Rabbit Corpus Cavernosum Strips Research Articles

Effects of eupatilin on the contractility of corpus cavernosal smooth muscle through nitric oxide-independent pathways.

Eupatilin (5,7-dihydroxy-3,4,6-trimethoxyflavone) is one of the main compounds present in Artemisia species. Eupatilin has both antioxidative and anti-inflammatory properties and a relaxation effect on vascular contraction regardless of endothelial function. We evaluated the relaxant effects of eupatilin on the corpus cavernosum (CC) of rabbits and the underlying mechanisms of its activity in human corpus cavernosum smooth muscle (CCSM) cells. Isolated rabbit CC strips were mounted in an organ bath system. A conventional whole-cell patch clamp technique was used to measure activation of calcium-sensitive K+ -channel currents in human CCSM cells. The relaxation effect of eupatilin was evaluated by cumulative addition (10-5 m ~ 3×10-4 m) to CC strips precontracted with 10-5 m phenylephrine. Western blotting analysis was performed to measure myosin phosphatase targeting subunit 1 (MYPT1) and protein kinase C-potentiated inhibitory protein for heterotrimeric myosin light chain phosphatase of 17-kDa (CPI-17) expression and to evaluate the effect of eupatilin on the RhoA/Rho-kinase pathway. Eupatilin effectively relaxed the phenylephrine-induced tone in the rabbit CC strips in a concentration-dependent manner with an estimated EC50 value of 1.2±1.6×10-4 m (n=8, p<0.05). Iberiotoxin and tetraethylammonium significantly reduced the relaxation effect (n=8, p<0.001 and p=0.003, respectively). Removal of the endothelium or the presence of L-NAME or indomethacin did not affect the relaxation effect of eupatilin. In CCSM cells, the extracellular application of eupatilin 10-4 m significantly increased the outward currents, and the eupatilin-stimulated currents were significantly attenuated by treatment with 10-7 m iberiotoxin (n=13, p<0.05). Eupatilin reduced the phosphorylation level of MYPT1 at Thr853 of MLCP and CPI-17 at Thr38. Eupatilin-induced relaxation of the CCSM cells via NO-independent pathways. The relaxation effects of eupatilin on CCSM cells were partially due to activation of BKCa channels and inhibition of RhoA/Rho-kinase.

Effect of a novel BKCa opener on BKCa currents and contractility of the rabbit corpus cavernosum.

Large-conductance Ca(2+)-activated K(+) (BKCa) channels are thought to play a key role in the regulation of corpus cavernosum smooth muscle (CCSM) excitability. Few BKCa channel openers have been accepted for clinical development. The effect of the novel BKCa channel opener GoSlo-SR5-130 on electrical activity in isolated rabbit CCSM cells and mechanical activity in strips of rabbit CCSM was examined. Single-channel currents were observed in inside-out patches. These channels were sensitive to Ca(2+), blocked by penitrem A, and had a conductance of 291 ± 20 pS (n = 7). In the presence of GoSlo-SR5-130, the number of open BKCa channels increased. Using voltage-ramp protocols, GoSlo-SR5-130 caused currents to activate at more negative potentials in a concentration-dependent manner, shifting the half-maximal activation voltage potential to the left on the voltage axis. Therefore, BKCa channels were open within the physiological range of membrane potentials in the presence of GoSlo-SR5-130. GoSlo-SR5-130 also resulted in an increase in the activity of spontaneous transient outward currents in myocytes isolated from CCSM, and this effect was reversed by iberiotoxin. In current-clamp mode, GoSlo-SR5-130 hyperpolarized the cell membrane. Isometric tension recording of strips of rabbit corpus cavernosum showed that GoSlo-SR5-130 inhibited spontaneous contractions in a concentration-dependent manner. This effect was reversed in the presence of iberiotoxin, suggesting that GoSlo-SR5-130 exerts its effect through BKCa channels. These findings suggest that GoSlo-SR5-130 is an effective tool for the study of BKCa channels and that these channels can modulate CCSM activity and are possible targets for the treatment of erectile dysfunction.

Nicotine in high concentration causes contraction of isolated strips of rabbit corpus cavernosum.

