Correspondence: Akira Furuta M.D., Ph.D., Department of Urology, Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo 105-8461, Japan. Email: email@example.com
Urethral closure mechanisms under stress conditions consist of passive urethral closure involving connective tissues, fascia and/or ligaments in the pelvis and active urethral closure mediated by hypogastric, pelvic and pudendal nerves. Furthermore, we have previously reported that the active urethral closure mechanism might be divided into two categories: (i) the central nervous control passing onto Onuf's nucleus under sneezing or coughing; and (ii) the bladder-to-urethral spinal reflex under Valsalva-like stress conditions, such as laughing, exercise or lifting heavy objects. There are over 200 million people worldwide with urinary incontinence, a condition that is associated with a significant social impact and reduced quality of life. Therefore, basic research for urinary continence mechanisms in response to different stress conditions can play an essential role in developing treatments for stress urinary incontinence. It has been clinically shown that the etiology of stress urinary incontinence is divided into urethral hypermobility and intrinsic sphincter deficiency, which could respectively correspond to passive and active urethral closure dysfunction. In this review, we summarize the representative stress urinary incontinence animal models and the methods to measure leak point pressures under stress conditions, and then highlight stress-induced urinary continence mechanisms mediated by active urethral closure mechanisms, as well as future pharmacological treatments of stress urinary incontinence. In addition, we introduce our previous reports including sex differences in urethral closure mechanisms under stress conditions and urethral compensatory mechanisms to maintain urinary continence after pudendal nerve injury in female rats.
Over 200 million people worldwide have urinary incontinence. It has been reported that the prevalence of urinary incontinence in women increases during young adult life (20–30%), reaches a near peak around middle age (30–40%) and steadily increases in the elderly (30–50%). In all groups, SUI is the most common (49%), followed by mixed incontinence (29%) and urgency urinary incontinence (21%). Risk factors for female SUI include parity, aging and obesity. In particular, the damage of pelvic floor muscles, ligaments and pudendal nerves induced by childbirth is the most prominent as a pathophysiological basis of SUI. Approximately 30% of mothers become urinary incontinent after their first vaginal delivery. In addition, the prevalence of SUI in women (5.9%) was higher than in men (0.49%) in 2008, although there are few data of sex differences in urethral closure mechanisms under stress conditions in humans.
Urethral muscles are controlled by three sets of peripheral nerves: sacral parasympathetic nerves (pelvic nerves) and thoracolumbar sympathetic nerves (hypogastric nerves) innervating the urethral smooth muscles, and sacral somatic nerves (pudendal nerves) innervating the urethral striated muscles including the external urethral sphincter and pelvic floor muscles. Urethral closure mechanisms under different stress conditions and their pathophysiological changes that contribute to the emergence of SUI have been summarized in a recent thorough review by Yoshimura and Miyazato. Thus, in the present article, we will review and update the information on the representative SUI animal models and the methods to measure leak point pressures under stress conditions, and then highlight stress-induced urinary continence mechanisms mediated by active urethral closure mechanisms, as well as future pharmacological treatments of SUI, using an online literature search for the past 20-year period. In addition, we introduce our previous reports including sex differences in urethral closure mechanisms under stress conditions and urethral compensatory mechanisms to maintain urinary continence after pudendal nerve injury in female rats.[10, 11]
Urethral closure mechanisms under stress conditions in SUI animal models
Two crucial urethral closure mechanisms under stress conditions have been shown using SUI animal models (Table 1). It is well known that passive urethral closure consists of urethral compression through connective tissues, fascia and/or ligaments without nerve activities. In contrast, nerve-mediated active urethral closure that maintains urinary continence during elevation of abdominal pressure can be divided into the two categories: (i) the central nervous control passing onto Onuf's nucleus under sneezing or coughing; and (ii) the bladder-to-urethral spinal reflex under Valsalva-like stress conditions, such as laughing, exercise or lifting heavy objects. It has been reported that the bladder-to-urethral spinal reflex is mediated by hypogastric (sympathetic), pelvic (parasympathetic) and pudendal (somatic) nerves. Therefore, the bladder-to-sympathetic reflex not only achieved passively, but also triggered by bladder afferents (distension), is included in the bladder-to-urethral spinal reflex.
