Rabbit Corpus Cavernosum Smooth Muscle Research Articles

The role of Ca2+-activated Cl- current in tone generation in the rabbit corpus cavernosum.

Rabbit corpus cavernosum smooth muscle (RCCSM) cells express ion channels that produce Ca2+-activated Cl- (IClCa) current, but low sensitivity to conventional antagonists has made its role in tone generation difficult to evaluate. We have reexamined this question using two new generation IClCa blockers, T16Ainh-A01 and CaCCinh-A01. Isolated RCCSM cells were studied using the perforated patch method. Current-voltage protocols revealed that both L-type Ca2+ current and IClCa T16Ainh-A01 and CaCCinh-A01 (10 μM) reduced IClCa by ~85%, while 30 μM abolished it. L-type Ca2+ current was unaffected by 10 μM CaCCinh-A01 but was reduced by 50% at 30 μM CaCCinh-A01, 46% at 10 μM T16Ainh-A01, and 78% at 30 μM T16Ainh-A01. Both drugs reduced spontaneous isometric tension in RCCSM strips, by 60-70% at 10 μM and >90% at 30 μM. Phenylephrine (PE)-enhanced tension was also reduced (ED50 = 3 μM, CaCCinh-A01; 14 μM, T16Ainh-A01). CaCCinh-A01 at 10 μM had little effect on 60 mM KCl contractures, though they were reduced by 30 μM CaCCinh-A01 and T16Ainh-A01 (10 μM and 30 μM) consistent with their effects on L-type Ca2+ current. Both drugs also reversed the stimulatory effect of PE on intracellular Ca2+ waves, studied with laser scanning confocal microscopy in isolated RCCSM cells. In conclusion, although both drugs were effective blockers of IClCa, the effect of T16Ainh-A01 on L-type Ca2+ current precludes its use for evaluating the role of IClCa in tone generation. However, 10 μM CaCCinh-A01 selectively blocked IClCa versus L-type Ca2+ current and reduced spontaneous and PE-induced tone, suggesting that IClCa is important in maintaining penile detumescence.

Relaxation Effects of Eucomiae Cortex in Isolated Rabbit Corpus Cavernosum Smooth Muscle

This study was aimed to investigate the relaxation effects of Eucomiae Cortex (EC) extract in isolated rabbit corpus cavernosum smooth muscle and its mechanism. To evaluate the relaxation of EC extract in rabbit corpus cavernosum, EC extract was treated in corporal strips which were precontracted with phenylephrine(PE). To study its mechanism, Nω-nitro-L-arginine (L-NNA) was pretreated after infuse of EC extract and compared with non-treated. In calcium chloride (CaSUP2+/SUP) -free krebs solution, EC extract and CaSUP2+/SUP 1 mM were infused by turns after CaSUP2+/SUP 1 mM was treated into corporal strips contracted by PE. Cell ability, nitric oxide (NO) and epithelial nitric oxide synthase (eNOS) on human umbilical vein endothelial cell (HUVEC) were measured by MTT assay, Griess reagent system and histochemical, immunohistochemical methods. EC extract showed a significant relaxation effects on the corporal strips, this effects were inhibited by pretreatment of L-NNA. EC extract inhibited the increase of contraction by CaSUP2+/SUP influx in CaSUP2+/SUP-free krebs solution, and eNOS positive reaction in corpus cavernosum, NO production in HUVEC increased by treatment of EC extract. These result suggest that the relaxation effects of EC extract in isolated corpus cavernosum smooth muscle are involved in increase of eNOS and NO production, blocking of extracellular CaSUP2+/SUP influx.

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.