It is well known that cigarette smoke can cause erectile dysfunction by affecting the penile vascular system. However, the exact effects of nicotine on the corpus cavernosum remains poorly understood. Nicotine has been reported to cause relaxation of the corpus cavernosum; it has also been reported to cause both contraction and relaxation. Therefore, high concentrations of nicotine were studied in strips from the rabbit corpus cavernosum to better understand its effects. The proximal penile corpus cavernosal strips from male rabbits weighing approximately 4 kg were used in organ bath studies. Nicotine in high concentrations (10-5~10-4 M) produced dose-dependent contractions of the corpus cavernosal strips. The incubation with 10-5 M hexamethonium (nicotinic receptor antagonist) significantly inhibited the magnitude of the nicotine associated contractions. The nicotine-induced contractions were not only significantly inhibited by pretreatment with 10-5 M indomethacin (nonspecific cyclooxygenase inhibitor) and with 10-6 M NS-398 (selective cyclooxygenase inhibitor), but also with 10-6 M Y-27632 (Rho kinase inhibitor). Ozagrel (thromboxane A2 synthase inhibitor) and SQ-29548 (highly selective TP receptor antagonist) pretreatments significantly reduced the nicotine-induced contractile amplitude of the strips. High concentrations of nicotine caused contraction of isolated rabbit corpus cavernosal strips. This contraction appeared to be mediated by activation of nicotinic receptors. Rho-kinase and cyclooxygenase pathways, especially cyclooxygenase-2 and thromboxane A2, might play a pivotal role in the mechanism associated with nicotine-induced contraction of the rabbit corpus cavernosum.

Endothelium-Independent Relaxant Effect of Rubus Coreanus Extracts in Corpus Cavernosum Smooth Muscle

Rubus coreanus is a perennial shrub native to the southern part of the Korean peninsula. Although it is known that R. coreanus has a dose-dependent relaxation effect on rabbit corpus cavernosum (CC), the exact mechanism of action by which R. coreanus work is not fully known. To elucidate the direct effects of unripe R. coreanus extract (RCE) on CC smooth muscle cells. Dried unripe R. coreanus fruits were pulverized and extracted with 95% ethanol. Isolated rabbit CC strips were mounted in an organ-bath system, and the effects of RCE were evaluated. To estimate [Ca(2+)]i , we used a Fura-2 fluorescent technique. The effects of unripe RCE on ion channels and the intracellular Ca(2+) concentration ([Ca(2+)]i ) of CC. RCE effectively relaxed phenylephrine (PE)-induced tone in rabbit CC, and removal of the endothelium did not completely abolish the relaxation effect of RCE. Tetraethylammonium (1 mM) did not inhibit RCE-induced relaxation in strips precontracted by PE in the organ bath. However, CaCl2 -induced constriction of CC strips, bathed in Ca(2+)-free buffer and primed with PE, was abolished by RCE. In addition, RCE decreased basal [Ca(2+)]i in corporal smooth muscle cells. The increases of [Ca(2+)]i evoked by 60 mM K(+)-containing solution in A7r5 cells were suppressed by RCE, and RCE relaxed KCl-induced tone in endothelium-free CC, which indicated that RCE blocked the voltage-dependent Ca(2+) channels (VDCCs). RCE decreased basal [Ca(2+)]i and the [Arg8]-vasopressin-induced [Ca(2+)]i increases in A7r5 cells, and RCE inhibited the contraction of endothelium-free CC induced by PE in Ca(2+)-free solution, which suggested that RCE might act as a modulator of corporal smooth muscle cell tone by inhibiting Ca(2+) release from sarcoplasmic reticulum. RCE acts through endothelium-independent and endothelium-dependent pathways to relax CC. RCE may inhibit VDCCs and Ca(2+) release from sarcoplasmic reticulum.

Changed Phosphodiesterase Selectivity and Enhanced in vitro Efficacy by Selective Deuteration of Sildenafil