Mechanical urethral compression induced by supportive ligaments and fascia attached to the pelvic floor muscles
Active urethral closure mechanisms
Central nervous control passing onto Onuf's nucleus under sneezing or coughing mediated by pudendal nerves
Bladder-to-urethral spinal reflex under Valsalva-like stress conditions mediated by hypogastric, pelvic and pudendal nerves
Representative SUI animal models are shown in Table 2. SUI animal models with PULT have been reported as models of passive urethral closure dysfunction, because the pubo-urethral ligament plays an important role in closing the urethra under Valsalva-like stress conditions, and increments of posterior urethrovesical angle are known to induce SUI in a clinical setting. However, it is unclear whether passive urethral closure mechanisms in rats are similar to those in humans, because animals usually walk with four legs, whereas humans walk with two legs in a standing position. In contrast, SUI animal models with PNC, PNL and PNT have been reported as models of active urethral closure dysfunction, because pudendal nerve injury during pregnancy or childbirth can induce SUI in women. A SUI animal model induced by VD with a dilated 4 mL balloon in the vagina for 3 h seems to have passive and active urethral closure dysfunction, although the injured condition remains to be clarified. The RB model is also included in the group of “passive and active urethral closure dysfunction”, because this model showed impairment of the sneeze-induced urethral continence reflex, suggesting the possibility that RB might affect the active urethral reflex mechanisms.
Measurements of LPP using Crede, vertical tilt and sneeze methods have also been developed to elucidate stress-induced urethral closure mechanisms. In the Crede method, gentle pressure is slowly applied to the abdominal wall for 10 s until fluid leakage occurred from the urethral orifice, and then immediately removed. However, it seems to be difficult to measure LPP exactly, because intravesical pressure is increased by manual abdominal wall compression. To solve the problem, the vertical tilt method was developed to measure LPP more precisely. In the vertical tilt method, animals are mounted on a tilt table and placed in the vertical position. Intravesical pressure is increased upward in 2-cmH2O steps for 60 s from zero, and LPP are defined as a pressure at which visual identification of a leak from the urethra is observed. In addition, spinal cord transection at the level of Th8-9 is required to eliminate the voiding reflex mediated by spino-bulbo-spinal pathways, so that the passive urethral closure function and bladder-to-urethral spinal reflex can be evaluated using the Crede and vertical tilt methods. In contrast, in the sneeze method, a rat's whisker is cut and inserted into the nostrils to induce sneezing (approximately 30 trials) while examining whether or not urine leakage occurs at the urethral orifice. Pelvic nerve transection is required to eliminate the voiding reflex, because active urethral closure during sneezing is mediated by spinal descending pathways from supraspinal neural centers.
Future pharmacological treatments of SUI
As described earlier, the passive urethral closure consisting of urethral compression through connective tissues, fascia and/or ligaments without nerve activities, the active urethral closure consisting of central nervous controls passing onto Onuf's nucleus during sneezing or coughing and the bladder-to-urethral spinal reflex under Valsalva-like stress conditions are crucial mechanisms for maintaining urinary continence. Clinically, the etiology of SUI is divided into urethral hypermobility and intrinsic sphincter deficiency, which might correspond to passive and active urethral closure dysfunction, respectively. Mid-urethral sling procedures are effective for the treatment of urethral hypermobility (passive urethral closure dysfunction). In contrast, the treatment of intrinsic sphincter deficiency (active urethral closure dysfunction) remains to be established except for some drugs, such as duloxetine and clenbuterol, which are only approved for use in a few countries.[20, 21]
We have previously reported that the central nervous control passing onto Onuf's nucleus during sneezing is mediated by activation of pudendal nerves, and that the bladder-to-urethral spinal reflex under Valsalva-like stress conditions is mediated by activation of hypogastric, pelvic and pudendal nerves (Table 1). The fast contraction of pelvic floor striated muscles mediated by pudendal nerves is important to prevent urine leakage during sneezing or coughing, because the duration of these events is less than 0.15 s. In contrast, the contraction of urethral smooth muscles mediated by pelvic and hypogastric nerves also participates in urinary continence under Valsalva-like stress conditions, such as laughing, exercise or lifting heavy objects, because the duration of these stress conditions are longer than that of sneezing or coughing.
