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Rho-kinase inhibitors in the treatment of erectile dysfunction
249 RHO-KINASE INHIBITORS DYSFUNCTION Reea
Rowland’Cellek ,
‘Urology, University
Selim’.
IN THE
Moncada
TREATMENT
Salvador’.
OF ERECTILE
Ralph David’
Institute of Urology, London, United Kingdom, College London, London, United Kingdom
250 ALFUZOSIN POTENTIATES THE PROERECTILE APOMORPHINE IN SPONTANEOUS HYPERTENSIVE Ramirez-Gil
‘Wolfson
Institute,
J.F.*, Pouyet T.I. Mavoux
E.‘, Galzin A.M.‘,Arbilla
‘Research-Internal Medicine, sanofi-synthelabo, ‘Medical, sanofi-synthelabo, Gentilly, France
EFFECTS RATS
OF
S.’
Rueil-Malmaison,
France,
INTRODUCTION & OBJECTIVES: Noradrenergic neurotransmiasion plays an important role in the regulation of cavernosal smooth muscle tone. It is now known that noradrenergic contraction in the cavernosal smooth muscle cell? involves activation of the Rho-kinase pathway. which is a calcium-sensitisinc mechanism. Inhibition of Rho-kinase causes relaxation of the smooth muscle, particularly in the organs with a high basal tone. Therefore. Rho-kinase antagonism has been suggested as a novel therapeutic approach to erectile dysfunction. We aimed to investigate the presence and the activity of this pathway in penile ca\‘ernosal smooth muscle.
INTRODUCTION & OBJECTIVES: With the aging of the population, the prevalence of benign prostate hypertrophy (BPH) increases. In addition there is growing evidence suggesting that BPH is a risk factor for erectile dysfunction (ED). While the uroselective al-adrenoceptor antagonist, alfuzosin, is commonly used in the treatment of BPH patients, apomorphine emerged recently as a new treatment for ED. Therefore the purpose of this study was to evaluate the combination of these two compounds on the erectile function in rats.
MATERIAL & METHODS: Primary culture of smooth muscle cells from human and rabbit penile corpus cavernosum was dcvcloped. The presence of Rho-kinase was investigated by indirect imtnunolluoreacence and Western blotting. The activity of the immunoprecipitatcd protein wab determined usmp non-specific kinase assay. The effect of an inhibitor of Rho-kinaTe, (+)-(R)-ran+4.(I-aminoethyl)-N-(4.pyridyl) cyclohexane carboxamidedihydrochloridcmonohydrate (Y-27632), on the contractions elicited by noradrenergic nerve stimulation and by phenylephrine wan investigated in the human and rabbit penile corpus cavernosum in vitro.
MATERIAL & METHODS: lntracavemous pressure (ICP) and blood pressure (BP) were recorded continuously in male anaesthetised spontaneous hypertensive rats (SHR) after subcutaneous administration of apomorphine (IO250 pgikg). Alfuzosin (3-30 ygikg) was then given (20 min after apomorphine) by i.v. route and the effects followed over 30 minutes. RESULTS: Apomorphine, at IO and 20 pgikg s.c., produced erections in many rats but not in all, while rhythmic erection was triggered in all rats from the dose
RESULTS: The cells in culture showed characteristic myocyte morphology and a-actin immunoreactivity. Positive immunostaining for Rho-hinase was observed in the pcrinuclear space. Western blotting showed positive bands corresponding to Rhokmase in human and rabbit material. A specific inhibitor of Rho-kinasc, Y-27632, inhibned in a concentration-dependent manner the kinase activity of the protein immunoprecipitnted wth anti-Rho-kinase antibody. In both human and rabbit corpus caverno\um. EFS-induced noradrenergic contractions were inhibIted by Y-27632 in a concentration-dependent manner. Y-27632 caused concentration-dependent relaxation of phenylephrine-contracted tissues.
50 ;dgikg. Alfuzosin alone did not induce any erectile event, whatever the dose. However, when combining apomorphine (IO-100 pgikg) with alfuzosin (30 /‘g/kg). the number of erectile events increased significantly when compared to
CONCLUSIONS:
alfuzosin
apomorphine alone. Apomorphine, at 50@g/kg, increased ICP up to 29*7 mmHg while decreasing the blood pressure which gave an ICPiBP ratio of 36*60/o. In the presence of alfuaosin. at doses (3-30 p&/kg) known to exhibit a marked relaxant effect on urethra in rats, the ICPiBP ratio was increased up to 58+3’S, In agreement with these observations. the overall ICP measured as the area under the curve was increased by 46 to 275% with alfuzosin. The average number of erections per rat was increased from 2.5Sti.37 to 4*0.4 with (30 Irgikg)
when combined
with SO pglkg
of apomorphine.
