Tohoku J. Exp. Med., 2009, 217, 239-242
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Stress and Erectile Dysfunction
Restraint Stress Impairs Erectile Responses in Rats ERSIN BAL,1 NERGIS MURAT,2 OMER DEMIR,3 BURAK CEM SONER,4 ERTAN CAN,1 SEDEF GIDENER4 and ADIL ESEN3 1
Urology Department, Tepecik Training Hospital, Izmir, Turkey Advanced Professional School of Health Sciences, Dokuz Eylül University, Izmir, Turkey 3 Department of Urology, Medical Faculty, Dokuz Eylül University, Izmir, Turkey 4 Department of Pharmacology, Medical Faculty, Dokuz Eylül University, Izmir, Turkey 2
It has been established that various forms of physical and psychological stress reduce sexual functions. However, there is no study yet evaluating the functional changes over cavernosal pressure in rats exposed to restraint stress. In this study, we aimed to investigate the convenience of the restraint stress model that may be used to determine the disruptive effects of stress on erectile function. Sprague Dawley rats were randomized into two groups as control (n = 7) and stress (n = 7) groups. In the stress group, rats were placed for 60 minutes in a cylindrical plastic tube with holes for fresh air supply (restraint stress). Following the stress application, several parameters for erectile responses were evaluated immediately. The control animals were maintained at room temperature without any procedure until the measurement. During the electrical stimulation of cavernous nerve, we measured the intracavernous pressure (ICP), the ratio of ICP to the mean arterial pressure (MAP), and detumescence time. There were significant decreases in ICP (24.4 ± 4.1 vs 53.4 ± 4.5 mmHg, p < 0.01), ICP/MAP (34.4 ± 7.8% vs 55.7 ± 3.9%, p < 0.05), and detumescence time (31.7 ± 6.1 vs 78.6 ± 12.8 sec, p < 0.01) in stress group when compared to control group. Thus, restraint stress declined detumescence time and decreased intracavernosal pressure in male rats. In conclusion, restraint stress model in rats may be useful for determining the effects of stress on erectile response. Even a short-term restraint stress may cause erectile dysfunction. ──── Erectile Dysfunction; Restraint Stress; Intracavernosal Pressure; Rat; In vivo. Tohoku J. Exp. Med., 2009, 217 (3), 239-242. © 2009 Tohoku University Medical Press
Erectile dysfunction (ED) is a worldwide clinical problem with tens of thousands of new cases in each year. The risk of ED increases with age, comorbid diseases, smoking and sedantery life style (Feldman et al. 1994). The literature review revealed that lots of studies were performed to clarify the underlying pathophysiology for development of ED (Sivalingham et al. 2006). Several experimental and animal models were also used to investigate the association between risk factors and penile erection. Penile erection is a neurovascular phenomenon that involves neurologically mediated increase in penile arterial inflow, relaxation of cavernosal smooth muscle and restriction of venous outflow from the penis (Wagner and Saenz De Tejada 1998; Bivalacqua et al. 2003). Various diseases deteriorate erectile function via altering the nervous, vascular or hormonal systems that may produce changes in the smooth muscle tissue of the corpus cavernosum and influence the patient’s psychological mood and behaviour. Furthermore, multiple forms of stress are believed to reduce sexual functions. The relationship between stress and sexual behavior in both male and female rats using immobilization, restraint
models, has been shown in some studies (Sato et al. 1996; Yoon et al. 2005). The link of chronic stress to sexual function disorders was demonstrated in experimental behavior studies (Almeida et al. 2000). The association between psychogenic ED and stress, depression and anxiety has also been shown (Bodie et al. 2003). However, there has been no study evaluating the functional changes over cavernosal pressure in rats exposed to stress. Also, there has been no animal model investigating the functional effects of stress on cavernosal structures. Thus, the aim of this study was to investigate the convenience of the restraint stress model for evaluating the effects of stress on erectile function in rats. We also aimed to investigate whether stress induces changes on the erectile response of the penis.