腎氣丸의 남성 생식기능 및 성기능 개선효과 : 정자생성 촉진, 고환독성 완화 및 음경해면체 평활근의 이완

Objectives : This study aimed to investigate the effects of Shingi-whan(SG) on the male reproductive and sexual function, so we measured the spermatogenesis and the testicular toxicity in mice and the vasorelaxation in isolated rabbit corpus cavernosum smooth muscle. Methods : To evaluate effect on the spermatogenesis in mice, we prepared two groups, control group and SG group that was orally administered SG(1,000mg/kg) for 20 days, and compared. To analyze testicular toxicity in mice, we also prepared two groups, doxo group that was injected with doxorubicin (3mg/kg) on three times and doxo + SG group that was injected with doxorubicin and SG for 20 days, and compared. To investigate sexual function of SG in mice, we prepared three groups, normal group and aging elicited group consisting of 18-month-old mice, SG treated aging group that was orally administered SG for 60 days, and compared using histochemical staining on mice corpus cavernous tissues. In order to define the relaxation effects of SG, rabbit corpus cavernous tissues were prepared in <TEX>$2{\times}2{\times}6mm$</TEX> sized strip. Then the dose-dependent relaxation responses of SG at 0.01-3.0 mg/ml in contracted strips induced by phenylephrine were measured. Results : The sperm density in dutus epididymis and the diameter of seminiferous tubules of SG group was significantly increased when compared to control group. The testicular weight and the diameter and height of epithelial layer of seminiferous tubules of doxo + SG group was significantly increased when compared to doxo group. The cavernous strips were significantly relaxed by SG extract In SG treated aging group, ratio of smooth muscles to collagen fibers and red blood cell count in venous sinus was increased as compared to aging elicited group. Conclusions : Our findings have shown that SG extract have effect on spermatogenesis and mitigating effect on doxo-induced testicular toxicity. Further, it also have the vasorelaxant effect on rabbit corpus cavernosum.

Effects of Shingi-whan on the Male Reproductive and Sexual Function : Enhancing Spermatogenesis, Reducing Testicular Toxicity, and Relaxing Smooth Muscle of Corpus Cavernosum

Objectives : This study aimed to investigate the effects of Shingi-whan(SG) on the male reproductive and sexual function, so we measured the spermatogenesis and the testicular toxicity in mice and the vasorelaxation in isolated rabbit corpus cavernosum smooth muscle. Methods : To evaluate effect on the spermatogenesis in mice, we prepared two groups, control group and SG group that was orally administered SG(1,000mg/kg) for 20 days, and compared. To analyze testicular toxicity in mice, we also prepared two groups, doxo group that was injected with doxorubicin (3mg/kg) on three times and doxo +SG group that was injected with doxorubicin and SG for 20 days, and compared. To investigate sexual function of SG in mice, we prepared three groups, normal group and aging elicited group consisting of 18-month-old mice, SG treated aging group that was orally administered SG for 60 days, and compared using histochemical staining on mice corpus cavernous tissues. In order to define the relaxation effects of SG, rabbit corpus cavernous tissues were prepared in 2 × 2 × 6 mm sized strip. Then the dose-dependent relaxation responses of SG at 0.01-3.0 mg/ml in contracted strips induced by phenylephrine were measured. Results : The sperm density in dutus epididymis and the diameter of seminiferous tubules of SG group was significantly increased when compared to control group. The testicular weight and the diameter and height of epithelial layer of seminiferous tubules of doxo + SG group was significantly increased when compared to doxo group. The cavernous strips were significantly relaxed by SG extract In SG treated aging group, ratio of smooth muscles to collagen fibers and red blood cell count in venous sinus was increased as compared to aging elicited group. Conclusions : Our findings have shown that SG extract have effect on spermatogenesis and mitigating effect on doxo-induced testicular toxicity. Further, it also have the vasorelaxant effect on rabbit corpus cavernosum.

Relaxation Effects of Epimedium Koreanum Nakai in Isolated Rabbit Corpus Cavernosum Smooth Muscle

