Urethral sphincter mechanism incompetence is the most common cause of acquired urinary incontinence in spayed female dogs.1 Among dogs, the prevalence of the disease in spayed females is higher than it is in sexually intact females.2–5 Indeed, Thrusfield et al6 determined an incidence rate per animal-year of 1.7% in neutered dogs and 0.22% in sexually intact dogs; the relative risk for development of USMI was 7.8 times greater in neutered dogs. Among female dogs with USMI, 90% are spayed.2,4 Clinical signs of USMI, such as urine leakage during sleep or periods of excitement, may develop as long as 10 years after spaying.1,3,7 Results of urodynamic studies8–10 have indicated that MUCP and UFPL values in female dogs with USMI are significantly decreased, compared with findings in female dogs without urinary incontinence.
Urethral sphincter mechanism incompetence has been compared to postmenopausal urinary incontinence in women because both conditions are associated with a state of estrogen deficiency.7 In women, the lack of estrogens induces major changes in the lower portion of the urinary tract, including bladder and urethral mucosal atrophy, reduction of urethral and periurethral collagen content and urethral submucosal vascularity, and decreased sensitivity of the urethral smooth musculature to α-adrenergic stimulation.11 These postmenopausal structural changes lead to functional urodynamic modifications such as decreases in MUCP12–17 and UFPL14 and increases in bladder and premicturition detrusor pressures.17
Structural and functional changes that develop in the urogenital tract of dogs after spaying have been sparsely studied in veterinary medicine. In dogs, vaginal length and volume are decreased in spayed females, compared with findings in sexually intact females.18,19 Reichler et al20 detected a significant decrease in MUCP in spayed female dogs 10 months after sterilization. One might hypothesize that decreased urethral resistance in spayed dogs could be related to the lack of estrogens and could predispose these animals to the development of USMI. Therefore, estrogen replacement has been widely used in the treatment of stress incontinence in women or USMI in female dogs.2,21–31 The extensive use of estrogens in the treatment of these conditions was reinforced by identification of highaffinity estradiol receptors in the urethra, vagina, and pubococcygeal muscle and, to a lesser extent, in the bladder of humans, rabbits, and rats.32–35 In female dogs, estrogen receptors have been identified in the transitional epithelium and subjacent stroma of the proximal portion of the urethra, but were not detected in the kidneys, ureters, bladder, and distal portion of the urethra36
Estriol, a naturally occurring estrogen, is commonly used in humans and is currently the estrogenic substance of choice for treatment of female dogs. It differs from other estrogen compounds, such as estradiol and stilbestrol, because it occupies the nuclear-bound receptors for a relatively short period,37 thereby decreasing the risk of associated adverse effects. Estriol has been used for decades in hormone replacement therapy for women and has not been associated with bone marrow depletion.38 Long-term administration of estriol in female dogs has not been accompanied by adverse effects such as pyometra and medullar aplasia.39
Unexpectedly, hormone replacement in postmenopausal women has led to controversial and contradictory results.21–25 Whereas some studies24,26–29 have indicated improvement of the symptoms of urinary incontinence associated with urogenital atrophy and recurrent urinary tract infections after oral or intravaginal administration of estriol, other studies25,40–42 did not reveal any improvement of the urinary symptoms after estrogen treatment. Although increased urethral pressure was detected by Dessole et al29 after intravaginal administration of estriol, other investigators detected minimal23 or no changes22,43 in the values of urodynamic parameters with the exception of an increased ratio of pressure transmission to the proximal portion of the urethra.23,24,29,43
In female dogs with USMI, the success rate for complete urinary continence with administration of estriol is approximately 65%.30,31 The urodynamic effects of estriol administration have been sparsely investigated in the veterinary literature. In 1 study,44,a an increased UFPL and threshold bladder pressure were detected following estriol administration in Beagles.
On the basis of evidence that estrogens increase the sensitivity of the α-adrenergic receptors to the sympathetic stimulation,7,45–48 it appears reasonable to combine an α-adrenergic agent with estrogen to try to obtain a synergetic therapeutic effect. Results of several studies40,49–53 have indicated that administration of such drug combinations is superior to the administration of estriol alone, in terms of improving urinary symptoms attributable to stress incontinence in postmenopausal women. However, results are contradictory regarding the urodynamic effects of this type of combination treatment. In some studies,50–52,54 values of various urodynamic parameters increased after the combined administration, compared with effects after administration of estriol alone, whereas another study52 did not reveal any modification in values of the urodynamic parameters, although the symptoms in postmenopausal women with stress urinary incontinence were greatly improved.
Investigations of the effectiveness of estrogen combined with an α-adrenergic agent in female dogs are almost nonexistent. Creed47 identified an increased UFPL after administration of estrogens and phenylephrine in female dogs without urinary incontinence, whereas this effect was not detected in incontinent females. The purpose of the study reported here was to evaluate and compare the urodynamic and morphologic effects of the administration of estriol alone and in combination with phenylpropanolamine on the lower portion of the urogenital tract in sexually intact and spayed female Beagles that did not have urinary incontinence.
Materials and Methods
Dogs—Six adult (mean ± SD age, 11 ± 1 years old) female Beagles were included in the study. The dogs were allocated to 1 of 2 groups on the basis of their reproductive status; group 1 included 3 sexually intact females in anestrus (determined on the basis of visual examination of the vulva and cytologic examination of a vaginal smear55), and group 2 included 3 ovariectomized females. All dogs were born and housed at the Small Animal Theriogenology facilities of the Department of Clinical Sciences of the College of Veterinary Medicine, University of Liège. Animal housing, care, and experimentation were in accordance with Belgian governmental regulations and with the NIH Guide for Care and Use of Laboratory Animals.56 Complete physical examinations were performed on each dog each week. Dogs were housed in groups of 2 to 5 in indoor-outdoor runs (2.5 × 10 m), exposed to natural light, fed a commercial dry food once a day in amounts sufficient to maintain body weight, and provided with water ad libitum. The dogs weighed 12 to 17 kg (mean ± SD weight, 15 ± 0.5 kg). Prior to each experiment, each dog underwent a physical examination, which included measurements of heart rate, respiratory rate, and rectal temperature, and a urine sample was obtained via cystocentesis for urinalysis and bacteriologic culture. Urinalysis included determination of specific gravity and pH and detection of blood, protein, bilirubin, and glucose when present. A vaginal smear was also performed during the course of the treatments to identify potential cytologic changes induced by the estrogenic supplementation.