In order to explore whether selective deuteration of sildenafil affects selectivity and efficacy of the drug, the inhibitory activity of sildenafil (1-[4-ethoxy-3-(6,7-dihydro-1-methyl-7-oxo-3-propyl-1H-pyrazolo[4,3-d]pyrimidin-5-yl)phenyl-sulfonyl] -4-methylpiperazine citrate, CAS 139755-83-2) and three deuterated sildenafil derivatives, D8-piperazine-sildenafil (BDD-10402), D3-methyl-D8-piperazine-sildenafil (BDD-10403) and D5-ethoxy-Sildenafil (BDD-10406) against phosphodiesterases 1-6 was compared. Furthermore, the relaxant effect of sildenafil and its deuterated derivatives in a contractility assay on rabbit corpus cavernosum strips from New Zealand rabbits was investigated. BDD-10406 exhibits a 2-fold higher selectivity for phosphodiesterase 5 versus phosphodiesterase 6 than sildenafil. BDD-10406 and sildenafil inhibited cGMP formation with IC50 values of 6 nmol/L and 9 nmol/L, respectively. The corresponding IC80 values for BDD-10406 and Sildenafil were 33 nmol/L and 40 nmol/L, respectively. Sildenafil, BDD-10402, BDD-10403 and BDD-10406 relaxed the rabbit corpus cavernosum strips in a dose-dependent manner with ED50 values of 245 nmol/L, 91 nmol/L, 121 nmol/L and 85 nmol/L, respectively. Deuteration of sildenafil on the ethoxy group (BDD-10406) leads to enhanced selectivity for phosphodiesterase 5 versus phosphodiesterase 6 and higher efficacy in vitro. This is the first example of a deuteration effect on the inhibitory activity of a reversible enzyme inhibitor.

Characterizing Behavior of Corpus Cavernosum in Chloride-free Condition

To elucidate the role of chloride currents in erectile function through characterizing the behavior of corpus cavernosum (CC) in chloride-free (Cf) medium, which has not been evaluated before. Isolated rabbit CC strips were suspended in thermo-regulated organ baths containing oxygenated Tyrode for isometric tension recording. Cf Tyrode was prepared by substituting sodium chloride, calcium chloride, and potassium chloride (KCl) with equivalent molar concentrations of sodium acetate, calcium acetate, and potassium acetate salts. Resting cavernosal tone and contractions by noradrenaline, histamine, and KCl were assessed in Cf Tyrode with or without chloride channel blockers, niflumic acid (NFA), and anthracene-9-carboxylic acid (A9C). Withdrawal of extracellular chloride caused myogenic contractions in the unstimulated CC strips (n = 18). In addition, peak contractions by noradrenaline (n = 14) and histamine (n = 13) were augmented in Cf buffer by 47.2 ± 5.9% and 85.4 ± 13.2%, respectively (P <.05), whereas KCl contractions were not significantly altered (17.6 ± 4.6%; n = 7). Interestingly, Cf buffer exerted opposing effects, potentiation and reduction, respectively, on the plateau phase of contractions mediated by noradrenaline and histamine. The stimulatory effect of Cf buffer on the intrinsic myogenic tone was diminished by NFA (30 μM), and A9C (300 μM-1 mM). NFA (30-100 μM), however, specifically reduced the plateau phase without significantly modifying the peak contraction of noradrenaline in Cf buffer. These results reiterate the importance of chloride currents as a mechanism underlying the maintenance of penile cavernosal tone. Thus, chloride channel could be an effective alternative target to regulate penile erection.

1541 DISTAL CORPUS CAVERNOSUM SMOOTH MUSCLE FIBROSIS DUE TO NON-ISCHAEMIC PRIAPISM- SHOULD EMBOLISATION BE PERFORMED EARLIER?

INTRODUCTION AND OBJECTIVES: Non-ischaemic priapism is due to unregulated cavernous arterial inflow. Although rare, it is commonly associated with perineal or penile trauma. Since the venous outflow mechanism is unaffected the corpus cavernosum is perfused with oxygenated blood and therefore these patients can be managed conservatively. We report on our series of patients with non-ischaemic priapism who have developed fibrosis of the distal corpus cavernosum following a period of conservative treatment. METHODS: Over a 5 year period, 7 patients with non-ischaemic priapism presented to our unit. The mean age was 37.4 years (range 20 56). The etiology in 6 cases was secondary to a perineal injury and in 1 case conversion of an ischaemic priapism to a non ischaemic priapism following surgical intervention. The diagnosis of non-ischaemic priapism in all patients was based on the clinical history and examination, cavernosal blood gas analysis, colour duplex ultrasonography of the penis and pudendal angiography. 4 cases (57%) also underwent a penile MRI. RESULTS: Although the diagnostic tests confirmed non-ischaemic priapism, follow up Doppler studies and MRI penis showed the development of distal corpus cavernosum fibrosis. Pudendal artery super selective embolization was eventually performed in all of the patients, of which 2 (28.5%) patients underwent a second attempt at embolisation. On follow up these patients reported the development of erectile dysfunction due to poor distal penile tumescence in all cases and the penile MRI demonstrated distal corpus cavernosum fibrosis. Four patients (57%) eventually underwent insertion of a penile implant due to failed pharmacotherapies. A further 3 (43%) patients are using PDE-5 inhibitors. The conditions were reproduced in an in vitro model using precontracted strips of rabbit corpus cavernosum superfused at high pO2 levels. This showed that the smooth muscle tone reduced by 43% of the initial tone (n 4) after superfusion for 12 hours indicating irreversible smooth muscle dysfunction. CONCLUSIONS: The risk of ischaemic damage in non-ischaemic priapism is expected to be minimal due to the oxygenation of the corpus cavernosum being maintained. However, corpus cavernosum perfusion with higher than normal pO2 levels may result in oxidative stress due to the formation of reactive oxygen species (ROS). Irreversible damage to the smooth muscle is followed by the development of fibrosis, Based on these findings we suggest that superselective embolisation of these cases should be performed at the earliest opportunity.