MATERIAL AND METHOD Experimental animals and stress conditioning This study was approved by Dokuz Eylul University Medical Faculty Experimental Animals’ Ethic Committee. Sprague Dawley rats, weighing 300 - 400 g, were kept under standardized laboratory conditions (temperature 23°C, 12h light-dark cycle) with free access
Received October 21, 2008; revision accepted for publication February 17, 2009. Correspomdence: Omer Demir, Department of Urology, Medical Faculty, Dokuz Eylül University, Izmir, Turkey. e-mail:
[email protected]
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to food and water. Animals were kept in cages measuring 42 cm length × 26 cm width × 18 cm height with four rats per cage. Rats were randomized into two groups as control group (n = 7) and stress group (n = 7). The restraint stress applied to the stress group consisted of immobilization. Rats were subjected to restraint stress for 60 minutes on scheduled day (McDougall et al. 2000). Animals were placed in a cylindrical polyvinyl chloride tube (6 cm diameter) with holes for fresh air supply. The length of the tubes was adjusted that the animals were unable to move/turn around. The restraint stress was applied to rats at the same time of the day. Following the stress application, in vivo measurements were done immediately. The control group was maintained at room temperature with free access to food and water. Animals in control group did not undergo any procedure until the experiment. In vivo measurements of the control animals were also done at the same time of the day, with the stress group on the scheduled day. Measurement of erectile responses The rats from each group were anesthetized with urethane and alpha-chloralose (450 and 100 mg/kg intraperitoneally, respectively) and placed on a surgical table. Trachea was cannulated (PE 14 polyethylene tubing) to maintain the airway. Right carotid artery was cannulated (PE 50 tubing) for the measurement of systemic arterial pressure. The bladder and prostate was exposed through a midline abdominal incision. The cavernosal nerve was identified posterolateral aspect of the prostate. Bipolar hook (Harward Dastre electrode, USA) was placed around the cavernosal nerve. Electrical stimulation at 20 Hz was performed with pulse duration of 1 msec (ms) for 1 minute at 5 V. These measurements were obtained as a result of a preliminary study designed to determine the optimal stimulatory conditions. The penis was denuded of skin and the tip of a catheter, with a 25-gauge needle at one end, inserted into the corpus cavernosum for measurement of intracavernous pressure (ICP). Both the arterial and cavernous catheters were filled with heparinized saline (250 U/ml). Systemic arterial and cavernosal pressure were recorded continuously with a MP30 BPT300 transducer connected to a computerized system for data acquisition (MP30 Biopac systems Inc., Santa Barbara California, U.S.A). Baseline measurements of ICP and MAP were recorded just before the stimulation of cavernous nerve in both control and stress groups. Then erectile responses to the cavernous nerve stimulation (CNS) were recorded for each animal. Statistical analysis was performed with the following parameters that shows the erectile response to the CNS: 1) maximum ICP (mmHg), defined as the maximum value of the ICP at the point of ICP curve during CNS interval that indicates the maximal relaxation capacity of deep arteries in the penis, 2) the ratio of the ICP to the MAP, defined as the ratio of the ICP to the MAP values at the point of maximal ICP values during CNS interval that indicates the extent of relaxation of corpus cavernosum, while MAP represents the capacity of pumping blood into the penis, 3) area under the curve (AUC), to define the ICP alterations with time during CNS, which gives erection capability, and 4) detumescence time, defined as the period from the end of CNS to a point showing 50% of maximal ICP, which may reflect the bioavailability of cyclic guanosine monophosphate (cGMP). Statistical analysis All data were expressed as mean ± SEM. Statistical analyses were performed by Mann-Whitney U test at a significance level of p
0.05). The maximal ICP values were significantly decreased in stress group (24.4 ± 4.1 mmHg) when compared to controls (53.4 ± 4.5 mmHg) ( p < 0.01, Fig. 1). The AUC showed that stress group had a significantly lower AUC than the control (1061 ± 157.2 vs 2249 ± 253.7, respectively) ( p < 0.01, Fig. 2). The ratio of ICP/MAP of the stress group (34.4 ± 7.8%) was significantly lower than the control group (55.7 ± 3.9%) ( p < 0.05, Fig. 3). Detumescence time was shortened in stress group when compared with control group (31.7 ± 6.1 sec vs 78.6 ± 12.8 sec) ( p < 0.01, Fig. 4).
Fig. 1. Comparison of the erectile function in groups. An electrostimulation of 20 Hz with a pulse duration of 1 ms for 1 minute at 5 V was applied to control (n = 7) and stress group (n = 7) in order to elicit penile erection. The maximum ICP (mmHg) were significantly lower in stress group than in controls. Data are expressed as means ± SEM. Significantly different from the control a: p < 0.01.
