Extracorporeal Shock Waves Therapy Delivered by Aries Improves Erectile Dysfunction in Spontaneously Hypertensive Rats Through Penile Tissue Remodeling and Neovascularization.

BACKGROUND: Low-intensity extracorporeal shock wave therapy (Li-ESWT) has been reported to improve erectile function in patients with moderate-to-severe erectile dysfunction (ED) or even convert phosphodiesterase type 5 inhibitors nonresponders to responders. ED is highly prevalent in hypertensive patients. The effect of Li-ESWT on an animal model of hypertension-associated ED has not been reported. AIM: To investigate the effect of Li-ESWT on hypertension-associated ED and provide plausible mechanisms of action of Li-ESWT on local mechanisms of penile erection. METHODS: Spontaneously hypertensive rats (SHRs) in the active group (n = 13) received Li-ESWT at energy flux density 0.06 mJ/mm2 (Aries; Dornier MedTech, Wessling, Germany) twice weekly for 6 weeks. The emitter was set to zero for SHRs in the sham group (n = 12). Erectile function was assessed 4 weeks post-treatment by monitoring intracavernosal pressure (ICP) in response to electrical stimulation of cavernous nerve before and after single dose of 0.3 mg/kg intravenous sildenafil. Cavernosal tissue was then evaluated for collagen/smooth muscle content, neuronal nitric oxide synthase (nNOS), and vascular endothelial factor (CD31) expression. OUTCOMES: Erectile function was assessed with ICP, erectile tissue remodeling was studied by smooth muscle/collagen ratio, nNOS and CD31 were semiquantitatively evaluated on cavernosal sections. RESULTS: The improvement of ICP parameters was greater in Li-ESWT-treated rats compared with controls with and without sildenafil. Sildenafil led to 20% increase in area under the intracavernosal pressure curve measured during the entire response/mean arterial pressure at 10 Hz in ESWT_SHR + sildenafil compared with ESWT_SHR. The smooth muscle/collagen ratio increased 2.5-fold in Li-ESWT compared with sham. Expression of CD31 tended to be increased whereas nNOS was unchanged. CONCLUSIONS: Li-ESWT by Aries may represent an effective noninvasive therapeutic alternative and a relevant add-on therapy to phosphodiesterase type 5 inhibitors for ED in hypertensive patients, and it is suggested that it acts via remodeling of the penile tissue and promoting cavernosal vascularization. Assaly R, Giuliano F, Clement P, et al. Extracorporeal Shock Waves Therapy Delivered by Aries Improves Erectile Dysfunction in Spontaneously Hypertensive Rats Through Penile Tissue Remodeling and Neovascularization. Sex Med 2019;7:441-450.

The Favorable Effect of Empagliflozin on Erectile Function in an Experimental Model of Type 2 Diabetes.

INTRODUCTION: Following the results of the EMPA-REG Outcome trial, we hypothesized that empagliflozin, a highly potent and specific sodium/glucose cotransporteur 2 inhibitor, could improve type 2 diabetes mellitus (T2DM)-associated erectile dysfunction (ED), a highly prevalent complication of T2DM, very often coexisting with cardiovascular complications and considered as a prognostic factor of cardiovascular disease in men with diabetes. AIM: To investigate the effects of chronic treatment with empagliflozin on ED in a T2DM rat model in the presence or absence of sildenafil. METHODS: Male Goto-Kakizaki (GK), a model of T2DM, and age-matched Wistar rats received placebo or empagliflozin treatment at 25.3 ± 0.9 mg/kg/d for 4 weeks. Then, the in vivo effect of empagliflozin on erectile function was assessed by electrical stimulation of the cavernous nerve at different frequencies under anesthesia in the presence or absence of acute intravenous injection of sildenafil. Endothelium-dependent, -independent, and nitrergic relaxations of cavernosal strips from the rats were studied. MAIN OUTCOME MEASURES: Body weight, food consumption, metabolic parameters, plasma inflammation biomarkers, and in vivo erectile responses elicited by electrical stimulation of the cavernous nerve in empagliflozin-treated and untreated GK rats and control Wistar rats were assessed and followed by concentration or frequency response curves to endothelium-dependent, -independent, and nitrergic relaxations of cavernosal strips from these rats. RESULTS: Chronic empagliflozin followed by acute sildenafil significantly improved erectile responses in adult GK rats (n = 12-15/group). Ratios of intracavernous pressure and area under the curve/mean arterial pressure during the electrical stimulation were significantly increased in empagliflozin-treated vs untreated GK rats. Nitrergic relaxations of cavernosal strips from GK rats were significantly increased with empagliflozin compared with placebo. Moreover, the effect of sildenafil on erectile function was not altered by empagliflozin treatment. CLINICAL IMPLICATIONS: Empagliflozin may benefit T2DM patient with ED. STRENGTHS & LIMITATIONS: The mechanism(s) by which empagliflozin shows favorable effect on erectile function in GK rats needs to be further elucidated. CONCLUSION: Empagliflozin shows favorable effect on erectile function in diabetic GK rats mediated by an improvement of nitrergic relaxation of erectile tissue. Whether this favorable effect on ED in the experimental context of T2DM is due to better glycemic control or to another effect of empagliflozin deserves further investigation. Assaly R, Gorny D, Compagnie S, et al. The Favorable Effect of Empagliflozin on Erectile Function in an Experimental Model of Type 2 Diabetes. J Sex Med 2018;15:1224-1234.