This study aimed to investigate the relaxation effects and its underlying mechanisms of Epimedium koreanum Nakai(EK) in phenylephrine(PE) treated isolated rabbit corpus cavernosum smooth muscle. The dose-dependent relaxation responses of phenylephrine(PE, 1×10<SUP>-6</SUP> M)-precontracted strips to EK at 0.01-3.0 ㎎/㎖ were measured and also observed after endothelial denudation using organ bath. To analyze the underlying mechanisms of EK-induced relaxation, Nω-nitro-L-arginine(L-NNA), methylene blue(MB), tetraethylammonium chloride(TEA), indomethacin(IM) were pretreated before EK extract infused into precontracted strips induced by PE. To investigate cytotoxic activity and nitric oxide(NO) concentration of EK extract on EA.hy926 cells, mitochondrial dehydrogenase activity(MTT) assay and nitric oxide detection kit were used. The cavernous strips were significantly relaxed by EK extract at 0.3 ㎎/㎖, 1.0 ㎎/㎖, 3.0 ㎎/㎖ and the relaxation responses of PE-precontracted strips denuded endothelium also inhibited in comparison with intact endothelium. The pretreatment of L-NNA, MB, TEA reduced EK extract-induced endothelium-dependent relaxation, but the pretreatment of IM didn"t affect EK extract-induced endothelium-dependent relaxation. When EK extract was applicated on EA.hy926 cells, the NO concentration was increased. Our findings have shown that EK extract exerts a relaxing effect on corpus cavernosum in part by suppressing influx of extracellular Ca<SUP>2+</SUP> through activating the NO-cGMP system.

A medico-legal problem of child maltreatment among pediatric cases admitted to the Burn Unit of Alexandria Main University Hospital

To investigate expression of ryanodine receptors (RyRs) in rabbit penile corpus cavernosum smooth muscle (CCSM) cells. New Zealand White Rabbit CCSM cells were cultured by primary tissue culture. Using CCSM cells and fibroblast have different adherence velocity, CCSM cell can be purified. Identification of CCSM cell was by inverted microscope and immunofluorescence technique. The cDNA sequence of RyRs was found out by searching genebank. Three pair of primers were designed by Primer Premier 5.0. The RyRs subunits mRNA was detected by reverse transcription PCR in cultured CCSM cells. After 7d, we found growth of cultured cells. While 15 to approximately 20 d, cells filled the bottom of culture flask. They were identified by inverted microscope and immunofluorescence technique. After purification, purity of CCSM cells was near to 100%. It suggested only RyRs1 subunit was expressive in CCSM by RT-PCR. RyRsl subunit is expressed in CCSM cells. It suggests that RyRs contribute to the regulation of Ca2+ in CCSM cells.

Relaxation Effects of Rubus coreanus in Isolated Rabbit Corpus Cavernosum Smooth Muscle

This study aimed to investigate the relaxation effects and its underlying mechanisms of Rubus coreanus(RC) extract in contracted rabbit corpus cavernous tissues by phenylephrine(PE) 1 μM. In order to define the relaxation effects of RC, rabbit corpus cavernous tissues were prepared in 2×2×6 ㎜ sized strip. The dose-dependent relaxation responses of RC at 0.01-3.0 ㎎/㎖ in contracted strips induced by PE were measured and also observed after endothelial denudation. To analyze the underlying mechanisms of RC-induced relaxation, indomethacin(IM), tetraethylammonium chloride(TEA), Nω-nitro-L-arginine (L-NNA), methylene blue(MB) were treated before RC extract infused into precontracted strips induced by PE. To study the effect of RC extract on influx of extracellular Ca<SUP>2+</SUP> in corpus cavernous strips, calcium chloride(Ca) 1 mM infused into precontracted strips induced by PE after pretreatment of RC extract in Ca<SUP>2+</SUP>-free krebs-ringer solution. To investigate cytotoxic activity and nitric oxide(NO) concentration of RC extract on human umbilical vein endothelial cell(HUVEC), cell viability on HUVEC was measured by MTT assay, and NO concentration was measured by Griess reagent system. The cavernous strips were significantly relaxed by RC extract at 1.0 ㎎/㎖, 3.0 ㎎/㎖ and the relaxation responses to RC were inhibited significantly by endothelial disruption. The pretreatment of IM, TEA didn"t affect RC extract-induced endothelium -dependent relaxation, but the pretreatment of L-NNA, MB reduced RC extract-induced endothelium-dependent relaxation. When Ca<SUP>2+</SUP> was supplied the cavernous strips which were precontracted by PE in a Ca<SUP>2+</SUP>-free krebs-ringer solution, contraction of strips was increased, but pretreatment of RC inhibited contractile response to Ca<SUP>2+</SUP>. When RC extract was applicated on HUVEC, NO concentration was increased. Our findings show that RC extract exerts a relaxing effect on corpus cavernosum in part by suppressing influx of extracellular Ca<SUP>2+</SUP> through activating the NO-cGMP system.