Study design—Dogs received estriolb (2 mg) PO once a day for 7 days (treatment A), immediately followed by the combined administration PO of estriolb (2 mg) and phenylpropanolamine hydrochloridec in an immediate-release formulation (1.5 mg/kg, PO) once a day for 7 additional days (treatment B). Urodynamic and morphologic measurements were performed in each dog the day before the first estriol treatment (baseline data; day 0) and on days 7 and 14 within 2 hours of drug administration. Each animal was used as its own control, to decrease variations attributable to interindividual differences and increase the statistical power, which is important when experimental groups are composed of small numbers of animals.
Prior to data collection, food was withheld from the dogs for 12 hours; dogs were allowed to urinate prior to each urodynamic measurement. Anesthesia was induced by use of an IV bolus of propofold (5 mg/kg) and was maintained with a continuous IV infusion of propofol at a rate of approximately 30 mg/kg/h. All dogs were intubated and monitored with a pulse oxymetere; oxygen supplementation was provided if oxygen saturation was < 85%. When dogs were in a light plane of anesthesia, they were placed in right lateral recumbency.57 After cystocentesis and collection of a sample for preparation of a vaginal smear, 3 successive simultaneous urethral pressure profiles and diuresis cystometry were performed, as previously described.58 After the urodynamic testing, each dog remained anesthetized in right lateral recumbency and vaginourethrography was performed, as described.59
Data interpretation—By use of the urodynamic test results and examination of vaginourethrograms (lateral views), specific variables were assessed (Figure 1). These included MUP, MUCP (ie, the difference between MUP and bladder pressure), IP (ie, the functional area under the UFP, calculated as [MUCP × UFPL]), UFPL (defined as the distance between the point in the urethra where intraurethral pressure exceeds bladder pressure and the point where either a pressure plateau is detected or where the intraurethral pressure is less than bladder pressure), urethral anatomical profile length (ie, the distance between the point in the urethra where intraurethral pressure exceeds bladder pressure and the point where the intraurethral pressure decreases to atmospheric pressure), plateau (the distance between the end of the UFPL and the point where the intraurethral pressure falls to atmospheric pressure), the distance before MUP (defined as the distance between the point in the urethra where intraurethral pressure exceeds the total bladder pressure and the point of maximal urethral pressure), maximum meatus pressure (defined as the maximal pressure at the level of the external urethral orifice), threshold pressure (ie, urinary bladder pressure at the time of micturition), threshold volume (ie, fluid volume retrieved from the urinary bladder at the time of micturition reflex), compliance, urethral length (ie, distance between the bladder neck and the external urethral orifice), vaginal length, vaginal width, the length of the dorsal median paracervical fold (distance A), the distance between the dorsal median paracervical fold and the vestibulovaginal junction (distance B), and the distance between the vestibulovaginal junction and the external urethral orifice (distance C').
Drawing of the features of interest on a vaginourethrogram (lateral view) in a female dog. The urethral length is defined as the distance between the bladder neck and the external urethral orifice. The vaginal length is defined as the distance between the cervix and the external urethral orifice. Distance A is defined as the length of the dorsal median paracervical fold (a to b); distance B is defined as the distance between point b and the vestibulovaginal junction (c); distance C' is defined as the distance between point c and the external urethral orifice (d). VW = Vaginal width. (Adapted from Hamaide AJ, Verstegen JP, Snaps FR, et al. Influence of the estrus cycle on urodynamic and morphometric measurements of the lower portion of the urogenital tract in dogs. Am J Vet Res 2005;66:1075–1083. Reprinted with permission.)
Citation: American Journal of Veterinary Research 67, 5; 10.2460/ajvr.67.5.901
Drawing of the features of interest on a vaginourethrogram (lateral view) in a female dog. The urethral length is defined as the distance between the bladder neck and the external urethral orifice. The vaginal length is defined as the distance between the cervix and the external urethral orifice. Distance A is defined as the length of the dorsal median paracervical fold (a to b); distance B is defined as the distance between point b and the vestibulovaginal junction (c); distance C' is defined as the distance between point c and the external urethral orifice (d). VW = Vaginal width. (Adapted from Hamaide AJ, Verstegen JP, Snaps FR, et al. Influence of the estrus cycle on urodynamic and morphometric measurements of the lower portion of the urogenital tract in dogs. Am J Vet Res 2005;66:1075–1083. Reprinted with permission.)
Citation: American Journal of Veterinary Research 67, 5; 10.2460/ajvr.67.5.901
Drawing of the features of interest on a vaginourethrogram (lateral view) in a female dog. The urethral length is defined as the distance between the bladder neck and the external urethral orifice. The vaginal length is defined as the distance between the cervix and the external urethral orifice. Distance A is defined as the length of the dorsal median paracervical fold (a to b); distance B is defined as the distance between point b and the vestibulovaginal junction (c); distance C' is defined as the distance between point c and the external urethral orifice (d). VW = Vaginal width. (Adapted from Hamaide AJ, Verstegen JP, Snaps FR, et al. Influence of the estrus cycle on urodynamic and morphometric measurements of the lower portion of the urogenital tract in dogs. Am J Vet Res 2005;66:1075–1083. Reprinted with permission.)