Human spinal ejaculation generator.

OBJECTIVE: A spinal ejaculation generator (SEG) has been identified in the rat with lumbar galaninergic interneurons playing a pivotal role (Science 2002;297:1566-1569). The aim was to evidence a SEG in humans. METHODS: Spatial distribution of galaninergic neurons was studied in postmortem spinal cord segments of 6 men and compared with that of 6 women for evidencing sexual dimorphism. Based on the identified segmental distribution of galaninergic neurons, the ability for penile vibratory stimulation (PVS) to elicit ejaculation when the concerned spinal segments were injured was studied in 384 patients with clinically complete spinal cord injury (SCI) and consequent anejaculation. Such patients represent a unique model to investigate the role of defined spinal segments in the control of ejaculation. RESULTS: Galaninergic neurons were mostly located between L2 and L5 segments in medial lamina VII, with a maximal density within L4. Three-dimensional 3D reconstruction showed that these neurons were grouped into single columns bilaterally to the central canal. In addition, galaninergic neuron density was found higher in L3 and L4 segments in men as compared to women supporting sexual dimorphism. In the patients' cohort, injury of L3-L5 segments was the sole independent predictor for failure of PVS to induce ejaculation. Although evidence from clinical observations was indirect, there is close correspondence to neuroanatomical data. INTERPRETATION: Organization and sexual dimorphism of human spinal galaninergic neurons were similar to the rat's SEG. Neurohistological data, together with clinical results, corroborate the existence of an SEG in humans in L3-L5 segments. Such a generator could be targeted to treat neurogenic and non-neurogenic ejaculatory disorders. ANN NEUROL 2017;81:35-45.

Low Intensity Extracorporeal Shock Wave Therapy Improves Erectile Function in a Model of Type II Diabetes Independently of NO/cGMP Pathway.

PURPOSE: Erectile dysfunction is highly prevalent in type II diabetes mellitus. Low intensity extracorporeal shock wave therapy improves erectile function in patients with erectile dysfunction of vasculogenic origin, including diabetes. However, its mode of action remains unknown. We investigated the effects of low intensity extracorporeal shock wave therapy compared to or combined with sildenafil on erectile dysfunction in a type II diabetes mellitus model. Our purpose was to test our hypothesis of a mode of action targeting the cavernous nitric oxide/cyclic guanosine monophosphate pathway. MATERIALS AND METHODS: GK rats, a validated model of type II diabetes mellitus, and age matched Wistar rats were treated with low intensity extracorporeal shock wave therapy twice weekly for 3 weeks. Treatment was repeated after a 3-week no-treatment interval. The penis was stretched and dipped in a specifically designed water-filled cage. Shock waves were delivered by a calibrated probe yielding a controlled energy flux density (0.09 mJ/mm(2)). The probe was attached to an electrohydraulic unit with a focused shock wave source, allowing for accurate extrapolation to humans. Following a 4-week washout period erectile function was assessed as well as endothelium dependent and independent, and nitrergic relaxations of the corpus cavernosum of GK rats. RESULTS: Low intensity extracorporeal shock wave therapy significantly improved erectile function in GK rats to the same extent as sildenafil. Treatment effects were potentiated when combined with sildenafil. Shock wave effects were not associated with improved cavernous endothelium dependent or independent, or nitrergic reactivity. CONCLUSIONS: Low intensity extracorporeal shock wave therapy improved erectile function in GK rats. Unexpectedly, this was not mediated by a nitric oxide/cyclic guanosine monophosphate dependent mechanism. Sildenafil increased shock wave efficacy. This preclinical paradigm to deliver low intensity extracorporeal shock wave therapy to the rat penis should help further exploration of the mode of action of this therapy on erectile tissue.

