. • Cystometry experiments and evaluation of erectile function were performed. Prostate enlargement was evaluated. RESULTS: • SHR displayed a significant decrease in the intercontraction interval (ICI) and in the voided volume (VV) whereas non-voiding contractions (NVC) were increased. SHR-T exhibited a further decreased ICI and VV and an increased frequency of NVC. • Erectile responses to electrical stimulation of the cavernous nerve were significantly impaired in both SHR (-66%) and SHR-T (-58%). • The prostate weight was similar in WKY and SHR, but significantly increased in SHR-T. CONCLUSIONS: • The testosterone-supplemented SHR represents an experimental model for urodynamic impairment combining both static and dynamic components of voiding symptoms with erectile dysfunction and prostate enlargement. • This model is suitable for the assessment of sexual side effects of LUTS/BPH treatments and efficacy of new therapeutic agents in LUTS/BPH and associated erectile dysfunction.

Minimal effective dose of dysport and botox in a rat model of neurogenic detrusor overactivity.

BACKGROUND: Two botulinum toxins A have been evaluated for the treatment of refractory neurogenic detrusor overactivity (NDO) in humans: Dysport (abobotulinumtoxinA) and Botox (onabotulinumtoxinA). However, these two distinct commercialized products have different potency units and are not interchangeable. OBJECTIVE: Assessment of the dose response and determination of minimal effective dose (MED) for Dysport and Botox in spinal cord-injured (SCI) rats with NDO. DESIGN, SETTING, AND PARTICIPANTS: Female, adult, Sprague-Dawley rats (n=98) underwent T8-T9 spinal cord transection. Nineteen days after spinal cord injury, rats received intradetrusor injections (25µl injected, eight sites) of vehicle (V); Dysport 2, 5, 7.5, 10, and 12.5 U; and Botox 0.8, 2, 5, 7.5, and 10 U. Two days after injection, continuous cystometry was performed in conscious rats. MEASUREMENTS: Voiding contractions (VC) were assessed by duration of VC, intercontraction interval, voided volume, maximal pressure, pressure threshold change, and intravesical baseline pressure (BP), while nonvoiding contractions (NVC) were evaluated by amplitude, frequency, and volume threshold to elicit NVC. MEDs for Dysport and Botox were determined by analysis of variance step-down trend test. RESULTS AND LIMITATIONS: MEDs for Dysport and Botox were 10 U and 7.5 U, respectively. Regarding VC, only BP significantly decreased after 10 U Dysport and 7.5 U Botox compared to V (from 3.7±0.6 to 1.5±0.1 and 1.4±0.3mm Hg, respectively; p<0.01 and p<0.001, respectively). Dysport (10 U) and Botox (7.5 U) significantly inhibited NVC by decreasing their amplitude (from 7.4±1.1 to 5.8±0.5 and 5.4±0.6mm Hg, respectively; p<0.05); frequency (from 2.2±0.4 to 1.5±0.2 and 1.3±0.3 NVC per minute, respectively; p<0.01); and increasing volume threshold to elicit NVC (from 29.8±3.7 to 47.6±6.9 and 47.7±6.3%, respectively; p<0.05 and p<0.001, respectively). CONCLUSIONS: This is the first preclinical dose-ranging study with Dysport and Botox under standardized conditions showing similar inhibiting effects on NDO, albeit at different MEDs. It highlights the importance of distinguishing each preparation for predicted outcomes and doses to be used. Further studies in patients with NDO are warranted to confirm these experimental results.

Vardenafil decreases bladder afferent nerve activity in unanesthetized, decerebrate, spinal cord-injured rats.