Citation: American Journal of Veterinary Research 67, 5; 10.2460/ajvr.67.5.901
Compliance was calculated by use of the following equation:
where Vo and Po are the urinary bladder volume and pressure at the start of cystometric evaluation, respectively. Definitions of urodynamic measurements were in accordance with those of the International Continence Society.60
Statistical analysis—Data are expressed as mean ± SE. Differences in age and weight between groups were determined by use of a 1-way ANOVA. Two-way ANOVAs with interaction (treatment or group) were performed by use of statistical software.f Within-treatment comparisons (differences between successive times of testing), between-treatments comparisons, and between-groups comparisons were made. A value of P < 0.05 was considered significant.61
Results
Physical examination, urinalysis, and bacteriologic culture of urine revealed no signs of disease of the lower portion of the urinary tract in any of the dogs included in the study. Mean ± SE values of urodynamic and morphologic measurements were calculated (Table 1). At days 7 and 14, cytologic examination of vaginal smears from 2 sexually intact and 2 spayed female dogs had evidence of estrogenic maturation including the presence of RBCs, mitotic figures, and a few keratinized cells. Cytologically, vaginal smears of the remaining sexually intact and spayed females were compatible with smears from female dogs in the proestrous phase.
Mean ± SE values of urodynamic and morphometric variables in 3 sexually intact and 3 spayed female dogs before (day 0) treatment, after treatment with estriol (2 mg, PO, q 24 h) for 7 days (day 7), and after the same treatment with estriol combined with phenylpropanolamine (1.5 mg/kg, PO, q 24 h) for another 7 days (day 14).
| Variable | Day 0 | Day 7 | Day 14 | |||
|---|---|---|---|---|---|---|
| Sexually intact | Spayed | Sexually intact | Spayed | Sexually intact | Spayed | |
| MUP (cm H20) | 23 ± 9 | 30 ± 9 | 65 ± 9* | 46 ± 9 | 43 ± 9 | 46 ± 9 |
| MUCP (cm H20) | 19 ± 8 | 27 ± 8 | 60 ± 8* | 38 ± 8 | 37 ± 8 | 41 ± 8 |
| Integrated pressure (cm × cm H20) | 100 ± 39 | 145 ± 39 | 255 ± 39* | 234 ± 39 | 197 ± 39 | 214 ± 39 |
| UFPL (mm) | 79 ± 6 | 82 ± 6 | 86 ± 6 | 89 ± 6 | 87 ± 6 | 87 ± 6 |
| UAPL (mm) | 104 ± 6 | 100 ± 6 | 98 ± 6 | 100 ± 6 | 107 ± 6 | 93 ± 6 |
| BMUP (mm) | 52 ± 10 | 47 ± 10 | 62 ± 10 | 63 ± 10 | 63 ± 10 | 66 ± 10 |
| MMP (cm H20) | 23 ± 12 | 14 ± 12 | 23 ± 12 | 25 ± 12 | 13 ± 12 | 20 ± 12 |
| Plateau (mm) | 15 ± 3 | 8 ± 3 | 12 ± 3 | 7 ± 3 | 14 ± 3 | 9 ± 3 |
| Threshold pressure (cm H20) | 49 ± 7 | 33 ± 7 | 46 ± 7 | 50 ± 7 | 46 ± 7 | 48 ± 7 |
| Threshold volume (mL) | 401 ± 81 | 407 ± 81 | 406 ± 81 | 452 ± 81 | 457 ± 81 | 422 ± 81 |
| Compliance (mL/cm H20) | 10 ± 2 | 16 ± 2 | 10 ± 2 | 11 ± 2 | 11 ± 2 | 10 ± 2 |
| Urethral length (cm) | 11 ± 2 | 10 ± 1 | 10 ± 1 | 11 ± 1 | 11 ± 2 | 11 ± 2 |
| Distance A (cm) | 1 ± 0.2† | 0.8 ± 0.3 | 1.2 ± 0.2 | 0.5 ± 0.3 | 1.3 ± 0.2 | 0.5 0.3 |
| Distance B (cm) | 12 ± 1† | 7 ± 2 | 12 ± 1 | 9 ± 2 | 12 ± 1 | 8 ± 2 |
| Distance C′ (cm) | 1 ± 0.3 | 1 ± 0.4 | 2 ± 0.3 | 1.2 ± 0.4 | 1.3 ± 0.3 | 1.2 0.4 |
| Vaginal length (cm) | 15 ± 2† | 8 ± 2 | 16 ± 2 | 11 ± 2 | 15 ± 2 | 10 ± 2 |
| Vaginal width (cm) | 4 ± 0.5† | 1 ± 0.6 | 3 ± 0.5 | 1.5 ± 0.7 | 3 ± 0.5 | 1.8 ± 0.6 |
Value significantly (P < 0.05) different from baseline value (day 0) for that group.
Value significantly (P < 0.05) different from value for spayed females at day 0.
UAPL = Urethral anatomic profile length. BMUP = Distance between the point in the urethra where intrau-rethral pressure exceeds the total bladder pressure and the point of MUP. MMP = Maximum meatus pressure.
Urodynamic measurements—At day 0, no significant differences in the urodynamic variables were detected between sexually intact and spayed female dogs. Compared with findings at day 0, MUP, MUCP, and integrated pressure values were increased from baseline in both groups of female dogs after treatment A; however, these differences from baseline values were significant (P = 0.004, P = 0.005, and P = 0.006, respectively) only in sexually intact females. Compared with values obtained after treatment A, treatment B did not induce significant differences in MUP, MUCP, and IP values.
Radiographic measurements of the urethra and vagina—At day 0, vaginal length and width, as well as distances A and B, were significantly (P = 0.008, P = 0.006, P = 0.01, and P = 0.004, respectively) shorter in spayed female dogs, compared with values in sexually intact female dogs. After treatment A (day 7), no significant differences in the values of the radiographic measurements were identified between the 2 groups. Similarly, after treatment B (day 14), no significant differences in the values of the radiographic measurements were identified between the 2 groups. At days 7 and 14, radiographic measurements did not differ from baseline values in either group.