The relaxation mechanisms of tetrandrine on the rabbit corpus cavernosum tissue in vitro

The relaxation mechanisms of tetrandrine (Tet) on the rabbit corpus cavernosum tissue in vitro were investigated. Strips of rabbit corpus cavernosum were mounted in organ chambers. The effects of Tet were examined on isolated muscle strips pre-contracted with phenylephrine (PE) alone, in the presence of N W-nitro-L-arginine (LNNA, a nitric oxide synthase inhibitor), 1-H-[1,2,4]oxadiazolo[4,3-α]quinoxalin-1-one(ODQ, a guanylyl cyclase inhibitor), indomethacin (cyclooxygenase inhibitor), tetraethylammonium (TEA, Ca2+-activated K+ channel blocker), 4-aminopiridine (4-AP, voltage dependent K+ channel blocker) and glibenclamide (ATP sensitive K+channel blocker). The effects of Tet on KCl-induced contraction of isolated muscle strips were also investigated. The procedure of calcium absence–calcium addition was designed to observe the effect of Tet on the two components of the contractile responses to PE based on the source of Ca2+ (extracellular vs. intracellular). Corpus cavernosum strips showed relaxation in response to Tet (10−8 ∼ 10−3 mol L−1) in a concentration-dependent manner with an IC50 of 3.73 × 10−5 mol L−1. However, they were not affected by LNNA, ODQ, indomethacin and K+-channel blockers. Tet (10 μmol L−1, 30 μmol L−1) concentration dependently reduced the maximal contraction response of isolated strips induced by KCl to (73.0 ± 3.8) and (41.5 ± 3.4)%, respectively (p < 0.01). In the procedure of calcium absence–calcium addition, Tet 100 μmol L−1 inhibited both intracellular calcium-dependent and extracellular calcium-dependent contraction induced by PE (20 μmol L−1) (p < 0.05). The inhibition ratios were (23.8 ± 7.1) and (40.7 ± 11.2)%, respectively. The results of the present study suggest that Tet possesses a relaxant effect on rabbit corpus cavernosum tissues, which is attributable to the inhibition of extracellular Ca2+ influx and the inhibition of release of intracellular-stored Ca2+, but not mediated by the release of nitric oxide, prostaglandins or by the activation of potassium channels.

Effect of hypothyroidism on the purinergic responses of corpus cavernosal smooth muscle in rabbits

Several studies have reported evidence of hormonal abnormalities in 25-35% of impotent men. Hypothyroidism has been reported to occur in 6% of impotent men. In the present study, we examined purinergic relaxation responses in hypothyroidism in an experimental rabbit model and compared them with controls to evaluate the possible involvement of the purinergic pathway. The study comprised 20 male New Zealand white rabbits. The rabbits were divided into two equal groups. We tested the effects of ATP, alpha beta ATP, and adenosine precontracted with phenylephrine on the isolated corpus cavernosum preparations from control and hypothyroid rabbits. We also evaluated the effects of ATP, alpha beta ATP, and adenosine on the cGMP levels in the isolated corpus cavernosum preparations from control and hypothyroid rabbits. T3, T4, and testosterone levels were significantly lower in hypothyroid rabbits. ATP, alpha beta ATP, carbachol, and electrical field stimulation (EFS)-induced frequency-dependent relaxation responses in the isolated rabbit corpus cavernosum strips precontracted with phenylephrine reduced significantly (P<0.05). Adenosine-induced relaxation responses did not change significantly in hypothyroid rabbits. Reduction of relaxation response in hypothyroid rabbits corpus cavernosum can depend on a decreased release of nitric oxide (NO) from nitrergic nerves and endothelium.