Chronic administration of imipramine but not agomelatine and moclobemide affects the nitrergic relaxation of rabbit corpus cavernosum smooth muscle

Sexual dysfunction is a common and underestimated effect of antidepressants. However, the mechanism by which these drugs cause erectile dysfunction is unclear. We investigated the reactivity of the corpus cavernosum of rabbits that were treated with either chronic imipramine, which is a tricyclic agent; agomelatine, which is a melatonergic agonist and serotonin 5HT2c antagonist; or moclobemide, which is a reversible inhibitor of monoamine-oxidase A. Twenty rabbits were randomly divided into four groups: the control group (n=5), the imipramine-treated group (n=5), which received i.p. injections of 10mg/kg/day of imipramine, the moclobemide-treated group (n=5), which received i.p. injections of 20mg/kg/day of moclobemide, and the agomelatine-treated group (n=5), which was orally administered 10mg/kg/day of agomelatine. The reactivities of corpus cavernosum tissue obtained from the antidepressant-treated and the control groups were studied in organ chambers after the animals were subjected to 21 days of drug administration. The acetylcholine-induced endothelium-dependent and the electrical field stimulation (EFS)-induced neurogenic relaxation of the corpus cavernosum of the imipramine-treated group was significantly decreased compared with the control group. However, neither the acetylcholine- nor EFS-induced relaxation was changed in the moclobemide- or agomelatine-treated groups. There were no change in the relaxant response to the nitric oxide (NO) donor sodium nitroprusside and contractile response to KCl between the groups. This study suggests that chronic imipramine treatment but not agomelatine and moclobemide treatments causes significant functional changes in the penile erectile tissue of rabbits and that these changes may contribute to the development of impotence.

Effects of Korean ginseng berry extract (GB0710) on penile erection: evidence from in vitro and in vivo studies

Several reports have promoted the root-derived Korean red ginseng (KRG; Panax ginseng) as alternative treatment for erectile dysfunction (ED), and ginsenosides are known to be the principal active ingredients of ginseng. Recent studies showed that ginseng berries produce more ginsenosides than KRG; thus, we investigated the ability of the Korean ginseng berry extract GB0710 to relax the penile corpus cavernosum smooth muscle (CCSM) in this study. As a comparative control, the results were compared to those obtained using KRG. In addition, possible mechanisms of action for GB0710 were investigated. While KRG and GB0710 both displayed dose-dependent relaxation effects on precontracted rabbit CCSM in vitro, GB0710 was shown to be more potent than KRG. The GB0710-induced relaxation could be partially reduced by removing the endothelium. In addition, pre-treatment with several nitric oxide (NO) inhibitors significantly inhibited the relaxation of muscle strips. Furthermore, administration of GB0710 increased intracavernosal pressure (ICP) in a rat in vivo model in both a dose- and duration-dependent manner. Intracellular NO production in human microvascular endothelial cells could be induced by GB0710 and inhibited by N(G)-monomethyl-L-arginine. In conclusion, GB0710 had a greater relaxation effect on rabbit CCSM than did KRG extract, and increased ICP in a rat model in both a dose- and a duration-dependent manner. This relaxing effect might be mediated by NO production.

In vitro and in vivo aphrodisiac properties of Corchorus depressus Linn. on rabbit corpus cavernosum smooth muscle relaxation and sexual behavior of normal male rats