BACKGROUND: Phosphodiesterase type 5 inhibitors (PDE5-Is) improve storage symptoms in benign prostatic hyperplasia patients, despite a lack of effect on peak urinary flow rate. Moreover, vardenafil improves urodynamic parameters in spinal cord-injured (SCI) patients with neurogenic detrusor overactivity (NDO). SCI rats also display NDO characterized by nonvoiding contractions (NVCs) during bladder filling, resulting in an increased bladder afferent nerve firing (BANF). OBJECTIVE: We postulated that vardenafil could improve urodynamic parameters by reducing BANF. The effect of vardenafil has been investigated on intravesical pressure by cystometry experiments while recording BANF in response to bladder filling. DESIGN, SETTING, AND PARTICIPANTS: Complete T7-T8 spinalization was performed in 15 female adult Sprague-Dawley rats (250-275 g). MEASUREMENTS: At 21-29 d postspinalization, fine filaments were dissected from the L6 dorsal roots and placed across a bipolar electrode. Bladder afferent nerve fibers were identified by electrical stimulation of the pelvic nerve and bladder distension. SCI rats were decerebrated before cystometry experiments. Bladders were filled to determine the maximal bladder filling volume (BFV) for each rat. Then, after bladder stabilization at 75% of maximal BFV, saline (n=7) or vardenafil 1 mg/kg (n=8) was delivered intravenously. NVCs and BANF were recorded for 45 min. RESULTS AND LIMITATIONS: In all SCI rats, BANF was already present and regular at resting conditions (26.2±4.1 spikes per second). During bladder filling, intravesical pressure (IVP) slowly increased with transient NVCs superimposed. Concomitantly, BANF progressively increased up to 2.4-fold at maximal BFV (2.08±0.24 ml). After stabilization at submaximal BFV, BANF was increased by 186±37%. Vardenafil injection induced an immediate decrease in NVCs compared to saline (p<0.001) and BANF (52% decrease vs 28% in saline after 45 min; p<0.001). CONCLUSIONS: Systemic vardenafil reduced both NVCs and BANF in unanesthetized, decerebrate, SCI rats. These findings provide new insights into the mechanism of action by which PDE5-Is improve storage symptoms in SCI patients.

Effects of potassium channel modulators on myogenic spontaneous phasic contractile activity in human detrusor from neurogenic patients.

OBJECTIVE: To characterize the spontaneous contractile activity (SCA) developed by detrusor from patients with neurogenic detrusor overactivity (NDO) because the alteration of detrusor properties plays a critical role in the pathogenesis of detrusor overactivity, as well as to evaluate the role of K(ATP) and K(Ca) channels on this SCA because these channels regulate detrusor SCA in many species, including humans without overactive bladder (OAB). PATIENTS AND METHODS: Human bladder samples were obtained from 44 patients undergoing cystectomy for bladder cancer with no known OAB symptoms and from 38 patients suffering from urodynamically diagnosed NDO. Detrusor strips with or without urothelium/suburothelium were mounted isometrically in organ baths filled with Krebs-HEPES (37 °C; 95% O(2) /5% CO(2) ). Strips were incubated with 10 µm pinacidil (K(ATP) opener) followed by 10 µm glibenclamide (K(ATP) blocker). In another set of experiments, strips were incubated with 30 µm NS-1619 (BK(Ca) opener) followed by 100 nm iberiotoxin (BK(Ca) blocker) or with 100 nm apamin (SK(Ca) blocker). RESULTS: SCA occurred more frequently with larger amplitude and area under the curve in detrusor strips from NDO patients compared to control patients. The presence of urothelium/suburothelium did not significantly modify SCA in either patient population. Pinacidil markedly inhibited SCA of detrusor strips from control and NDO patients. This effect was reversed by glibenclamide. By contrast, NS-1619 followed by iberiotoxin did not elicit any significant changes in SCA from NDO patients, contrary to control patients. CONCLUSIONS: K(ATP) and SK(Ca) channels regulate SCA of NDO patients' detrusor strips. By contrast, BK(Ca) channels are not involved in the regulation of detrusor SCA in NDO patients, whereas they regulate SCA in control patients. These results should be considered in the development of K(+) channels openers for the treatment of NDO. Moreover, SCA observed in vitro should be regarded as an in vitro modelling of human NDO.

Effect of a local delivery of triiodothyronine (T3) within neuroregenerative guide on recovery of erectile function in a rat-model of cavernous nerve injury.