Relaxant effects of an alkaloid-rich fraction from Aspidosperma ulei root bark on isolated rabbit corpus cavernosum

We described earlier that an alkaloid-rich fraction (F(3-5)) from Aspidosperma ulei (Markgr) induces penile erection-like behavioral responses in mice. This study verified a possible relaxant effect of this fraction on isolated rabbit corpus cavernosum (RbCC) strips precontracted by phenylephrine (1 microM) or K+ 60 mM. F(3-5) (1-300 microg ml(-1)) relaxed the RbCC strips in a concentration-dependent and reversible manner. The relaxant effect of F(3-5) (100 microg ml(-1)) on phenylephrine contraction was unaffected in the presence of atropine, N-omega-nitro-L-arginine methyl ester or 1H-[1,2,4]oxadiazole[4,3-a] quinoxalin-1-one and by preincubation with tetrodotoxin, glibenclamide, apamine and charybdotoxin suggesting that mechanisms other than cholinergic, nitrergic, sGC activation or potassium channel opening are probably involved. However, the phasic component of the contraction induced by K+ 60 mM as well as the maximal contraction elicited by increasing external Ca2+ concentrations in depolarized corpora cavernosa was inhibited by F(3-5). We conclude that F(3-5) relaxes the RbCC smooth muscle, at least in part, through a blockade of calcium influx or its function.

Involvement of α-receptors and potassium channels in the mechanism of action of sildenafil citrate

Modulation of the adrenergic activity and interfering with channels such as potassium channels may affect relaxation and contraction of the corpus cavernosum. Sildenafil is a selective phosphodiesterase-5 inhibitor, proven effective in treating erectile dysfunction. In this study, the effect of sildenafil citrate on alpha-receptors modulation and potassium channels was tested. The direct relaxant effect of sildenafil citrate was studied by measuring changes in isometric tension in isolated strips of rabbit corpus cavernosum and rat aortic ring precontracted with phenylephrine or KCl compared to that of diazoxide in the presence and absence of tetraethylammonium. The inhibitory effect of sildenafil on electrical field stimulation-induced contraction of rabbit corpus cavernosum and rat anococcygeus muscle was also studied compared to that of phentolamine. Muscle relaxant effect of sildenafil (1 x 10(-9)-1 x 10(-6) M on phenylephrine-precontracted rabbit corpus cavernosum strips was not attenuated by N(G)-nitro-L-arginine (3 x 10(-5) M). Cumulative addition of sildenafil (1 x 10(-9)-1 x 10(-6) M) and phentolamine (1 x 10(-9)-1 x 10(-6) M) to the organ bath dose-dependently inhibited electrical field stimulation-induced contraction of rabbit corpus cavernosum and rat anococcygeus muscle, with almost similar EC(50) values. Sildenafil (1 x 10(-7) M) also inhibited phenylephrine-induced contraction of rat aortic rings by 39.83+/-3.01%. In addition, tetraethylammonium (1 x 10(-3) M) significantly attenuated the muscle relaxant effect of sildenafil (1 x 10(-9)-1 x 10(-6) M) on phenylephrine-precontracted strips of rabbit corpus cavernosum. Sildenafil citrate is capable of producing cavernosal smooth muscle relaxation by an additional mechanism that may involve alpha-receptors and potassium channel opening.

토끼 음경해면체평활근 이완 및 10주간 경구투여한 흰쥐의 생식기능에 미치는 KH204의 효과

KH204가 음경해면체 이완과 생식기능에 미치는 영향을 규명하기 위해 효능을 실험한 결과 토끼의 음경해면체를 1×10<SUP>-6</SUP> M pheylephrine으로 수축시킨 후 KH204를 61, 183 및 549 ㎎/L의 축적용량으로 처리하여 각각 6.8, 10.1 및 16%의 용량의존적인 이완효과를 관찰하였다. 흰쥐를 이용한 생식기능 실험은 KH204를 61, 183 및 549 ㎎/㎏으로 10주간 장기투여에서 생식장기인 고환조직의 중량 및 정자 수 증가에 유의한 효과를 나타내었다. 정자 형태검사는 대조군과 비슷한 수치로 나타내어 고농도로 장기투여에도 정자 기형에 문제가 없음을 확인하였다. 이상의 결과로 KH204가 특별한 부작용이 없이 음경해면체를 농도의존적으로 이완시키 효과가 있어, 음경발기를 개선시키는 효과뿐만 아니라 고환성 원인 또는 정자생산 기능부전으로 보이는 남성불임에서 또한 효능이 있음을 시사하였다.