Ethnopharmacological relevanceCorchorus depressus Linn. has been used as an aphrodisiac in traditional Indian medicine to treat male sexual dysfunction and impotency. Aim of the studyThe petroleum ether, chloroform, ethyl acetate, n-butanol and aqueous fractions of 95% methanol extract of Corchorus depressus were screened initially for their in vitro aphrodisiac activity on rabbit corpus cavernosum smooth muscle. The chloroform fraction (CDC) was found to be the most active and therefore investigated further on general mating behavior, libido and potency of normal male Wistar albino rats in comparison with the standard drug, Sildenafil citrate. Materials and methodsAnimals were divided into the following groups: Control, SC CDC 100, CDC 200, and CDC 400, who received saline, Sildenafil citrate or the chloroform fraction of Corchorus depressus at doses of 100, 200 or 400mg/kg b.wt., respectively. The route of administration for all the groups was oral dosing, which was once in a day for 45 days. To analyze the mating behavior, female rats with estrus phase were used. ResultsThe chloroform fraction of methanolic extract of Corchorus depressus significantly reduced ML, IL, PEI and III. There was a significant increase in the MF, IF and EL and serum testosterone levels throughout the study period. The potency test significantly increased erections, quick flips, long flips and total reflex. In vitro aphrodisiac activity was significantly higher in chloroform fraction at a concentration of 25.0mg/ml, which induced 71.4% relaxation. The combined results of the above mentioned models indicate that the chloroform fraction of Corchorus depressus produces a significant increase in sexual activity as exhibited by 25mg/ml in vitro and 400mg/kg in vivo. In comparison with the control, all the drug-treated groups have shown drug-induced effects for a few parameters. ConclusionsIn vitro and in vivo studies provide valuable experimental evidence that the chloroform fraction of methanolic extract of Corchorus depressus possesses aphrodisiac property. This study further substantiates the ethnopharmacological claims of Corchorus depressus as a sexual stimulating agent and offers a significant potential for studying the effect on male sexual response and its dysfunctions.

The relaxant effect of methanolic extract of yohimbe bark (&amp;lt;i&amp;gt;Pausinystalia yohimbe&amp;lt;/i&amp;gt;) on rabbit corpus cavernosum smooth muscles

Background: Development and maintenance of penile erection requires the relaxation of corpus cavernosum smooth muscle (CCSM) cells and it is essentially mediated by nitric oxide (NO). Pausinystalia yohimbe bark is traditionally used for the treatment of erectile dysfunction (ED). Objectives and Methods: This study aimed to evaluate the effects of the methanolic extract of P. yohimbe bark (10 mg/ml) on isolated rabbit corpus cavernosum smooth muscle (CCSM). Results: Methanolic extract of P. yohimbe bark blocked contractions in CCSM induced by adrenergic agonists, through facilitating NO release from the endothelial cells. Conclusion: The obtained results revealed that, methanolic extract of P. yohimbe bark caused relaxation of CCSM that validates its traditional aphrodisiac property, thus it could be used to initiate and maintain erection in cases of erectile dysfunction.

Chronic administration of fluoxetine impairs neurogenic and endothelium-dependent relaxation of the rabbit corpus cavernosum smooth muscle

Antidepressants, including selective serotonin reuptake inhibitors (SSRIs), cause erectile dysfunction; however, the mechanism by which they cause erectile function is unclear. We investigated the reactivity of the corpus cavernosum after chronic fluoxetine treatment in rabbits. Twelve rabbits were randomly divided into two groups: control (n=6) or 20mg/kg/day of fluoxetine delivered i.p. (n=6). The reactivity of the corpus cavernosum tissue from the fluoxetine-treated and control groups was studied in organ chambers after 21 days of fluoxetine injection. In the fluoxetine-treated group, endothelium-dependent relaxation of the corpus cavernosum in response to acetylcholine was significantly decreased compared to the control group. However, the sensitivity (i.e., pD(2)) of the fluoxetine-treated cavernosal tissue strips to acetylcholine was not changed with respect to controls. Electrical field stimulation (EFS)-induced neurogenic relaxation was also significantly reduced in the fluoxetine-treated group. Relaxation in response to the nitric oxide (NO) donor sodium nitroprusside was similar between the cavernosal tissues from the two groups. There was also no change in agonist potency between the two groups. Additionally, chronic fluoxetine treatment had no effect on KCl-induced contractile responses. When tissue contraction was produced with phenylephrine to study relaxation in response to various stimuli, the tension induced was similar between the fluoxetine-treated and control groups. This study suggests that chronic fluoxetine treatment causes significant functional changes to the penile erectile tissue of rabbits, and these changes may contribute to the development of impotence.