α-AR agonists showed modest improvement for the treatment of SUI by urethral smooth muscle contraction, but these drugs have not been commonly used because of the risk of hypertension and/or hemorrhagic stroke. Thus, pharmacological therapy for Onuf's nucleus controlling pelvic floor striated muscles by pudendal nerves will be effective, although the side-effects induced by duloxetine, a norepinephrine/serotonin reuptake inhibitor, remain to be apprehensive. The representative future drugs, which can act on Onuf's nucleus, are shown in Table 3.[24-29] It has also been documented that hybrid therapy with the combined use of these drugs might be useful to reduce side-effects.
Sex differences in urethral closure mechanisms under stress conditions
Sex differences in urethral function have been well known in rats.[30, 31] In a previous report, the effects of α1-AR antagonists on lower urinary tract obstruction in women are weak compared with men with benign prostatic hyperplasia, because the prostate dominantly innervated by sympathetic nerves is situated along the proximal urethra only in men. Therefore, we investigated sex differences in the urethral continence reflex during bladder compression and neurotransmitters involved in the maintenance of urinary continence reflexes using the Crede-LPP method. In the experiment, we directly manually compressed the bladder wall, because lower abdominal wall compression might stimulate abdominal wall muscles and/or the colon to induce spinal reflexes other than the bladder-to-urethral reflex. Changes in LPP with or without C6 application were investigated to determine the autonomic nerve-sensitive components in LPP in response to bladder compression. It is assumed that the administration of C6 can block ganglionic transmission of the sympathetic and parasympathetic efferent pathways contained in the hypogastric and pelvic nerves, including nerves from sympathetic chain ganglia. According to the results of C6 administration, in which LPP is either decreased or increased, we tested the effects of an α1-AR antagonist and a MR antagonist, or a β-AR antagonist and a NOS inhibitor, respectively, to identify the excitatory or inhibitory transmitters involved in autonomic nerve-mediated urethral responses.
In male rats, the Hg-Hg reflex and the Pel-Pel reflex, which were evaluated after the transection of pelvic and hypogastric nerves, respectively, in the pudendal nerve-transected condition, are mediated by α1-AR and MR, respectively, to induce urethral contraction (Fig. 1a,c). The pelvic-to-pelvic and pudendal nerve (Pel–Pel/Pud) reflex, which was evaluated after the hypogastric nerve transection, activates both α1-ARs and MR because pudendal nerves also receive postganglionic axons from the caudal sympathetic chain ganglia (Fig. 1d). On the other hand, the hypogastric-to-hypogastric and pudendal nerve (Hg-Hg/Pud) reflex, which was evaluated after the pelvic nerve transection, activates only α1-ARs in the hypogastric nerve, suggesting that the activation of hypogastric nerve cannot influence the activation of sympathetic nerves originating from the caudal sympathetic chain ganglia (Fig. 1b). In addition, the LPP in Pel-Pel/Pud reflex was significantly higher than that in the Pel-Pel reflex before C6 application, although there was no significant difference between the LPP in the Hg-Hg reflex and Hg-Hg/Pud reflex before C6 application in male rats. These findings suggest that somatic contribution for LPP through bladder compression will be induced by the Pel-to-Pud reflex, although somatic contribution (urethral striated muscle) for urethral pressure in both the bladder resting phase and contraction phase is known to be quite significant.[33, 34] Furthermore, the role of MR to maintain urinary continence remains to be clarified in men, although urethral contraction mediated by MR activation is induced in the Pel-Pel reflex in male rats (Fig. 2).