Epimedium brevicornum Maxim extract relaxes rabbit corpus cavernosum through multitargets on nitric oxide/cyclic guanosine monophosphate signaling pathway

Epimedium brevicornum Maxim (EbM) has been reputed to have sexual stimulation effects on males. The study is aimed to test the hypothesis that EbM extracts relaxed the corpus cavernosum (CC) smooth muscle through activation of multitargets on nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signaling pathway. Water extract of EbM and its subfraction (EP-20) were prepared and standardized by high-performance liquid chromatography. Isolated rabbit CC strips were mounted in organ baths and isometric tension was recorded in the presence or absence of specific inhibitors related to NO/cGMP signaling such as L-N(G)-nitro-arginine methyl ester (L-NAME), 1H-[1,2,4]oxadiazolo-[4,3-a] quinoxalin-1-one (ODQ, a guanylyl cyclase inhibitor) or phosphodiesterase 5 (PDE 5) inhibitors. cGMP level was determined in EP-20-treated CC strips. The results showed that EP-20 enriched the content of L-arginine in the process of purification and relaxed the CC smooth muscle precontracted with phenylephrine (PE, 1 microM) in a concentration-dependent manner. Besides, EP-20 increased the amount of cGMP production in rabbit CC tissues. Coincubation with EP-20 and L-NAME or ODQ significantly decreased EP-20-induced relaxation whereas EP-20 increased sodium nitroprusside-induced relaxation in PE-precontracted CC strips. Besides, EP-20 increased the potency and the duration of the relaxation effects caused by electrical field stimulation. Finally, EP-20 could potentiate PDE 5 inhibitors in relaxation of PE-precontracted CC strips. We concluded that extract of EbM relax the CC smooth muscle through multitargets in NO/cGMP/PDE 5 pathway and might bring into perspective the treatment strategy for those patients with erectile dysfunction.

Effect of SA1, a Herbal Formulation, on Sexual Behavior and Penile Erection

SA1 is a mixture of 9 Oriental herbs (Korean red ginseng, fermented soybean, Tribulus terrestris, Fructus Rubi, Fructus Lycii, Semen Cuscutae, Dioscorea Rhizome, Fructus Corni and Fructus Crataegi) that are widely used as energizers and vitalizers in the indigenous system of medicine and have been alleged to improve the sexual functions in men. This study evaluated SA1 using both in vitro and in vivo experiments on laboratory animals in order to determine its effect on the sexual behavior and penile erection. The male rats used to examine the copulatory behavior were administered either the vehicle or SA1 (30, 100, 300, 600 mg/kg) orally for 2 weeks. The intracavernous pressure and systemic blood pressure were recorded in anesthetized rats. The responses to acetylcholine and SA1 of rabbit corpus cavernosum strips were also examined. There was an overall increase in the copulatory behavior parameters in the SA1-treated rats, which was reflected by a decrease in the mount and intromission latencies and an increase in the ejaculation latency and mount frequency. SA1 significantly increased the ratio of the intracavernous pressure to mean arterial pressure. In vitro, SA1 significantly enhanced the relaxation responses to acetylcholine. These results suggest that SA1 improves the sexual activity and erectile function.

Effects of ligustrazine on the contraction of isolated rabbit corpus cavernosum strips

To investigate the role of ligustrazine on relaxation of the isolated rabbit corpus cavernosum tissue in vitro, the effects of ligustrazine on the corpus cavernosum were observed by using experimental method of smooth muscle strips. Concentration-responses to phenylephine (PE) and KCl were recorded. The results showed that ligustrazine concentration-dependently depressed the contraction response of smooth muscle strips induced by PE. The maximum percentage relaxation of cavernosal strips by ligustrazine was 74.1% +/- 6.2% (compared with control: 21.9% +/- 5.6%, P < 0.01). Ligustrazine concentration-dependently reduced the amplitude of the contraction induced by cumulative doses of PE or KCl, shifted the cumulative concentration response curves of PE and KCI to the right and depressed their maximal responses. It was concluded that ligustrazine could significantly relax the cavernosal muscle contraction induced by PE in vitro. The results suggested that ligustrazine inhibited calcium ion influx.