Discussion
The first purpose of the present study was to evaluate the effects of administration of estriol on the function of the lower portion of the urogenital tract in sexually intact and spayed female dogs that did not have urinary incontinence. The dosage of estriol used was chosen on the basis of previous reports30,31 in which a starting dosage of 0.5 to 2 mg/dog, PO, once daily for 7 days was recommended. Because estriol undergoes metabolism via the enterohepatic cycle, important variations in plasma concentrations are detected among dogs, which may explain the lack of relationship between the recommended dosage and the weight of the dog.30,31
Although estriol supplementation has been associated with improvement or complete cure of clinical signs in women and female dogs with urethral sphincter deficiency,24,26–31 treatment-associated urodynamic changes in humans have been variable.23,29 In the present study, a significant increase in urethral resistance in female dogs was detected after 7 days of treatment with estriol, as indicated by significant increases in MUP, MUCP, and integrated pressure values (compared with pretreatment baseline values). An integrated pressure value defines the functional area under the urethral profile curve, which is a better measure of urethral sphincter competence than MUCP or UFPL alone because it more fully characterizes sphincter deficiency in humans and is a useful parameter in assessing urethral function.62 Although MUP, MUCP, and integrated pressure values were increased after estriol treatment in both groups of dogs in our study, these values were significantly increased from baseline only in the sexually intact females. We believe that the lack of significance associated with the increases in the spayed group could be related to the small number of dogs or to a reduced capacity of spayed animals to respond to the treatment. It must be also acknowledged that the plane of anesthesia was not standardized among dogs in terms of identical infusion rates. The goal was to achieve a similar light plane of anesthesia in each dog with persistence of eye blinking and jaw tone. Although good reproducibility of urodynamic measurements in individual female dogs by use of propofol has been described,58 interindividual variations attributable to the systemic effects of propofol could have influenced the urodynamic results.
In a previous study63 of the influence of the endogenous hormones on the urinary tract in dogs, slight increases in MUP and MUCP values during proestrus, compared with values during anestrus, were detected, which could be explained by the effects of the increased estrogen plasma concentration associated with the proestrous phase on the lower portion of the urinary tract.
In the present study, the increased urethral resistance detected after estrogen administration could be explained by several mechanisms. In humans, dogs, and rabbits, estrogens stimulate α-adrenergic receptors of the urethra and increase their sensitivity to sympathetic stimulation,7,45–48,64,65 leading to increased smooth muscle contractility.66 In the urethra of rabbits, Larsson et al67 detected an increase in the number of α2-adrenergic receptors after estrogen supplementation, compared with the pretreatment value. In women, it is suggested that estrogens stimulate collagen production and increase collagen density in the urethral wall and supporting structures of the urethra.68,69 Blood flow in the sinusoidal plexus, which plays an important role in urethral function,70,71 is enhanced after estrogen administration.72,73 Estrogen replacement in women also induces maturation of the vaginal and urethral epithelium74 with an associated increased thickness of the urethral wall, which potentially contributes to a better seal of the urethral lumen. In the present study, maturation changes were detected via cytologic examination of vaginal smears from all dogs during the 14 days of treatment, suggesting that the observed increased urethral resistance could be related to a thickening of the wall of the vagina and subsequently that of the urethra. In 1 sexually intact female dog, the findings indicative of proestrus after estriol supplementation could have been an early return to estrus. In female dogs, sterilization induces a decrease in smooth muscle mass and urethral connective tissue density,75 which could be associated with the decrease in MUCP values detected after spaying.20 Estrogen supplementation could therefore be beneficial in reversing these structural connective changes and enhance urethral resistance. However, in rats, administration of 17β-estradiol does not modify the volume of the smooth musculature nor the innervation to the proximal portion of the urethra.76
Interestingly, the significant increase in functional area detected after estriol supplementation was mainly a result of the modification of the urethral pressure profile curve in its distal portion, corresponding to the distal fourth of the urethra, which is rich in striated musculature. After ovariectomy in dogs, density of type I fibers from the striated urethral musculature decreases to a greater extent than type II fibers,77 which could play a role in the weakness of the sphincter mechanism because type I fibers are supposed to be involved in the maintenance of urinary continence at rest. The administration of estrogens in adult female cats has induced motoneuron axonal growth, suggesting that sex steroid hormones can influence motoneurons78 and possibly influence the number of muscular fibers, via their innervation. This urodynamic observation suggests that estrogen action on the urethra could take place in all components of the urethral wall, including not only the smooth musculature, vascular plexus, connective tissue, and urothelium but also the striated musculature.
The values of the cystometric measurements were not significantly different after the administration of estriol (treatment A) in either the sexually intact or spayed female dogs. However, spayed dogs responded to some extent to the estriol supplementation, whereas values from the sexually intact dogs did not vary from baseline after treatment A. Although not significant, threshold pressure was increased and compliance values decreased from baseline after treatment A in the spayed dogs, whereas no difference was detected in the sexually intact dogs. These results could corroborate previous findings,44,a but once again, investigation of a larger number of dogs would be necessary to draw definite conclusions.
The action of estrogens on the detrusor muscle is controversial. Although Levin et al79 determined that there was an increased density of cholinergic receptors and an increased response to α-adrenergic, muscarinic cholinergic, and purinergic agonists after administration of estrogens in rabbits, other authors found a decrease in the density of muscarinic cholinergic receptors in the urinary bladder muscle80,81 and inhibition of the intracellular influx of calcium ions.82 Whereas the former effects could explain a possible decreased bladder compliance after estrogen supplementation, the latter effects could lead to a decreased detrusor tone and an increased bladder capacity,82,83 which could account for the improvement of the urinary signs attributable to bladder hyperreflexia that are observed in postmenopausal women taking estrogen supplements.22,84 As the actual structural changes occurring in the detrusor muscle in female dogs after sterilization are still unknown, it is difficult to speculate on the cystometric changes to be expected after estrogen replacement in sexually intact or spayed dogs.