Erectogenic Effects ofClerodendron capitatum: Involvement of Phosphodiesterase Type-5 Inhibition

Clerodendron capitatum (Willd) (family: verbenaceae) is locally named as Gung and used traditionally to treat erectile dysfunction. Therefore, the current study was designed to investigate the erectogenic properties of C. capitatum. The relaxation effect of this plant was tested on phenylephrine precontracted rabbit corpus cavernosum smooth muscle (CCSM). The effects of C. capitatum were also examined on isolated Guinea pig atria alone, in the presence of calcium chloride (Ca2+ channel blocker), atropine (cholinergic blocker), and glibenclamide (ATP-sensitive K+ channel blocker). These effects were confirmed on isolated rabbit aortic strips. The extract, when tested colorimetrically for its inhibitory activities on phosphordiesterase-5 (PDE-5) in vitro towards p-nitrophenyl phenyl phosphate (PNPPP), was observed to induce significant dose-dependent inhibition of PDE-5, with an ID50 of 0.161 mg/ml (P < .05). In conclusion, our results suggest that C. capitatum possesses a relaxant effect on CCSM, which is attributable to the inhibition of PDE-5, but not mediated by the release calcium, activation of adrenergic or cholinergic receptors, or the activation of potassium channels.

Three Phases of Corporal Tracing Elicited by Electrical Field Stimulation on Rabbit Corpus Cavernosum Smooth Muscle in Penile Perfusion Model

Nitric oxide (NO) has been shown to mediate electrical field stimulation (EFS)-caused smooth muscle relaxation. It is known that the neural control of penile erection involves adrenergic, cholinergic, and non-adrenergic-non-cholinergic (NANC) neuro-effector systems; however, the effects of EFS on adrenergic and cholinergic nerves are not clear. To elucidate EFS-induced signal transductions involved in adrenergic, cholinergic, and NANC neuro-effector systems by using an in vitro penile perfusion model. EFS was performed on penile corpus cavernosum smooth muscle from male New Zealand White rabbits, which was pre-contracted with L-phenylephrine (10 µM). We investigated the penile tracing elicited by EFS on tissues pre-incubated with guanethidine (Guan, 50 µM), tetrodotoxin (TTX, 10 µM), Nω nitro-L-arginine-methyl ester (L-NAME, 1 mM), atropine (50 µM), or eserine (10 µM). The time-to-peak of each phase, the percentage of relaxation, and the area under the curve (AUC). We discovered an extraordinary phenomenon: three distinct phases elicited by EFS. Phase I was abolished by L-NAME. Phase II was decreased by eserine and Guan, but increased by L-NAME. Phase III was abolished by atropine, but enhanced by eserine and Guan. TTX diminished all three phases. The time to reach the top of phase I was delayed by TTX. The time to attain the peak of phase II was shortened by L-NAME, but delayed by TTX and atropine. The time to reach the top of phase III was shortened by L-NAME, eserine, and Guan. AUC was significantly decreased by L-NAME and TTX. EFS stimulated adrenergic, cholinergic, and NANC neuro-effector systems simultaneously. Phase I was related to the NO pathway. Phase II was multiply affected by self-recovery properties, and adrenergic and cholinergic nerves. Phase III was related to cholinergic nerves. The corporal tracing elicited by EFS was the balanced result of multiple factors.

Changes in the neurogenic and endothelium-dependent relaxant responses of rabbit corpus cavernosum smooth muscle after cavernous nerve neurotomy

The present study was conducted to investigate the reactivity of the corpus cavernosum smooth muscle after unilateral cavernous nerve neurotomy in rabbits. Rabbits (18) were randomly divided into two groups: sham-operated (n = 9) and those subjected to unilateral neurotomy of a 5-mm segment of the cavernous nerve (n = 9). The reactivity of the corpus cavernosum tissue from the neurotomized and sham groups was studied in organ chambers at 4 weeks postoperation. In the neurotomized group, endothelium-dependent relaxation of the corpus cavernosum smooth muscle to carbachol was significantly increased when compared to the sham group. In addition, the sensitivity (i.e., pD(2)) of neurotomized strips to carbachol was also increased when compared to controls. Electrical field stimulation-induced neurogenic relaxation was significantly reduced in the neurotomized group. Relaxation to the nitric oxide (NO) donor sodium nitroprusside and to papaverine was similar in the cavernosal tissue of both groups. There was no change in agonist potency. Furthermore, neurotomy had no effect on KCl-induced contractile responses. When tissue contraction was induced with phenylephrine to study relaxation to various stimuli, the tension induced was similar in the neurotomized and the sham control groups. We conclude that unilateral, chronic cavernous nerve neurotomy causes significant functional changes to the penile erectile tissue of rabbits, which may contribute to the development of impotence.