These rebulth suggest that Rho-kinaae he involved in noradrenrrgic contractile pathway in the cavemosal smooth muscle of the penis. Cavernosal smooth muscle cells can provide a cellular model for the study of enzymes involved in Ca’+-scnsitising pathways regulatmg cavernosal smooth muscle tone. Rho-kinabe antagonism represents a potential therapeutic use for the treatment of erectlIe dysfunction.
CONCLUSIONS: These results indicate that alfuaosin improves the proerectile properties of apomorphine by increasing the number and the amplitude of erections. Therefore the association of both drugs could be of interest in the treatment of BPH patients facing erectile dysfunction.
251
252
PATHOPHYSIOLOGICAL MECHANISMS OF ERECTILE DYSFUNCTION IN HYPERTENSIVE RATS Behr-Roussel Katell’. Benoit
Delphine’,Chamiot-Clerc GCrard’, Giuliano
Philippe’, Francois’
EFFECTS OF HORMONAL REPLACEMENT ON GENITAL AROUSAL IN OVARIECTOMISED FEMALE RATS
BemabC Jacques’.
‘Domaine INRA, Pelvipharm Laboratories. BuresiYvette, France, Groupe de Recherche en Urologic. Le Kremlin-Bic&tre, France
Mevel
‘Urology,
INTRODUCTION:
Many studies have demonstrated that men with hypertension (HTN) have a significantly higher prevalence of erectile dysfunction than the general population. Moreover, the question has been raised as to whether the higher rate of sexual dysfunction in hypertensive individuals is due to HTN and/or to drug treatment of HTN.
OBJECTIVES
& METHODS: We aimed to evaluate the erectile function measured by the frequency-dependent increase in intracavemosal pressure (ICP) following electrical stimulation of the cavernous nerve (CN) in viva (O-IO Hz) and explore in vitro the corporal reactivity to contractile and relaxant agonists in male spontaneously hypertensive rats (SHR. I2 weeks) and age-matched nommotensive Wistar-Kyoto rats (WKY). RESULTS: CN stimulations induced frequency-dependent increases in ICP in both WKY (n=5) and SHR (n=S). These erectile responses were drastically reduced in SHR compared to WKY rats (responses to IO Hz-6V of 23.3+3.7% vs. 46.l*l.9%, p
Chamiot-Clerc Philippe’, Mevel Katell’, Benoit GCrard’. Giuliano Francoia’
Compagnie
SEXUAL
Sandrine' , BernabC
‘Domaine INRA, Pelvipharm Laboratories, BuresiYvette, de Recherche en Urologic. Le Kremlin-Bidtre. France
France. Qrology,
Jacques’, Groupe
OBJECTIVES: We aimed to assess the consequences of the hormonal deprivation and replacement in our model of rat female genital sexual arousal (FGSA) in response to pelvic nerve stimulation (PNS). Bilateral ovariectomy (OVX) was performed under MATERIAL & METHODS: isollurane anaesthesia in IO week old female Sprague-Dawley rats. Animals were allowed to recover during 2 weeks before being given subcutaneous pellets with estradiol (E2)(0.5 mg) or E2+ progesterone (PG)(33.5 mg) or vehicle (Veh) for 6 weeks. Then, under urethane anaesthesia. a catheter was inserted into the carotid artery for blood pressure (MAP) recording. The left PN was exposed and placed on bipolar platinum stimulating electrodes. Vagmal engorgement was measured by monitoring vaginal blood flow using Laser Doppler Perfusion Measurements (LDPM-arbitrary units). The variation in temperature (DT-“C) in the vagina was measured using miniature thermocouples located in the rectum and in the vaginal lumen. Vaginal contractile activity (VCA)(&) was recorded using force transducers. The effect of PNS (square-waves pulses. lms. 6V. 3Os)(2-IOHr) on the different parameters was studied. RESULTS: MAP is not significantly modified in by E2 or E2+PG comparatively to Veh group (84.5+2.5 mmHg. 85.8k4.4 and 85.l+J.Y mmHg respectively), whereas basal vaginal LDPM 1s significantly increased (162.0~4.0,201.4~3.5 and 250.9?10.7 in Veh, E2- and E2+PG-treated animals respectively; p
European
Urology
Supplements
1 (2002) No. 1, pp. 65
Rowland’Cellek ,
‘Urology, University
Selim’.