The combined administration of estriol and phenylpropanolamine (treatment B) did increase urethral resistance, compared with baseline values, but did not induce changes other than those associated with administration of estriol alone. Furthermore, the urodynamic values decreased after 2 weeks of treatment, despite the continued administration of estriol and the addition of phenylpropanolamine. As the combined treatment was administered to female dogs that had initially received estriol alone for 7 days, it was not possible to differentiate the effect that was attributable to the administration of estriol from the effect that was attributable to the combined treatment. The values obtained after the combined treatment were therefore not compared with baseline values, but were compared with values obtained after the estriol supplementation alone, to analyze the effects of the phenylpropanolamine administration. It appears that phenylpropanolamine added to the estriol supplementation did not induce the changes detected in a previous study,85 in which single daily administration of phenylpropanolamine was effective in increasing the urethral resistance in continent female dogs. One limitation of the present study was that treatment of the dogs with phenylpropanolamine alone was not investigated; if that had been included, we would have been able to compare the effects of phenylpropanolamine administered alone and in combination with the estriol. However, on the basis of our previous experience, we could speculate that the administration of phenylpropanolamine alone would have resulted in a significant increase in urethral pressure, compared with the baseline value. We choose to add the administration of phenylpropanolamine to the treatment with estriol after 7 days of the latter on the basis of the fact that initial treatment with estrogen would stimulate α-adrenergic receptors of the urethra and would increase their sensitivity to sympathetic stimulation.7,45–48,64,65 The reason why no additional increases in the values of the profilometric parameters were detected after the addition of phenylpropanolamine to the initial estriol treatment is unknown at this time. In a study52 in humans, no changes in urodynamic parameters were detected despite the addition of phenylpropanolamine, although clinical improvement of the symptoms was evident. Further studies to assess the clinical efficacy of the combination of phenylpropanolamine and estriol are therefore warranted. The fact that the values of the profilometric parameters were lower after 14 days of treatment could indicate that a certain degree of estrogenic receptor downregulation could have been present after 2 weeks of estriol supplementation. Indeed, Batra and Iosif86 determined that after an initial increase in estrogenic receptor density in the urogenital tract of female rabbits during the first week of estrogen treatment, this density greatly decreased; this suggested that after the initial stimulation of synthesis, a prolonged estrogen treatment could result in receptor downregulation. This theory is reinforced by the decreased activity of peroxidase, a marker of estrogenic activity, after prolonged estrogen treatment in rabbits.86
In the present study involving a limited number of animals, no urodynamic differences between the sexually intact and spayed female dogs were detected prior to treatment. Structural changes in the bladder wall that could possibly lead to modifications of bladder function, such as detrusor hyperreflexia, were not detected in the spayed dogs of our study. Also, modifications of urethral function were not detected in the spayed dogs, which is in disagreement with findings of Reichler et al,20 who detected decreased urethral resistance in female dogs 10 months after ovariectomy. However, compared with sexually intact dogs, threshold pressure was decreased and compliance was increased (albeit not significantly) in spayed dogs. It is interesting to note that these findings are similar to those reported for old sexually intact female dogs without urinary incontinence63 and in female dogs with USMI.87 It must be acknowledged again that the number of dogs in of our study groups was low and that a significant difference in threshold pressure and compliance could not have been detected because of the low power of the study and the fact that the groups were comprised of different animals.
The second purpose of our study was to evaluate the effects of the administration of estriol on the morphologic features of the lower portion of the urogenital tract in sexually intact and spayed female dogs without urinary incontinence. Prior to treatment, vaginal length and width were significantly smaller in spayed female dogs, compared with the values in sexually intact dogs, which is in agreement with previous findings.18,19 Administration of estriol, alone and in combination with phenylpropanolamine, did not modify the morphologic parameters of the urogenital tract in both groups of female dogs in the present study. This finding does not corroborate results of other studies in dogs18,59,88 that identified enlargement of the vagina under the influence of exogenous or endogenous estrogens. During the estrous cycle, morphologic changes are observed in the urogenital tract after 8 days of estrogenic impregnation.63 This influence of the endogenous estrogens on the urogenital tract has been described previously63; in that study, there was a significant increase in all vaginal measurements, as well as an increase in urethral length, during the proestrous phase. It must be noted that the urethral length was not modified by the estrogenic treatments of the present study. Previous studies79,81,82 in rabbits have revealed a significant increase in the weight of the urogenital tissues (including the bladder, which appeared large and distended) after 1 to 3 weeks of estrogen supplementation, with no further changes after 4 weeks of treatment. These changes were probably a result of mucosal hyperplasia. The fact that maturation changes were detected cytologically on the vaginal smears of all the female dogs in our study reinforced the idea that mucosal hyperplasia could have been present in the treated dogs. Therefore, we were assuming that the duration of the treatments in our study would have allowed us to determine morphologic changes in the urogenital tract. However, because of the short half-life of estriol, a longer treatment period is perhaps required to detect morphologic changes; in previous investigations,18,89 morphologic changes in the urogenital organs of dogs and rats were detected after 3 to 4 months of estrogen supplementation or cessation of estrogen supplementation. The findings of the present study question the hypothesis that the positive effects of estrogen supplementation on the resolution of signs of urinary incontinence in female dogs could be attributable to an increase in vaginal mass that is capable of modifying the bladder neck position or the urethral length.
Our data have indicated that there are significant differences in morphologic features of the lower portion of the genital tract (with no associated urodynamic differences) between sexually intact and spayed female dogs. In the present study, the administration of estriol at the recommended dosage for 7 days significantly increased the urethral resistance in sexually intact and spayed female dogs without urinary incontinence. This effect diminished over the subsequent 7-day period, despite the continuation of estriol administration and the addition of phenylpropanolamine to the treatment protocol. Significant changes in bladder function or in the morphologic parameters of the lower portion of the urogenital tract were not detectable after 14 days of estrogen supplementation. These results suggest that estriol mainly acts on the urethral sphincter mechanism and has minimal effects on the morphologic parameters of the urogenital tract. However, as Beagles are not predisposed to USMI,5 further studies involving large numbers of clinical cases are required to draw definite conclusions. Further investigations are also needed to better characterize the structural changes that develop in the lower portion of the urinary tract of female dogs after sterilization and the consequent related changes induced by estrogen replacement in spayed female dogs with urinary incontinence.