Fig. 2. Effects of stress on erectile responses. The data is expressed as means ± SEM of the AUC (mmHg × sec). (a) Significantly different from the control ( p < 0.01).
Stress and Erectile Dysfunction
Fig. 3. Comparison of ICP/MAP (%) of groups. Significantly different from the control a; p < 0.05.
Fig. 4. The effects of stress on erectile responses. The detumescence time was taken to be the period from the termination of the CNS (20 Hz) to the time when the ICP returned to 50% of mean baseline value. Significantly different from the control a: p < 0.01.
DISCUSSION Stress has long been a focus among researchers interested in psychological influences on health. Responses to stress are numerous and include physiological changes, cognitive and emotional reactions as well as behavioral responses (Curtis et al. 2005). Furthermore, various forms of stress models such as restraint or mental are known to cause organic responses. Several studies have examined the relationship between stress and sexual behavior in both male and female rats (Sato et al. 1996; Bodie et al. 2003; Yoon et al. 2005). In these studies it has been reported that sexual dysfunction occurs due to complex changes of serum sex hormones, catecholamines and NOS subtype expressions in the vaginal tissues and at the cerebral area (Sato et al. 1996; Yoon et al. 2005). However, in our literature review we did not find any published study investigating the changes of penile erection and cavernosal tissues due to physical stress. In our study, we hypothesized that restraint stress may cause organic ED besides psychogenic ED. Hence, we evaluated the relationship of restraint stress and ICP chang-
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es due to electrical stimulation in rats. In this study, we demonstrated that restraint stress diminished ICP, ICP/MAP, AUC, and detumescence time in male rats. These results suggest that restraint stress may cause organic ED in male rats. We found a significant decrease in the detumescence time in stressed animals. In another animal model of ED, it has been shown that hypercholesterolemia shortened detumescence time in rats. This shortening of the detumescence time is thought to be caused by the hyperlipidemia related activation of the degradation of cGMP, which provides corpus cavernosum smooth muscle relaxation. cGMP is a second messenger that is mainly activated by nitric oxide (NO) (Ryu et al. 2006). Synthesis of NO and the NO binding to soluble guanyl cyclase are essential for the erectile process, and several cardiovascular risk factors affect the NO/cGMP pathway (Andersson 2001). Although synthesis or activation of NO/cGMP was not evaluated, our findings of decreasing detumescence time in stressed animals are similar to the previous studies, in which the erectile functions were evaluated in diabetic and hypercholesterolemic rats (Kang et al. 2005; Yu et al. 2005). In these studies, the detumescence time was prolonged in diabetic and hypercholesterolemic rats with the use of phosphodiesterase type 5 inhibitors. According to our results, we suggest that decrease in ICP, AUC and ICP/MAP values may be due to reduction in NO synthesis and/or bioavailability. The parameter ICP, ICP/MAP and AUC show the maximal relaxation capacity of penile artery and corpus cavernosum smooth muscle. The maximum relaxation capacity of these erectile structures shows differences according to the amount of NO content. These differences were shown in hypercholesterolemic and diabetic animal model of ED (Gholami et al. 2003; Bivalacqua et al. 2004; Park et al. 2005). In these studies, it has been established that ICP, ICP/MAP and AUC values were significantly decreased in hypercholesterolemic or diabetic animals than those in controls. The decreases in these values were associated with the impairment of NO/cGMP pathway. The limitations of our study should be mentioned. First the association between stress and ED has not been studied in the molecular level. Second the effects of handling stress could not be ignored in control animal; namely, the possibility remains that handling stress alone may impair the erectile responses in rats. Another possible limitation was that plasma level of stress hormones was not measured. Further studies with eliciting the exact mechanisms in stress-related erectile responses in male rats are required. In conclusion, physical stress may cause ED in male rats by decreasing detumescence time and ICP. We suggest that stress primarily causes the cavernosal damage, which in turn impairs erectile functions. Further studies are needed to clarify the mechanism of stress-induced ED at the molecular level and time related changes in erectile responses.
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Acknowledgment This study was supported by a grant from the Turkish Society of Andrology.
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