INTRODUCTION: A promoting effect of thyroid hormones has been established on the maturation of central and peripheral nervous systems. However, effects on autonomic nerves have never been experimentally investigated. AIM: To assess the effect of a local treatment combining silicone guides and local administration of Triiodothyronine (T3) on the erectile function and the histological neuroregeneration of crushed cavernous nerves (CNs) in rats. METHODS: Forty-five rats were divided into five equal groups: SHAM surgery, guide without crush, crush, crush + guide, crush + guide + T3. All surgical procedures were bilateral. CNs were crushed with microvascular bulldog clamp of 100 g/cm(2). A silicone guide was placed around the nerves. The guides were filled with T3 neuroregenerative solution. MAIN OUTCOME MEASURES: Erectile function was assessed 10 weeks post-operatively. Intra-cavernous pressure (ICP) and mean arterial pressure (MAP) were monitored during electrical stimulation of CNs at various frequencies. The main outcome was hardness of erection defined as DeltaICP/MAP. Fluorescent immunohistochemical analysis of CNs was performed to assess regeneration of nerves morphologically. RESULTS: Electrophysiological data showed increased recovery of erectile function in the group with guide + T3 neuroregenerative solution compared with the empty guide. Immunohistochemical analysis of cavernous nerves demonstrated in morphology that regenerated axons were straighter in nerves with guide and more regular if guides had been filled with T3. CONCLUSION: The use of guides prevented axonal sprouting, facilitated functional neuroregeneration and enabled a local delivery of thyroid hormones. Triiodothyronine improved neuroregeneration and recovery of erectile function after a nerve-sparing-like injury in a rat model.

Apoptosis and effects of intracavernous bone marrow cell injection in a rat model of postprostatectomy erectile dysfunction.

OBJECTIVES: To investigate the pathophysiology of postprostatectomy erectile dysfunction (pPED) in a rat model of bilateral cavernous nerve ablation (BCNA) and to assess the effects of local bone marrow mononuclear cell (BMMNC) injection on erectile dysfunction (ED) and cavernosal cellular abnormalities caused by BCNA. DESIGN, SETTING, AND PARTICIPANTS: This was an experimental study in Fisher rats with BCNA. INTERVENTION: Intervention included BNCA, electrical stimulation of the pelvic ganglion, and local BMMNC injection. MEASUREMENTS: Erectile responses to electric pelvic ganglion stimulation were studied. Cavernous tissue was examined to determine the cell types undergoing apoptosis and to detect changes in protein and gene expression of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS) using real-time quantitative polymerase chain reaction (RTQ-PCR) and Western blotting. The effects of local BMMNC injection on these parameters were studied. RESULTS AND LIMITATIONS: Diffuse apoptosis was noted in the connective tissue mesenchymal cells and vascular smooth muscle and endothelial cells. Compared with sham-operated controls, nNOS and eNOS levels were decreased after 3 wk and were normal (eNOS) or increased (nNOS) after 5 wk, suggesting spontaneous nerve regeneration. Despite nNOS recovery, erectile responses to electrical stimulation remained impaired after 5 wk, when mesenchymal cell apoptosis was the main persistent biologic abnormality. BMMNC injection decreased apoptotic cell numbers, accelerated the normalisation of nNOS and eNOS, and partially restored erectile responses at week 5. CONCLUSIONS: Massive cell apoptosis may play a key role in the pathophysiology of pPED. In this animal model, apoptosis persisted despite spontaneous nerve regeneration, suggesting that the course of BCNA-induced cell dysfunction was independent of reinnervation. BMMNC improved erectile function by inhibiting apoptosis and may hold promise for repairing penile cell damage caused by radical prostatectomy (RP).

Daily treatment with sildenafil reverses endothelial dysfunction and oxidative stress in an animal model of insulin resistance.