ABBREVIATIONS
| USMI | Urethral sphincter mechanism incompetence |
| MUCP | Maximum urethral closure pressure |
| UFPL | Urethral functional profile length |
| MUP | Maximum urethral pressure |
Nickel RF. Studies on the function of the urethra and bladder in continent and incontinent female dogs. PhD thesis, University of Utrecht, Utrecht, Switzerland, 1998;111–126.
Incurin, Intervet Belgium, Malines, Belgium.
Propalin, Vetoquinol SA-BP, Lure Cedex, France.
Diprivan, AstraZeneca SA, Brussels, Belgium.
Nellcor, Nellcor Puritan Bennett Inc, Pleasanton, Calif.
SAS, Version 8, SAS Institute Inc, Cary, NC.
References
- 2.
Holt PE. Urinary incontinence in the bitch due to sphincter mechanism incompetence: prevalence in referred dogs and retrospective analysis of sixty cases. J Small Anim Pract 1985;26:181–190.
- 3.
Thrusfield MV. Association between urinary incontinence and spaying in bitches. Vet Rec 1985;116:695.
- 4.
Krawiec DR. Diagnosis and treatment of acquired canine urinary incontinence. Companion Anim Pract 1989;19:12–20.
- 5.↑
Holt PE, Thrusfield MV. Association in bitches between breed, size, neutering and docking, and acquired urinary incontinence due to incompetence of the urethral sphincter mechanism. Vet Rec 1993;133:177–180.
- 6.↑
Thrusfield MV, Holt PE, Muirhead RH. Acquired urinary incontinence in bitches: its incidence and relationship to neutering practices. J Small Anim Pract 1998;39:559–566.
- 7.↑
Rosin AH, Ross L. Diagnosis and pharmacological management of disorders of urinary continence in the dog. Compend Contin Educ Pract Vet 1981;3:601–610.
- 8.
Richter KP, Ling GV. Clinical response and urethral pressure profile changes after phenylpropanolamine in dogs with primary sphincter incompetence. J Am Vet Med Assoc 1985;187:605–611.
- 9.
Richter KP, Ling GV. Effects of xylazine on the urethral pressure profile of healthy dogs. Am J Vet Res 1985;46:1881–1886.
- 10.
Holt PE. “Simultaneous” urethral pressure profilometry: comparisons between continent and incontinent bitches. J Small Anim Pract 1988;29:761–769.
- 11.↑
Rekers H, Drogendijk AC & Valkenburg HA, et al. The menopause, urinary incontinence and other symptoms of the genitor-urinary tract. Maturitas 1992;15:101–111.
- 12.
Enhörning G. Simultaneous recording of intravesical and intra-urethral pressure: a study of urethral closure in normal and stress incontinent women. Acta Chir Scand Suppl 1961;276:1–68.
- 13.
Haubensak K, Schindler E, Rothaar J. Dependence of resting urethral pressure on age and sex of healthy subjects and on technical details. Urol Int 1975;30:29–32.
- 14.↑
Henriksson L, Andersson K-E, Ulmsten U. The urethral pressure profiles in continent and stress-incontinent women. Scand J Urol Nephrol 1979;13:5–10.
- 15.
Rud T. Urethral pressure profile in continent women from childhood to old age. Acta Obstet Gynecol Scand 1980;59:331–335.
- 16.
Susset J, Plante P. Studies of female urethral pressure profile. Part II. Urethral pressure profile in female incontinence. J Urol 1980;123:70–74.
- 17.↑
Sorensen S. Urethral pressure variations in healthy and incontinent women. Neurourol Urodyn 1992;11:549–591.
- 18.
Holt PE, Gibbs C, Latham J. An evaluation of positive contrast vaginourethrography as a diagnostic aid in the bitch. J Small Anim Pract 1984;25:531–549.
- 19.
Gregory SP, Holt PE & Parkinson TJ, et al. Vaginal position and length in the bitch: relationship to spaying and urinary incontinence. J Small Anim Pract 1999;40:180–184.
- 20.↑
Reichler IM, Pfeiffer E & Piche CA, et al. Changes in plasma gonadotropin concentrations and urethral closure pressure in the bitch during the 12 months following ovariectomy. Theriogenology 2004;62:1391–1402.
- 21.
Faber P, Heidenreich J. Treatment of stress incontinence with estrogen in postmenopausal women. Urol Int 1977;32:221–223.
- 22.
Walter S, Wolf H & Barlebo H, et al. Urinary incontinence in postmenopausal women treated with estrogens. Urol Int 1978;33:135–143.
- 23.↑
Rud T. The effects of estrogens and gestagens on the urethral pressure profile in urinary continent and stress incontinent women. Acta Obstet Gynecol Scand 1980;59:265–270.
- 24.
Hilton P, Stanton SL. The use of intravaginal oestrogen cream in genuine stress incontinence. Br J Obstet Gynaecol 1983;90:940–944.
- 25.
Wilson PD, Faragher B & Butler B, et al. Treatment with oral piperazine sulphate for genuine stress incontinence in postmenopausal women. Br J Obstet Gynaecol 1987;94:568–574.
- 26.
Samsloe G, Jansson I & Meelstrom D, et al. Occurrence, nature and treatment of urinary incontinence in a 75 year old female population. Maturitas 1985;7:335–342.
- 27.
Benness C, Wise BG & Cutner A, et al. Does low dose vaginal estradiol improve frequency and urgency in postmenopausal women? Int Urogynecol J Pelvic Floor Dysfunct 1992;3:281.
- 28.
Cardozo L, Rekers H & Tapp A, et al. Oestriol in the treatment of postmenopausal urgency: a multicentre study. Maturitas 1993;18:47–53.
- 29.↑
Dessole S, Rubattu G & Ambrosini G, et al. Efficacy of lowdose intravaginal estriol on urogenital aging in postmenopausal women. Menopause 2004;11:49–56.
- 30.
Janszen BP, Van Laar PH, Bergman JG. Treatment of urinary incontinence in the bitch: a pilot field study with Incurin. Vet Q 1997;19:S42.