IN THE
Moncada
TREATMENT
Salvador’.
OF ERECTILE
Ralph David’
Institute of Urology, London, United Kingdom, College London, London, United Kingdom
250 ALFUZOSIN POTENTIATES THE PROERECTILE APOMORPHINE IN SPONTANEOUS HYPERTENSIVE Ramirez-Gil
‘Wolfson
Institute,
J.F.*, Pouyet T.I. Mavoux
E.‘, Galzin A.M.‘,Arbilla
‘Research-Internal Medicine, sanofi-synthelabo, ‘Medical, sanofi-synthelabo, Gentilly, France
EFFECTS RATS
OF
S.’
Rueil-Malmaison,
France,
INTRODUCTION & OBJECTIVES: Noradrenergic neurotransmiasion plays an important role in the regulation of cavernosal smooth muscle tone. It is now known that noradrenergic contraction in the cavernosal smooth muscle cell? involves activation of the Rho-kinase pathway. which is a calcium-sensitisinc mechanism. Inhibition of Rho-kinase causes relaxation of the smooth muscle, particularly in the organs with a high basal tone. Therefore. Rho-kinase antagonism has been suggested as a novel therapeutic approach to erectile dysfunction. We aimed to investigate the presence and the activity of this pathway in penile ca\‘ernosal smooth muscle.
INTRODUCTION & OBJECTIVES: With the aging of the population, the prevalence of benign prostate hypertrophy (BPH) increases. In addition there is growing evidence suggesting that BPH is a risk factor for erectile dysfunction (ED). While the uroselective al-adrenoceptor antagonist, alfuzosin, is commonly used in the treatment of BPH patients, apomorphine emerged recently as a new treatment for ED. Therefore the purpose of this study was to evaluate the combination of these two compounds on the erectile function in rats.
MATERIAL & METHODS: Primary culture of smooth muscle cells from human and rabbit penile corpus cavernosum was dcvcloped. The presence of Rho-kinase was investigated by indirect imtnunolluoreacence and Western blotting. The activity of the immunoprecipitatcd protein wab determined usmp non-specific kinase assay. The effect of an inhibitor of Rho-kinaTe, (+)-(R)-ran+4.(I-aminoethyl)-N-(4.pyridyl) cyclohexane carboxamidedihydrochloridcmonohydrate (Y-27632), on the contractions elicited by noradrenergic nerve stimulation and by phenylephrine wan investigated in the human and rabbit penile corpus cavernosum in vitro.
MATERIAL & METHODS: lntracavemous pressure (ICP) and blood pressure (BP) were recorded continuously in male anaesthetised spontaneous hypertensive rats (SHR) after subcutaneous administration of apomorphine (IO250 pgikg). Alfuzosin (3-30 ygikg) was then given (20 min after apomorphine) by i.v. route and the effects followed over 30 minutes. RESULTS: Apomorphine, at IO and 20 pgikg s.c., produced erections in many rats but not in all, while rhythmic erection was triggered in all rats from the dose
RESULTS: The cells in culture showed characteristic myocyte morphology and a-actin immunoreactivity. Positive immunostaining for Rho-hinase was observed in the pcrinuclear space. Western blotting showed positive bands corresponding to Rhokmase in human and rabbit material. A specific inhibitor of Rho-kinasc, Y-27632, inhibned in a concentration-dependent manner the kinase activity of the protein immunoprecipitnted wth anti-Rho-kinase antibody. In both human and rabbit corpus caverno\um. EFS-induced noradrenergic contractions were inhibIted by Y-27632 in a concentration-dependent manner. Y-27632 caused concentration-dependent relaxation of phenylephrine-contracted tissues.
50 ;dgikg. Alfuzosin alone did not induce any erectile event, whatever the dose. However, when combining apomorphine (IO-100 pgikg) with alfuzosin (30 /‘g/kg). the number of erectile events increased significantly when compared to
CONCLUSIONS:
alfuzosin
apomorphine alone. Apomorphine, at 50@g/kg, increased ICP up to 29*7 mmHg while decreasing the blood pressure which gave an ICPiBP ratio of 36*60/o. In the presence of alfuaosin. at doses (3-30 p&/kg) known to exhibit a marked relaxant effect on urethra in rats, the ICPiBP ratio was increased up to 58+3’S, In agreement with these observations. the overall ICP measured as the area under the curve was increased by 46 to 275% with alfuzosin. The average number of erections per rat was increased from 2.5Sti.37 to 4*0.4 with (30 Irgikg)
when combined
with SO pglkg
of apomorphine.