OBJECTIVES: Patients with insulin resistance exhibit endothelial dysfunction with decreased nitric oxide (NO) production and increased oxidative stress. We postulated that daily sildenafil improved endothelial function in fructose-fed rats. METHODS AND RESULTS: Wistar rats were fed a standard or fructose-enriched diet (FFR) for 9 wk. From weeks 6-8, sildenafil was administered twice daily (sc, 20 m g/kg), followed by a 1-wk washout. Concentration-response curves (CRCs) to endothelium-dependent (acetylcholine [Ach] and A23187) and -independent (sodium nitroprusside [SNP]) relaxing agents were performed on isolated precontracted aortas and superior mesenteric arteries (SMAs). Vascular cyclic guanosine monophosphate (cGMP) content, urinary excretion of nitrates/nitrites (NOx) and 8-isoprostanes (IPT), and plasma levels of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were evaluated. Relaxations to ACh were significantly reduced in aortas and SMAs of FFR. Sildenafil restored ACh-induced relaxations in aortas and provoked a significant leftward shift of the CRC to ACh in SMAs, whereas it did not modify the enhanced relaxations to SNP in FFR. IL-6, TNF-alpha, vascular cGMP, and urinary NOx levels were not modified by the fructose or sildenafil treatment. Urinary IPT levels were significantly elevated in FFR and normalized by sildenafil. CONCLUSIONS: Endothelial dysfunction and oxidative stress associated with insulin resistance can be reversed by daily sildenafil, even 1 wk after treatment cessation.

The melanocortin agonist, melanotan II, enhances proceptive sexual behaviors in the female rat.

Melanocortins have been reported to play a role in the control of both male and female sexual behavior. The present study examined the effects of melanotan-II (MT-II), a cyclic peptide analogue of alpha-melanocyte stimulating hormone on appetitive and consummatory aspects of female sexual behavior, including aspects of sexual proceptivity (solicitations, hops and darts, ear wiggling, pacing) and receptivity (lordosis). One group of ovariectomized Long-Evans rats (n=7) was primed subcutaneously with estradiol benzoate (EB) and progesterone (P) (10 microg and 500 microg respectively) and another group (n=7) with EB (10 microg) and oil (EB alone). Paced mating tests were performed with sexually experienced males in unilevel chambers, which were bisected by a Plexiglas divider containing three holes, through which only the female could pass. MT-II (1 and 3 mg/kg) or saline was injected intravenously 10 min before each 30-min paced mating test. Each female received the 3 treatments. In females primed with EB+P both doses of MT-II increased the number of hops and darts and ear wiggling significantly, but did not alter pacing or lordosis. With EB alone, no effect of MT-II was observed on any of the parameters measured. These results suggest that P can interact with MT-II to increase proceptive behaviors. Because hops and darts are essentially solicitations, made in close proximity to the male, that indicate a desire on the part of females to receive mounts and intromissions, these data suggest that activation of melanocortin receptors may represent a promising mode of action for the treatment of women with hypoactive sexual desire.

L6-S1 spinal nerve stimulation reduces micturition frequency in anaesthetized rats with cyclophosphamide-induced cystitis.

OBJECTIVE: To further investigate the rationale for using spinal nerve stimulation (SNS) for treating bladder overactivity associated with cystitis in a rat model of cyclophosphamide-induced cystitis, as several studies suggested that symptoms associated with chronic cystitis could be treated using stimulation of sacral spinal nerves, but the mechanisms by which it works are unknown. MATERIALS AND METHODS: Cystitis was induced by i.p. injection of cyclophosphamide 48 h before the experiments in anaesthetized male rats. Neurograms were taken by placing a recording electrode onto the pelvic nerve and a stimulating electrode on either the L6 or S1 ipsilateral spinal nerves. Two selected intensities were then evaluated for SNS in control and cyclophosphamide-treated rats during cystometry. RESULTS: Cyclophosphamide resulted in significant bladder overactivity. There was no apparent difference in the neurograms generated in response to SNS of the S1 and L6 spinal nerves, and between cyclophosphamide and control rats. Intensities of 200 microA (Adelta-fibre-specific) and 2 mA (Adelta+ C-fibres) were chosen for SNS. Continuous SNS at 200 microA significantly reduced the frequency of voiding and non-voiding contractions in cyclophosphamide-treated rats. SNS at 2 mA resulted in the abolition of voiding contractions, accompanied by continuous leakage of urine. CONCLUSION: SNS recruiting only Adelta-fibre produced fewer voiding contractions in cyclophosphamide-treated rats, to a level similar to that from the control rats. These results support the ability of SNS to decrease bladder overactivity in a pathophysiological model of chemical irritation of the bladder.