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.

The spinal control of ejaculation revisited: a systematic review and meta-analysis of anejaculation in spinal cord injured patients.

BACKGROUND After spinal cord injury (SCI), most men cannot ejaculate without medical assistance. A major advance in the knowledge of the spinal control of ejaculation has been achieved with the discovery of a spinal generator of ejaculation (SGE) in the rat. The aim of this report was to review studies about ejaculation after SCI in order to revisit the spinal control of ejaculation and especially to assess the existence of an SGE in man. METHODS Studies were identified from Embase, PubMed, EBSCOhost and Cochrane Library. Studies were eligible when they specify the occurrence of antegrade ejaculation as a function of the neurological characterization of SCI. Studies were excluded when ejaculation was elicited by rectal electrical stimulation or when ejaculation could not be discriminated from climax. Meta-analyses were performed to assess the reference ejaculation rates for each procedure used to elicit ejaculation, i.e. masturbation or coïtus, penile vibratory stimulation (PVS) or acetylcholine esterase (AchE) inhibitors prior to masturbation. Subgroup analyses were performed according to the procedure used to elicit ejaculation on (i) the completeness of the SCI and (ii) the upper and lower limits of the SCI. To assess the existence of an SGE, the effect of concurrent lesions of different spinal segments was assessed by means of a stratified bivariate analysis. RESULTS From 523 studies, 45 were selected (n = 3851). Ejaculation occurred in response to masturbation or coïtus, PVS or AchE inhibitors followed by masturbation in, respectively, 11.8% (n = 1161), 47.4% (n = 597) and 54.7% (n = 309) of patients with complete SCI and in, respectively, 33.2% (n = 343), 52.8% (n = 305) and 78.1% (n = 32) of patients with incomplete SCI. Ejaculation, in the case of complete lesion of the sympathetic centres (T12 to L2), of the parasympathetic and somatic centres (S2-S4) or of all spinal ejaculation centres (T12 to S5) occurred in response to PVS in none of the patients (respectively, n = 5, n = 4 and n = 21) and in response to AchE inhibitors followed by masturbation in 4.9% (n = 61), 30.8% (n = 26) and 0% (n = 16) of the patients, respectively. Ejaculation in response to PVS or AchE inhibitors prior to masturbation was rhythmic forceful in 97.9% (n = 48) of the patients with complete lesion strictly above Onuf's nucleus (segments S2-S4). Complete lesion of the S2-S4 segments precluded the occurrence of rhythmic forceful ejaculation (n = 5). Controlling for the number of the injured segments between T12 and L2, the ejaculation rate sharply decreased when the lesion extended to the L3 segment and below. CONCLUSIONS The results reinforce the crucial roles of the spinal sympathetic and parasympathetic centres for emission and the somatic centre for expulsion. The spinal segments between L2 and S2 is more than a pathway to connect the ejaculation centres and likely harbours an SGE in man located in the L3, L4 and L5 segments.

Inhibition of ejaculation by the non-peptide oxytocin receptor antagonist GSK557296: a multi-level site of action.