These rebulth suggest that Rho-kinaae he involved in noradrenrrgic contractile pathway in the cavemosal smooth muscle of the penis. Cavernosal smooth muscle cells can provide a cellular model for the study of enzymes involved in Ca’+-scnsitising pathways regulatmg cavernosal smooth muscle tone. Rho-kinabe antagonism represents a potential therapeutic use for the treatment of erectlIe dysfunction.
CONCLUSIONS: These results indicate that alfuaosin improves the proerectile properties of apomorphine by increasing the number and the amplitude of erections. Therefore the association of both drugs could be of interest in the treatment of BPH patients facing erectile dysfunction.
251
252
PATHOPHYSIOLOGICAL MECHANISMS OF ERECTILE DYSFUNCTION IN HYPERTENSIVE RATS Behr-Roussel Katell’. Benoit
Delphine’,Chamiot-Clerc GCrard’, Giuliano
Philippe’, Francois’
EFFECTS OF HORMONAL REPLACEMENT ON GENITAL AROUSAL IN OVARIECTOMISED FEMALE RATS
BemabC Jacques’.
‘Domaine INRA, Pelvipharm Laboratories. BuresiYvette, France, Groupe de Recherche en Urologic. Le Kremlin-Bic&tre, France
Mevel
‘Urology,
INTRODUCTION:
Many studies have demonstrated that men with hypertension (HTN) have a significantly higher prevalence of erectile dysfunction than the general population. Moreover, the question has been raised as to whether the higher rate of sexual dysfunction in hypertensive individuals is due to HTN and/or to drug treatment of HTN.
OBJECTIVES
& METHODS: We aimed to evaluate the erectile function measured by the frequency-dependent increase in intracavemosal pressure (ICP) following electrical stimulation of the cavernous nerve (CN) in viva (O-IO Hz) and explore in vitro the corporal reactivity to contractile and relaxant agonists in male spontaneously hypertensive rats (SHR. I2 weeks) and age-matched nommotensive Wistar-Kyoto rats (WKY). RESULTS: CN stimulations induced frequency-dependent increases in ICP in both WKY (n=5) and SHR (n=S). These erectile responses were drastically reduced in SHR compared to WKY rats (responses to IO Hz-6V of 23.3+3.7% vs. 46.l*l.9%, p
Chamiot-Clerc Philippe’, Mevel Katell’, Benoit GCrard’. Giuliano Francoia’
Compagnie
SEXUAL
Sandrine' , BernabC
‘Domaine INRA, Pelvipharm Laboratories, BuresiYvette, de Recherche en Urologic. Le Kremlin-Bidtre. France
France. Qrology,
Jacques’, Groupe
OBJECTIVES: We aimed to assess the consequences of the hormonal deprivation and replacement in our model of rat female genital sexual arousal (FGSA) in response to pelvic nerve stimulation (PNS). Bilateral ovariectomy (OVX) was performed under MATERIAL & METHODS: isollurane anaesthesia in IO week old female Sprague-Dawley rats. Animals were allowed to recover during 2 weeks before being given subcutaneous pellets with estradiol (E2)(0.5 mg) or E2+ progesterone (PG)(33.5 mg) or vehicle (Veh) for 6 weeks. Then, under urethane anaesthesia. a catheter was inserted into the carotid artery for blood pressure (MAP) recording. The left PN was exposed and placed on bipolar platinum stimulating electrodes. Vagmal engorgement was measured by monitoring vaginal blood flow using Laser Doppler Perfusion Measurements (LDPM-arbitrary units). The variation in temperature (DT-“C) in the vagina was measured using miniature thermocouples located in the rectum and in the vaginal lumen. Vaginal contractile activity (VCA)(&) was recorded using force transducers. The effect of PNS (square-waves pulses. lms. 6V. 3Os)(2-IOHr) on the different parameters was studied. RESULTS: MAP is not significantly modified in by E2 or E2+PG comparatively to Veh group (84.5+2.5 mmHg. 85.8k4.4 and 85.l+J.Y mmHg respectively), whereas basal vaginal LDPM 1s significantly increased (162.0~4.0,201.4~3.5 and 250.9?10.7 in Veh, E2- and E2+PG-treated animals respectively; p
European
Urology
Supplements
1 (2002) No. 1, pp. 65