In contrast, in female rats, the Hg-Hg, Hg-Hg/Pud and Pel-Pel reflexes were mediated by NO, and induced the relaxing response of the urethra (Fig. 3a–c). Our finding that the Hg-Hg reflex is mediated by release of NO in female rats, however, opposes our understanding that NO is released from parasympathetic nerves carried by the pelvic nerve. It is known that sympathetic preganglionic or postganglionic fibers in hypogastric nerves have facilitatory and inhibitory effects on cholinergic ganglionic transmission in the pelvic ganglia. Therefore, postganglionic parasympathetic nerves that contain NOS can be stimulated by activation of hypogastric nerves; therefore, the Hg-Hg reflex can induce the release of NO in the urethra during abdominal compression. In addition, in the Pel-Pel/Pud reflex, the α1-AR-mediated urethral contractile response is more effective than the NO-mediated urethral relaxing response in female rats (Fig. 3d). Overall, on the basis of these findings, it is assumed that the Hg-Hg and Pel-Pel reflexes during bladder compression become dominant to induce the urethral relaxing response mediated by release of NO when pudendal nerves are injured during pregnancy or childbirth, thereby leading to the higher prevalence of SUI in women than in men (Fig. 4). Furthermore, the LPP in the Pel-Pel/Pud reflex was significantly higher than that in Pel-Pel reflex before C6 application, although there was no significant difference between the LPP in the Hg-Hg reflex and Hg-Hg/Pud reflex before C6 application in female rats, which is similar to the findings in male rats.
Urethral compensatory mechanisms to maintain urinary continence after pudendal nerve injury in female rats
SUI after childbirth is common, as approximately 30% of mothers become urinary incontinent after their first vaginal delivery; however, it is usually short-lasting and spontaneously disappears in most women. In contrast, damage of the pelvic floor nerves and musculature induced by pregnancy or parturition seems to continue for a lifetime in women. For example, a prolonged pudendal motor terminal latency to the external anal sphincter reportedly occurs in women 2 days postpartum, and this prolonged latency was present when the women were re-investigated after 5 years, despite the recovery from SUI 2 months postpartum. In another report, women with SUI show a time-dependent decline in maximum vaginal surface electromyogram activity, which is not seen in healthy women of the same age group with comparable parity.[37, 38] Thus, to clarify urethral compensatory mechanisms for the maintenance of urinary continence after childbirth, acute PNT and PNT-4w female SUI rats with bilateral pudendal nerve transection were examined using the Crede-LPP method. In addition, changes in LPP after C6 treatment, which blocks ganglionic transmission of sympathetic and parasympathetic efferent pathways in hypogastric and pelvic nerves, respectively, were also investigated.
The LPP in PNT-4w female rats were equal to those in naive rats, but significantly higher than those in acute PNT rats; however, the post-C6 LPP in PNT-4w rats were significantly lower when compared with those in naive rats and in acute PNT rats ( Fig. 5). These findings lead to the hypothesis that urethral smooth muscle activity might be enhanced by switching from the NO-mediated relaxation to the α1-AR and MR-mediated contractile responses 4 weeks after pudendal nerve injury to compensate for the deficient striated muscle function (Fig. 6). Therefore, further studies will be required to clarify the effect of α1-AR antagonists on the urethral smooth muscle activity in PNT-4w rats.
Because LPP in naive rats were not altered by C6 application in this study, NO-mediated urethral relaxation seen in acute PNT rats seems to be negated by activation of the pudendal nerves in the normal condition. As the pudendal nerves contain not only somatic nerves, but also postganglionic axons from caudal sympathetic chain ganglia, urethral smooth muscle contraction induced by sympathetic nerve activation in the pudendal nerves might offset NO-mediated urethral smooth muscle relaxation during bladder compression, resulting in no apparent effects of C6 on LPP in naive rats. However, further studies are required to clarify this point.
In the present review, we summarized urethral closure mechanisms under different stress conditions using the representative SUI animal models and the measurement methods of LPP to develop future pharmacological treatments of SUI. In addition, we described our previous reports including sex differences in urethral closure mechanisms under stress conditions and urethral compensatory mechanisms to maintain urinary continence after pudendal nerve injury in female rats. Thus, it seems reasonable to conclude that pathophysiology of SUI could be shown in murine models, and that new modalities of SUI treatment based on the different pathophysiology might also be shown in murine models in future.
Parts of authors’ research have been supported by National Institutes of Health grants (DK067226, AR049398 and DK055387) and Grant-in-Aid for Scientific Research (C) from Japan Society for the Promotion of Science (22591802).