BACKGROUND AND PURPOSE: Oxytocin (OT) plays a major role in the control of male sexual responses. Notably, blockade of OT receptors has been reported to inhibit ejaculation in animals. The study aimed to investigate the action of a highly selective, non-peptide OT antagonist GSK557296 in a model of pharmacologically induced ejaculation in anaesthetized rats. The site of action was assessed by investigating different delivery routes for this compound. EXPERIMENTAL APPROACH: Urethane-anaesthetized Wistar rats were implanted with a cerebral ventricle cannula for i.c.v. injections or with a subdural catheter for intrathecal (i.t.) GSK557296 injections. Occurrence of ejaculation was assessed following i.v. 7-hydroxy-2-(di-N-propylamino)tetralin (7-OH-DPAT), a dopamine D3 receptor agonist. In addition, seminal vesicle pressures (SVP) and bulbospongiosus muscle (BS) EMG were recorded as physiological markers of emission and expulsion phases of ejaculation respectively. KEY RESULTS: Highest i.v. GSK557296 dose reduced occurrence of ejaculation and increases in SVP but had no effect on BS-EMG. I.c.v. GSK557296 dose dependently inhibited ejaculation, increases in SVP and BS contractions. At spinal thoracic level, GSK557296 dose dependently inhibited ejaculation and increases in SVP but BS-EMG was impaired only with the highest dose. When delivered at lumbar level, GSK557296 dose dependently inhibited ejaculation, increases in SVP and BS contractions. CONCLUSIONS AND IMPLICATIONS: In the 7-OH-DPAT-induced ejaculation model, GSK557296 acts peripherally and centrally to inhibit ejaculation with different modalities. Blockade of brain OT receptors seems to be the most effective mechanism of action. Targeting central OT receptors with highly selective antagonist seems a promising approach for the treatment of premature ejaculation.

Effect of dapoxetine on ejaculatory performance and related brain neuronal activity in rapid ejaculator rats.

INTRODUCTION: A brain network specifically activated when ejaculation occurs has been described in rats. Increasing serotonin (5-hydroxytryptamine [5-HT]) tone impairs ejaculation and chronic 5-HT selective serotonin reuptake inhibitors (SSRIs) are known to inhibit ejaculation. However, efficacy of acute treatment with SSRI varies from one compound to another. The SSRI dapoxetine has been reported to delay ejaculation when given on demand to men with premature ejaculation (PE), although the mechanism of action is unclear. Effects of acute SSRIs on activity of the brain ejaculation circuit in relation with ejaculation have never been examined. AIM: To test the effects of acute administration of the short half-life SSRI dapoxetine on ejaculatory performance and activity in brain ejaculation circuit in rapid ejaculator rats taken as PE model. METHODS: Standard copulatory test was used to attribute one sexual category (sluggish, middle, or rapid) to male rats on the basis of their ejaculatory performance. Parameters of sexual, including ejaculatory, behavior, and Fos level of expression in discrete brain areas were assessed in the three sexual categories and in rapid category following acute oral treatment with dapoxetine. MAIN OUTCOME MEASURES: Ejaculation frequency (EF) and latency (EL) were measured as primary end points of ejaculatory behavior. Density of Fos-immunopositive cells in specific brain areas of brain stem, hypothalamus, and thalamus was determined as marker of neuronal activity. RESULTS: EL and Fos level of expression in hypothalamic and thalamic structures were found related. Dapoxetine acute oral administration (300 mg/kg) to rapid ejaculator rats resulted in (i) diminution of ejaculatory performance (lengthened EL and decreased EF); and (ii) modulation of Fos level of expression in hypothalamic and thalamic nuclei of the brain ejaculatory circuit. CONCLUSION: Acute treatment with dapoxetine, which reduced ejaculatory performance in rapid ejaculator rats, was also accompanied with changes in neuronal activity in components of the brain ejaculatory network.

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.

Combination of BAY 60-4552 and vardenafil exerts proerectile facilitator effects in rats with cavernous nerve injury: a proof of concept study for the treatment of phosphodiesterase type 5 inhibitor failure.