Chuanxiongzine relaxes isolated corpus cavernosum strips and raises intracavernous pressure in rabbits

It has been shown that there are many Chinese traditional herbals that can enhance sexual activity. Chuanxiongzine is a vasoactive ingredient that has been isolated and purified from Ligusticum chuanxiong Hort. In previous studies, it has been found that chuanxiongzine was effective in relaxing rabbit corpus cavernosum smooth muscle. We determined the effects of chuanxiongzine on relaxation of isolated corpus cavernosum strips in vitro and on increase of intracavernous pressure (ICP) in vivo in rabbits. Chuanxiongzine caused a concentration-dependent relaxation of phenylephrine precontracted isolated corpus cavernosum strips (EC50 1.58 × 10−4 mol l−1), which were endothelium independent and NO independent. However, the guanylyl cyclase inhibitor 1-H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one significantly shifted the chuanxiongzine concentration–response relationship to the right. Although there was no significant difference in the level of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) in isolated corpus cavernosum strips treated with chuanxiongzine or vehicle, chuanxiongzine caused a significant rise in the level of cGMP and cAMP in isolated corpus cavernosum strips pretreated with the activator of adenylyl cyclase forskolin and the source of NO sodium nitroprusside. In an in vivo study, chuanxiongzine dose-dependently raised ICP after the intracavernous injection of its cumulative doses (0.5, 1, 2 and 5 mg kg−1). The ICP increased from baseline to 19.1±3.7, 24.8±2.1, 30.2±4.8 and 39.7±6.1 mm Hg, respectively, and the duration of tumescence ranged from 8.5±2.8 to 22.9±7.3 min. Our results show that chuanxiongzine can relax isolated corpus cavernosum strips of rabbits in vitro and increase ICP of rabbits in vivo, which is neither endothelium dependent nor NO dependent, but may be partly mediated by the inhibition of cAMP phosphodiesterase or cGMP phosphodiesterase.

Expression and function of ryanodine receptors in rabbit penile corpus cavernosum smooth muscle cells

This study aimed to investigate the expression and function of ryanodine receptors (RyRs) in rabbit penile corpus cavernosum smooth muscle (CCSM) cells. New Zealand white rabbit CCSM cells were cultured by primary tissue culture and identified by immunofluorescence technique. mRNA of three RyRs subunits in cultured CCSM cells was detected by reverse transcription polymerase chain reaction (RT-PCR). After excitation by noradrenaline, the concentration of Ca(2+) in CCSM cells was detected by laser scanning confocal microscopy. It was found that only the RyRs1 subunit is expressed in CCSM by RT-PCR. The CCSM cells were divided into two groups: Group A, CCSM cells + 10 micromol l(-1) noradrenaline and Group B, CCSM cells + 50 micromol l(-1) procaine 10 micromol l(-1) noradrenaline. Compared with the base level and the level after noradrenaline excitation, fluorescence intensity improved 44.10 +/- 6.01% in the test group A (n = 8) and 32.92 +/- 4.92% (n = 8) in the procaine control group B, respectively. There were statistically significant differences between group A and group B (P < 0.01). It is concluded that RyRs1 subunit is expressed in CCSM cells and may contribute to regulating the cytoplasmic Ca(2+) level.