- 31.
Mandigers PJ, Nell T. Treatment of bitches with acquired urinary incontinence with oestriol. Vet Rec 2001;149:764–767.
- 32.
Batra SC, Iosif CS. Female urethra: a target for oestrogen action. J Urol 1983;129:418–420.
- 33.
Ingelman-Sundberg A, Rosen J & Gustafson SA, et al. Cytosol estrogen receptors in the urogenital tissues in stress incontinent women. Acta Obstet Gynecol Scand 1981;60:585–586.
- 34.
Iosif CS, Batra SC & Ek A, et al. Estrogen receptors in the human female lower urinary tract. Am J Obstet Gynecol 1981;141:817–820.
- 35.
Urner F, Weil A, Herrmann WL. Estradiol receptors in the urethra and the bladder of the female rabbit. Gynecol Obstet Invest 1983;16:307–313.
- 36.↑
Schulze H, Barrack ER. Immunocytochemical localization of estrogen receptors in the normal male and female canine urinary tract and prostate. Endocrinology 1987;121:1773–1783.
- 37.↑
Clark JH, Markaverich BM. The agonistic and antagonistic actions of estriol. J Steroid Biochem 1984;20:1005–1013.
- 38.↑
Molander U. Urinary incontinence and related urogenital symptoms in elderly women. Acta Obstet Gynecol Scand Suppl 1993;158:1–22.
- 39.↑
Hendriks S, Janszen B. Safety trial in bitches with tablets containing oestriol (Incurin), in Proceedings. 1st Cong Eur Vet Soc Small Anim Reprod 1998;318.
- 40.
Ek A, Andersson KE & Gullberg B, et al. Effects of oestradiol and combined norephedrin and oestradiol treatment on female stress incontinence. Zentralbl Gynakol 1980;102:839–844.
- 41.
Fantl JA, Bump RC & Robinson D, et al. Efficacy of estrogen supplementation in the treatment of urinary incontinence. Obstet Gynecol 1996;88:745–749.
- 42.
Jackson S, Shepherd A & Brookes S, et al. The effect of oestrogen supplementation on post-menopausal urinary stress incontinence: a double-blind placebo-controlled trial. Br J Obstet Gynaecol 1999;106:711–718.
- 43.
Iosif CS. Effects of protracted administration of estriol on the lower genito urinary tract in postmenopausal women. Arch Gynecol Obstet 1992;251:115–120.
- 44.↑
Nickel RF. Oestriol: pharmacology and effects on lower urinary tract function. ESVIM Newsletter 1999;9:13–15.
- 45.
Hodgson BJ, Dumas S & Bolling DR, et al. Effect of estrogen on sensitivity of rabbit bladder and urethra to phenylephrine. Invest Urol 1978;16:67–69.
- 46.
Osborne CA, Oliver JE, Polzin DE. Non-neurogenic urinary incontinence. In: Kirk RW, ed. Current veterinary therapy VII: small animal practice. Philadelphia: WB Saunders Co, 1980;1128–1136.
- 47.↑
Creed KE. Effect of hormones on urethral sensitivity to phenylephrine in normal and incontinent dogs. Res Vet Sci 1983;34:177–181.
- 48.
Callahan SM, Creed KE. The effects of oestrogens on spontaneous activity and responses to phenylephrine of the mammalian urethra. J Physiol 1985;358:35–46.
- 49.
Beisland HO, Fossberg E & Moer A, et al. Urethral insufficiency in postmenopausal females: treatment with phenylpropanolamine and estriol separately and in combination. Urol Int 1984;39:211–216.
- 50.
Kinn AC, Lindskog M. Estrogens and phenylpropanolamine in combination for stress urinary incontinence in postmenopausal women. Urology 1988;32:273–280.
- 51.
Ahlstrom K, Sandahl B & Sjoberg B, et al. Effect of combined treatment with phenylpropanolamine and estriol, compared with estriol treatment alone, in postmenopausal women with stress urinary incontinence. Gynecol Obstet Invest 1990;30:37–43.
- 52.↑
Hilton P, Tweddell AL, Mayne C. Oral and intravaginal estrogens alone and in combination with alpha-adrenergic stimulation in genuine stress incontinence. Int Urogynecol J Pelvic Floor Dysfunct 1990;1:80–86.
- 53.
Walter S, Kjaergaard B & Lose G, et al. Stress urinary incontinence in postmenopausal women treated with oral estrogen (Estriol) and an alpha-adrenoceptor-stimulating agent (Phenylpropanolamine): a randomized double-blind placebo-controlled study. Int Urogynecol J Pelvic Floor Dysfunct 1990;1:74–79.
- 54.
Schreiter F, Fuchs P, Stockamp K. Estrogenic sensitivity of alpha-receptors in the urethral musculature. Urol Int 1976;31:13–19.
- 55.↑
Johnston S, Root Kustritz M, Olson P. Vaginal cytology. In: Johnston S, Root Kustritz M, Olson P, eds. Canine and feline theriogenology. Philadelphia: WB Saunders Co, 2001;32–40.
- 56.↑
Institute of Laboratory Animal Research, Commission on Life Sciences, National Research Council. Guide for the care and use of laboratory animals. Washington, DC: National Academies Press, 1996;1–125.
- 57.↑
Holt PE, Gibbs C, Wathes CM. Simultaneous urethral pressure profilometry using a microtip transducer catheter in the bitch: effects of bitch position and transducer orientation. Neurourol Urodyn 1990;9:281–296.
- 58.↑
Hamaide AJ, Verstegen JP & Snaps FR, et al. Validation and comparison of the use of diuresis cystometry and retrograde filling cystometry at various infusion rates in female Beagle dogs. Am J Vet Res 2003;64:574–579.
- 59.↑
Holt PE. Positive-contrast vaginourethrography for diagnosis of lower urinary tract disease. In: Kirk RW, Bonagura JD, eds. Current veterinary therapy X: small animal practice. Philadelphia: WB Saunders Co, 1989;1142.