BACKGROUND: Radical prostatectomy (RP) is frequently responsible for erectile dysfunction (ED). Post-RP patients often show a failure to respond to phosphodiesterase type 5 (PDE5) inhibitors. OBJECTIVE: The acute effect of BAY 60-4552, the soluble guanylate cyclase (sGC) stimulator, and vardenafil were evaluated alone or in combination on erectile responses to electrical stimulation of the cavernous nerve (ES CN) in rats with cavernous nerve (CN) crush injury-induced ED. DESIGN, SETTING, AND PARTICIPANTS: Male adult Sprague-Dawley rats underwent laparotomy (sham, n=10) or bilateral CN crush injury (n=56). After 3 wk of recovery, erectile function was evaluated under urethane anaesthesia following ES CN at different frequencies. MEASUREMENTS: The acute effects of intravenous (IV) injection of vehicle, vardenafil 0.03 mg/kg, BAY 60-4552 0.03 mg/kg or 0.3 mg/kg, or a BAY 60-4552 0.03 mg/kg plus vardenafil 0.03 mg/kg combination were evaluated in CN-crushed rats. RESULTS AND LIMITATIONS: Bilateral CN crush injury followed by a 3-wk recovery period decreased erectile responses to ES CN by about 50%. In CN-crushed rats, IV vardenafil 0.03 mg/kg and BAY 60-4552 (0.03 or 0.3 mg/kg) increased erectile responses to ES CN to the same extent: Δ intracavernosal pressure/mean arterial pressure (ICP/MAP) at 10 Hz ES CN was 21±1% after vehicle, 25±3% (p<0.001) after vardenafil, and 26±5% and 27±5% after BAY 60-4552 0.03 mg/kg (p<0.01) and 0.3 mg/kg (p<0.001), respectively. The combination of vardenafil with BAY 60-4552 in CN-crushed rats totally restored erectile responses to ES CN equivalent to sham rats (ΔICP/MAP at 10 Hz ES CN: 34±4% after BAY 60-4552/vardenafil combination vs 39±4% in sham rats; not significant). CONCLUSIONS: The present study supports the concept that the combined administration of a sGC stimulator, BAY 60-4552, and vardenafil provides synergistic beneficial effects and might therefore salvage patients who experience treatment failures with PDE5 inhibitors after RP.

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.

Neuroanatomical distribution of the melanocortin-4 receptors in male and female rodent brain.

The melanocortin-4 receptor (MC4-R) plays a critical role in several physiological functions, from food intake, energy homeostasis, neuroendocrine and cardiovascular function, to sexual responses. The brain regions and the central neuronal pathways mediating the different actions of MC4-R remain largely unknown. We aimed to use immunocytochemistry using a specific antibody against rat MC4-R, to establish the detailed neuroanatomical distribution of MC4-R in brain slices of male and estrous female rats. We demonstrated that MC4-R-positive neurons were widely distributed in several brain regions including the cortex, thalamus, hypothalamus, and brainstem. In both male and female brains, MC4-R-positive cells were especially abundant in the hypothalamus, including the paraventricular hypothalamic nucleus, lateral septal nucleus, arcuate nucleus, supraoptic nucleus, medial preoptic area and lateral hypothalamic area. A moderate number of MC4-R-positive neurons were found in the piriform cortex, bed nucleus of the stria terminalis, medial and basolateral nuclei of amygdala, periaqueductal gray, red nucleus and raphe nucleus. A dimorphic sexual difference in the number of MC4-R-positive neurons was observed in some brain regions. In the medial preoptic area and arcuate nucleus, MC4-R-positive neurons were significantly more abundant in female than in males, whereas in the lateral hypothalamus the opposite proportion was observed. This is the first time the neuroanatomical distribution, and sex differences, of brain MC4-R localisation have been described. The distribution of MC4-R is consistent with the proposed roles of MC4-R-positive neurons and provides further information about the circuitry controlling food intake, energy balance and sexual responses in both males and females.

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.

How does chronic sildenafil prevent vascular oxidative stress in insulin-resistant rats?

INTRODUCTION: Insulin resistance features both endothelial dysfunction and increased oxidative stress. Both disorders are targeted by a chronic treatment with sildenafil. However, the mechanism of action by which chronic sildenafil exerts its effects on reactive oxygen species sources is still largely unknown. AIM: We therefore investigated how chronic sildenafil administration could impact vascular endothelial NO and superoxide release in a rat model of insulin resistance induced by fructose overload. METHODS: Adult male Wistar rats were fed a fructose-enriched diet (fructose-fed rats [FFR]) for 9 weeks. From weeks 6-8, sildenafil was administered subcutaneously twice daily (20 mg/kg), followed by a 1-week washout. MAIN OUTCOME MEASURES: Vascular endothelial NO and superoxide release were monitored in vitro in thoracic aortic segments using oxidative fluorescence. Specific inhibitors were used to distinguish the respective role of the main superoxide-producing systems within the vascular wall (i.e., mitochondrial respiratory chain and NADPH oxidases). The levels of expression of eNOS, Akt, and NADPH oxidase subunits were determined in the abdominal aorta. RESULTS: Chronic sildenafil administration corrected hyperglycemia, hyperinsulinemia, and hypertriglyceridemia in FFR. Moreover, after 9 weeks of diet, while global unstimulated aortic endothelial NO and superoxide release were unchanged in FFR, the relative contribution of the mitochondrial respiratory chain and NADPH oxidases was modified. Chronic sildenafil treatment, even after the 1-week washout period, was able to increase endothelial NO release independently of Akt-dependent phosphorylation by up-regulating eNOS expression, and restored the relative contribution of each superoxide-producing system examined, yielding endothelial superoxide release. Finally, in vitro incubation of aortic segments with sildenafil markedly decreased the endothelial aortic superoxide release. CONCLUSIONS: The present study showed that chronic sildenafil produced sustained vascular antioxidant effects in insulin-resistant rats by increasing NO release and regulating vascular superoxide release, supporting therefore further investigations using chronic sildenafil administration in preventing cardiovascular alterations associated with oxidative stress.