Spontaneous Ca2+ Waves in Rabbit Corpus Cavernosum: Modulation by Nitric Oxide and cGMP

Detumescent tone and subsequent relaxation by nitric oxide (NO) are essential processes that determine the erectile state of the penis. Despite this, the mechanisms involved are incompletely understood. It is often assumed that the tone is associated with a sustained high cytosolic Ca(2+) level in the corpus cavernosum smooth muscle cells, however, an alternative possibility is that oscillatory Ca(2+) signals regulate tone, and erection occurs as a result of inhibition of Ca(2+) oscillations by NO. The aim of this study is to determine if smooth muscle cells displayed spontaneous Ca(2+) oscillations and, if so, whether these were regulated by NO. Male New Zealand white rabbits were euthanized and smooth muscle cells were isolated by enzymatic dispersal for confocal imaging of intracellular Ca(2+) (using fluo-4AM) and patch clamp recording of spontaneous membrane currents. Thin tissue slices were also loaded with fluo-4AM for live imaging of Ca(2+). Cytosolic Ca(2+) was measured in isolated smooth muscle cells and tissue slices. Results. Isolated rabbit corpus cavernosum smooth muscle cells developed spontaneous Ca(2+) waves that spread at a mean velocity of 65 microm/s. Dual voltage clamp/confocal recordings revealed that each of the Ca(2+) waves was associated with an inward current typical of the Ca(2+)-activated Cl(-) currents developed by these cells. The waves depended on an intact sarcoplasmic reticulum Ca(2+) store, as they were blocked by cyclopiazonic acid (Calbiochem, San Diego, CA, USA) and agents that interfere with ryanodine receptors and IP(3)-mediated Ca(2+) release. The waves were also inhibited by an NO donor (diethylamine NO; Tocris Bioscience, Bristol, Avon, UK), 3-(5-hydroxymethyl-2-furyl)-1-benzyl indazole (YC-1) (Alexis Biochemicals, Bingham, Notts, UK), 8-bromo-cyclic guanosine mono-phosphate (Tocris), and sildenafil (Viagra, Pfizer, Sandwich, Kent, UK). Regular Ca(2+) oscillations were also observed in whole tissue slices where they were clearly seen to precede contraction. This activity was also markedly inhibited by sildenafil, suggesting that it was under NO regulation. These results provide a new basis for understanding detumescent tone in the corpus cavernosum and its inhibition by NO.

Sildenafil Inhibits Superoxide Formation and Prevents Endothelial Dysfunction in a Mouse Model of Secondhand Smoke Induced Erectile Dysfunction

We determined the effect of passive secondhand cigarette smoke on 1) erectile function in vivo, 2) molecular mechanisms involved in penile vascular function, and 3) erectile function and penile molecular signaling in the presence of phosphodiesterase type 5 inhibitor therapy. Four groups of mice were used, including group 1--controls, group 2--mice exposed to 3 weeks of secondhand smoke (5 hours per day for 5 days per week), group 3--control plus sildenafil (100 mg/kg per day) and group 4--smoke exposed plus sildenafil (100 mg/kg per day). Cavernous nerve electrical stimulation and intracavernous injection of acetylcholine were done to assess erectile function. Constitutive and inducible nitric oxide synthase activity, reactive oxygen species generation, nitrotyrosine formation and superoxide anion levels were assessed. Decreased erectile responses to cavernous nerve electrical stimulation and impaired endothelium dependent erectile responses to ACh in mice exposed to secondhand smoke were observed. Superoxide anion was increased in endothelial and corporeal smooth muscle cells of smoking mouse penises. In mice exposed to secondhand smoke constitutive nitric oxide synthase activity was decreased, and inducible nitric oxide synthase activity, reactive oxygen species generation and nitrotyrosine formation increased. Sildenafil therapy restored constitutive nitric oxide synthase activity in the penis of smoking mice, decreased inducible nitric oxide synthase activity, reactive oxygen species generation and nitrotyrosine formation, and improved erectile responses to cavernous nerve electrical stimulation and acetylcholine. Short-term exposure to secondhand smoke impairs erectile function through excessive penile reactive oxygen species signaling and inducible nitric oxide synthase activity. Decreased penile constitutive nitric oxide synthase activity may be attributable to the decreased endothelial nitric oxide synthase activity resulting from increased oxidative stress. Sildenafil therapy restored nitric oxide synthase activity and decreased reactive oxygen species signaling, resulting in improved erectile function.