Effects of platelet‐activating factor on nitrergic transmission in rabbit corpus cavernosum

The information currently available suggests that nonadrenergic noncholinergic (NANC) transmitters, particularly nitric oxide, are involved in the relaxation of penile erectile tissues. Platelet-activating factor (PAF) is a chemical mediator and is involved in many physiological and pathophysiological events. It is well known that several of the vascular actions of PAF are mediated by the generation of nitric oxide. We designed this study to test the hypothesis that PAF has an effect on NANC responses in rabbit corpus cavernosum strips. Rabbit corpus cavernosum strips were precontracted with phenylephrine (10(-5) mol/L). Isometric tension changes produced by carbachol (10(-9)-10(-5) mol/L), sodium nitroprusside (10(-8)-10(-5) mol/L) and electrical field stimulation (for 10 s at sequential frequencies of 2, 4, 8, 16, and 32 Hz as square-wave pulses of 50 mV) were recorded with a pressure transducer. These relaxations were compared to those obtained in the presence of PAF. PAF had no effect on endothelium-dependent, endothelium-independent or electrical field stimulation-induced NANC relaxation responses in isolated rabbit corpus cavernosum strips. There was no statistically significant difference between the pD(2) and E(max) values for carbachol or sodium nitroprusside in the presence of PAF. Our results suggest that PAF does not modify the endothelium-dependent, endothelium-independent or electrical field stimulation-induced NANC relaxation responses in isolated rabbit corpus cavernosum strips.

Relaxation effects of adrenomedullin in isolated rabbit corpus cavernosum smooth muscle.

To clarify the pharmacological effects of adrenomedullin, a potent vasodilator and hypotensive peptide isolated from human phaeochromocytoma cells, on corpus cavernosal smooth muscle in vitro, as the intracavernosal injection of adrenomedullin induces penile erection in the anaesthetized cat. The effects of adrenomedullin were investigated in isolated muscle strips from New Zealand rabbit corpus cavernosum smooth muscle pre-contracted with phenylephrine alone, in the presence of indomethacin (cyclooxygenase inhibitor), Nomega-nitro l-arginine methyl ester (L-NAME, a nitric oxide synthase inhibitor), and K+-channel blockers. Adrenomedullin caused relaxation of isolated pre-contracted rabbit corpus cavernosum strips in a concentration-dependent manner. The response of corpus cavernosum was unaffected L-NAME, indomethacin and K+-channel blockers. The relaxation exerted by adrenomedullin in rabbit corporal tissue may arise from the effect of the drug on its specific receptors and/or calcitonin gene-related peptide-1 receptors. The relaxant effect of adrenomedullin might lead to novel clinical applications for erectile dysfunction.

Investigation of the mechanism of nicotine induced relaxation in rabbit corpus cavernosum in vitro.

In order to determine whether nicotine acts on corporal smooth muscle, the mechanism of its effect on strips of rabbit corpus cavernosum was studied in vitro. Rabbit corpus cavernosum muscle strips were mounted in an organ bath with modified Krebs-Henseleit solution and aerated with 95% O(2) and 5% CO(2). Tension was measured with isometric force transducers, and muscle relaxation was expressed as the percent decrease of precontraction induced by phenylephrine. Nicotine produced concentration dependent relaxation when preparations were precontracted by phenylephrine (10(-5) M). The maximum nicotine-induced relaxation was 60.4 +/- 4.2% of the phenylephrine contraction and was not affected by indomethacin (10(-5) M), N(w)-nitro L-arginine methyl ester (3 x 10(-5) M), methylene blue (10(-5) M), glibenclamide (10(-5) M), clotrimazole (10(-6) M), tetraethylammonium (3 x 10(-4) M), or 4-aminopyridine (10(-3) M). Nicotine did not exhibit a calcium antagonizing effect. From these results, we conclude that nicotine-induced relaxation of the rabbit corpus cavernosum is not mediated by the release of nitric oxide, prostaglandins or a related substance, by the activation of potassium channels, or by the stimulation of nicotinic cholinoceptors. Further work is needed to determine the cellular mechanism(s) of the action by which nicotine acts on corporal smooth muscle.