- 60.↑
International Continence Society. First report on the standardisation of terminology of lower urinary tract function. Br J Urol 1976;48:39–42.
- 61.↑
Lentner C. Statistical methods. In: Introduction to statistics. Basel, Switzerland: Ciba-Geigy, 1982;182–236.
- 62.↑
Wolters M, Methfessel HD & Goepel C, et al. Computer-assisted virtual urethral pressure profile in the assessment of female genuine stress incontinence. Obstet Gynecol 2002;99:69–74.
- 63.↑
Hamaide AJ, Verstegen JP & Snaps FR, et al. Influence of the estrous cycle on urodynamic and morphometric measurements of the lower portion of the urogenital tract in dogs. Am J Vet Res 2005;66:1075–1083.
- 64.
Caine M, Raz S. Some clinical implications of adrenergic receptors in the urinary tract. Arch Surg 1975;110:247–250.
- 65.
Ekstrom J, Iosif CS, Malmberg L. Effects of long-term treatment with estrogen and progesterone on in vitro muscle responses of the female rabbit urinary bladder and urethra to autonomic drugs and nerve stimulation. J Urol 1993;150:1284–1288.
- 66.↑
Holt PE. “Simultaneous” urethral pressure profilometry in the bitch: methodology and reproducibility of the technique. Res Vet Sci 1989;47:110–116.
- 67.↑
Larsson B, Andersson KE & Batra S, et al. Effects of estradiol on norepinephrine-induced contraction, alpha-adrenoceptor number and norepinephrine content in the female rabbit urethra. J Pharmacol Exp Ther 1984;229:557–563.
- 68.
Jackson S, Avery N & Shepherd A, et al. The effect of oestradiol on vaginal collagen in post-menopausal women with stress urinary incontinence. Neurourol Urodyn 1996;15:327–328.
- 69.
James M, Avery N & Jackson S, et al. The pathophysiological changes of vaginal tissue in women with stress urinary incontinence: a controlled trial. Neurourol Urodyn 1999;18:283–284.
- 70.
Batra S, Bjellin L & Iosif S, et al. Effects of oestrogen and progesterone on the blood flow in the lower urinary tract of the rabbit. Acta Physiol Scand 1985;123:191–194.
- 71.
Versi E, Cardozo LD. Urethral instability: diagnosis based on variations in the maximum urethral pressure in normal climacteric women. Neurourol Urodyn 1986;5:535–541.
- 72.
Batra S, Bjellin L & Sjogren C, et al. Increases in blood flow of the female rabbit urethra following low dose estrogens. J Urol 1986;136:1360.
- 73.
Klutke JJ, Bergman A. Hormonal influence on the urinary tract. Urol Clin North Am 1995;22:629–639.
- 74.↑
Semmens JP, Tsai CC & Semmens EC, et al. Effects of estrogen therapy on vaginal physiology during the menopause. Obstet Gynecol 1985;66:15–18.
- 75.↑
Augsburger HR, Cruz-Orive LM. Stereological analysis of the urethra in sexually intact and spayed female dogs. Acta Anat (Basel) 1995;154:135–142.
- 76.↑
Smith PG, Bradshaw S. Innervation of the proximal urethra of ovariectomized and estrogen-treated female rats. Histol Histopathol 2004;19:1109–1116.
- 77.↑
Augsburger HR, Cruz-Orive LM. Influence of ovariectomy on the canine striated external urethral sphincter (M. urethralis): a stereological analysis of slow and fast twitch fibres. Urol Res 1998;26:417–422.
- 78.↑
Vanderhorst VG, Holstege G. Estrogen induces axonal outgrowth in the nucleus retroambiguus-lumbosacral motoneuronal pathway in the adult female cat. J Neurosci 1997;17:1122–1136.
- 79.↑
Levin RM, Shofer FS, Wein AJ. Estrogen-induced alterations in the autonomic responses of the rabbit urinary bladder. J Pharmacol Exp Ther 1980;215:614–618.
- 80.
Shapiro E. Effect of estrogens on the weight and muscarinic cholinergic receptor density of the rabbit bladder and urethra. J Urol 1986;135:1084–1087.
- 81.
Batra S, Anderson KE. Oestrogen-induced changes in muscarinic receptor density and contractile responses in the female rabbit urinary bladder. Acta Physiol Scand 1989;137:135–141.
- 82.↑
Elliott RA, Castleden CM & Miodrag A, et al. The direct effects of diethylstilboestrol and nifedipine on the contractile responses of isolated human and rat detrusor muscles. Eur J Clin Pharmacol 1992;43:149–155.
- 83.
Elliott RA, Castleden CM & Miodrag A, et al. The effect of in vivo oestrogen pre-treatment on the contractile response of rat isolated detrusor muscle. Br J Pharmacol 1992;107:766–770.
- 84.
Fantl JA, Wyman JF & Anderson RL, et al. Post-menopausal urinary incontinence: comparison between non-estrogen and estrogen supplemented women. Obstet Gynecol 1988;71:823–828.
- 85.↑
Carofiglio F, Hamaide AJ & Farnir F, et al. Evaluation of the urodynamic and hemodynamic effects of orally administered phenylpropanolamine and ephedrine in female dogs. Am J Vet Res 2006;67:723–730.
- 86.↑
Batra S, Iosif CS. Effect of estrogen treatment on the peroxidase activity and estrogen receptors in the female rabbit urogenital tissues. J Urol 1992;148:935–938.
- 87.↑
Nickel RF, van den Brom WE. Simultaneous diuresis cysto-urethrometry and multichannel urethral pressure profilometry in female dogs with refractory urinary incontinence. Am J Vet Res 1997;58:691–696.
- 88.
Sokolowski JH, Zimbelman RG, Goyings LS. Canine reproduction: reproductive organs and related structures of the nonparous, parous, and postpartum bitch. Am J Vet Res 1973;34:1001–1013.
- 89.
Longhurst PA, Kauer J & Leggett RE, et al. The influence of ovariectomy and estradiol replacement on urinary bladder function in rats. J Urol 1992;148:915–919.