Differences in the spinal command of ejaculation in rapid ejaculating rats.

INTRODUCTION: It has been hypothesized that lifelong premature ejaculation is part of a biological variation in the intravaginal ejaculation latency, but what causes this variation remains poorly understood. AIM: The aim of this study is to elucidate whether variations in ejaculation latencies in an experimental rat model for premature ejaculation are linked to differences in the spinal command of ejaculation. MAIN OUTCOME MEASURES: Electrical microstimulation of the spinal generator for ejaculation revealed an accelerated expulsion phase in rapid ejaculating rats. METHODS: Adult male Wistar rats were categorized as "sluggish,""normal," or "rapid" ejaculators on the basis of their ejaculation frequency in sexual mating tests. One to three weeks after selection, males were urethane anesthetized and electrically microstimulated in the spinal generator for ejaculation, evoking ejaculation. Bulbospongiosus muscle electromyographic and intraluminal vas deferens pressure were measured simultaneously, representing, respectively, the expulsion and emission phase in ejaculation. RESULTS: Electrical microstimulation of the spinal generator for ejaculation evoked ejaculation in "sluggish" (N = 9), "normal" (N = 13), and "rapid" (N = 11) ejaculating rats. Vas deferens contraction (emission phase) was evoked at different stimulation strengths, but response properties were not statistically different between "sluggish,""normal," and "rapid" ejaculator rats. Bulbospongiosus muscle contractions (expulsion phase) following microstimulation was significantly accelerated in "rapid" rats as compared with "sluggish" and "normal" rats. The total duration of bulbospongiosus muscle contractions remained unchanged between the three ejaculator groups. CONCLUSIONS: Our results provide the first scientific evidence supporting a neurophysiological difference between "rapid,""normal," and "sluggish" ejaculators, expressed as an accelerated expulsion phase in "rapid" ejaculator rats. This bridges the gap between a sexual behavior trait and the spinal command of ejaculation.

Role of the neurokinin-1 receptors in ejaculation in anesthetized rats.

INTRODUCTION: Several lines of evidence indicate a role for substance P in the control of ejaculation, although its mode of action needs to be clarified. AIM: The effects and sites of action of a selective antagonist for the substance P-preferred receptor (neurokinin-1 receptor subtype; NK1) were investigated in a pharmacological model of ejaculation. METHODS: Ejaculation was induced in anesthetized rats by intracerebroventricular (i.c.v.) delivery of the dopamine D3 receptor preferring agonist 7-hydroxy-2-(di-N-propylamino)tetralin (7-OH-DPAT). The effects of the selective NK1 antagonist RP67580 on 7-OH-DPAT-induced ejaculation were measured following intraperitoneal (i.p.), i.c.v., or intrathecal (i.t.) (third lumbar spinal segment; L3) administration. MAIN OUTCOME MEASURES: Intraseminal vesicle pressure (SVP) and electromyogram of the bulbospongiosus muscle (BS) were recorded as physiological markers of emission and expulsion phases of ejaculation, respectively. RESULTS: Upon i.p., i.c.v., or i.t. administration, RP67580 significantly reduced the occurrence of ejaculation elicited by 7-OH-DPAT. A mild decrease in the occurrence of SVP and BS responses was observed in rats treated ip with RP67580, whereas only SVP responses were moderately affected following i.c.v. or i.t. administration. CONCLUSION: These results show the multilevel regulation of 7-OH-DPAT-induced ejaculation by NK1 receptors.

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).