Effects of NCX 4050, a new NO donor, in rabbit and human corpus cavernosum.

The effects of NCX 4050, a drug belonging to a new class of NO donors, was investigated in isolated preparations of human and rabbit corpus cavernosum (CC) and in human foetal corpora cavernosa (hfCC) smooth muscle cells. In strips of rabbit CC, NCX 4050 (0.001-100 microM) induced a concentration-dependent relaxation which was influenced neither by Nw-nitro-l-arginine-methyl-ester (l-NAME; 100 microm) nor by endothelium deprivation. The NCX 4050-induced relaxation was significantly reduced by the guanylate cyclase inhibitor 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 1 microm) and enhanced by a specific phosphodiesterase 5 inhibitor, sildenafil (300 nm). Moreover, NCX 4050 (0.01-1 microm), induced a concentration-dependent potentiation of the relaxant response induced by electrical field stimulation (EFS) in rabbit preparations pre-treated with guanethidine and indomethacin. The relaxant effect of NCX 4050 was similar to that obtained by increasing concentrations (0.001-100 microm) of sodium nitroprusside (SNP) in either rabbit or human preparations. To further investigate the activity of NCX 4050 on human corpora cavernosa, we exposed cultured hfCC smooth muscle cells to increasing concentrations of NCX 4050 and SNP. We found that both compounds dose-dependently reduced cell proliferation. The antiproliferative effect of all the concentration tested of NCX 4050 was completely blocked by ODQ (1 microm). These results suggest that in rabbit and human corpora cavernosa NCX 4050 acts by activating guanylate cyclase activity, induces smooth muscle relaxation and quiescence. Our results provide a rationale for a possible future use of NCX 4050 in the pharmacotherapy of erectile dysfunction linked to an impaired release of NO from the endothelium.

The Effect of The Specific Phosphodiesterase Type V Inhibitor, “Sildenafil Citrate”, on Alloxan-Induced Diabetic Rabbit Corpus Cavernosal Smooth Muscle

This study was carried out to investigate the effect of sildenafil (S) together with the influence of sodium nitroprusside (SNP) and acetylcholine (Ach) on isolated strips of rabbit corpus cavernosum. The in vitro effects of sildenafil on non-adrenergic, noncholinergic and nitric oxide (NO)-mediated cavernosal smooth muscle relaxation in diabetic rabbits, were also investigated. Diabetes mellitus was induced in adult male New Zealand white rabbits with alloxan (65 mg/kg, i.v). Cavernosal strips from agematched control and 5 months diabetic animals were mounted in organ baths containing Krebs-bicarbonate solution (pH 7.4, bubbled continueously with a gas mixture of 95% O2 plus 5% CO2, and maintained at 37oC. Isolated strips of control rabbit corpus cavernosum were stimulated isotonically with phenylephrine (10-5M) and relaxations were induced using increasing concentrations of Ach, SNP and S alone and in combination. Also, relaxation responses of control and diabetic strips precontracted with phenylephrine (10-5M) to electrical field stimulation (EFS, 4-20 Hz) or SNP (10-9 – 10-3M) were assessed in the absence and presence of a submaximal dose of sildenafil (10-6M). It was found that Ach, SNP and S relaxed the control rabbit corpus cavernosal strips in a dose-dependent manner. The relaxant effect of the test agents were ranked (from least inhibitory to most inhibitory) as follows: S, Ach plus S, SNP, SNP plus S, Ach. Moreover, it was observed that SNP-stimulated relaxations were significantly impaired in the corpus cavernosum from diabetic group compared to age-matched controls. Sildenafil (10-6M) significantly enhanced SNP-stimulated relaxation in control and diabetic groups. Electrical field stimulation-mediated relaxations of the corpus cavernosum were significantly impaired after 5 months diabetes mellitus and enhanced by sildenafil (10-6M). In conclusion, sildenafil enhances the relaxing effect of both SNP and Ach on the phenylephrine-induced contraction of rabbit cavernosal tissue. It could also be suggested that the impairment of NO-mediated relaxation of the diabetic corpus cavernosum reflect, at least in part, a defect in guanyl cyclase activity. These findings support the use of sildenafil as an effective, orally administered, treatment for diabetic erectile dysfunction.