- Email: [email protected]
Increased blood flow after catheterization and drainage in the chronically obstructed rabbit urinary bladder
BASIC SCIENCE
INCREASED BLOOD FLOW AFTER CATHETERIZATION AND DRAINAGE IN THE CHRONICALLY OBSTRUCTED RABBIT URINARY BLADDER ¨ DER, BARRY A. KOGAN, JEREMY LIEB, ANNETTE SCHRO
AND
ROBERT M. LEVIN
ABSTRACT Objectives. To determine the effect of drainage on rabbit bladder blood flow after 4 weeks of partial outlet obstruction. Previous studies have shown that catheterization and drainage of the urinary bladder in control rabbits resulted in a significant nitric oxide-induced increase of blood flow to the bladder. It was also shown that 4 weeks’ partial outlet obstruction caused a significant decrease in blood flow to the bladder. Methods. Male New Zealand White rabbits underwent partial outlet obstruction by standard methods. After 4 weeks, the blood flow to the bladder muscle and mucosa was determined by a microsphere technique. Within 1 to 2 minutes after transurethral catheterization and complete drainage of the bladder, the blood flow was again determined. Unobstructed animals served as controls. Four other control animals underwent a repetitive blood flow study during 10 minutes to determine the time frame of blood flow changes after drainage. Blood flow was also measured in 2 control rabbits after transurethral catheterization without drainage and in 2 control rabbits after drainage by suprapubic puncture. To exclude the possibility that increased intravesical pressure alters the blood flow measurements, the relationship between the intravesical volume and the bladder pressure was examined in the obstructed rabbits. Results. After drainage of the bladder, the blood flow to the bladder muscle increased 4.5-fold in the decompensated obstructed group (bladder weights greater than 15 g) and 2.5-fold in the compensated animals (bladder weights less than 5 g) and control animals. Blood flow to the mucosa followed the same pattern but without reaching significance. Blood flow returned to near baseline values within 5 minutes. Catheterization without drainage did not alter the blood flow. In contrast, drainage by puncture increased the blood flow significantly. Higher intravesical volumes increased the intravesical pressure slightly, but after opening the abdominal fascia, the intravesical pressure did not change with increasing volumes. Conclusions. Although the previously shown decreased blood flow to the bladder smooth muscle may be an etiologic factor in bladder contractile dysfunction secondary to partial outlet obstruction, the bladder does have the ability to increase the blood flow after drainage. This ability could be a compensatory and possibly protective mechanism after outlet obstruction. UROLOGY 58: 295–300, 2001. © 2001, Elsevier Science Inc.
C
hronic bladder outlet obstruction (eg, resulting from benign prostatic hyperplasia) causes several changes in bladder physiology in humans and animal models.1– 6 The reason why the status of These projects were supported by grants from the Department of Veterans Affairs and National Institutes of Health grants RO1DK26508, RO1-DK53965, and RO1-DK47949. From the Department of Urology, Johannes-Gutenberg-University, Mainz, Germany; Albany Medical College, Albany College of Pharmacy, and Stratton Veterans Affairs Medical Center, Albany, New York Dr. Schro¨der is currently at the Department of Clinical Pharmacology, Lund University Hospital, Lund, Sweden. Reprint requests: Robert M. Levin, Ph.D., Albany College of Pharmacy, 106 New Scotland Avenue, Albany, NY 12208 Submitted: November 27, 2000, accepted (with revisions): March 23, 2001 © 2001, ELSEVIER SCIENCE INC. ALL RIGHTS RESERVED
decompensated bladder function is reversible in some individuals and irreversible in others remains unknown. Previous studies by other groups have shown that long-term partial obstruction of the urinary bladder leads to a significant decrease in blood flow to the bladder muscle,7–10 and this phenomenon is thought to be one reason for the damage to the bladder caused by outlet obstruction. We obtained similar results in a study of long-term outlet obstruction on blood flow to the rabbit bladder.11 The decrease in blood flow after obstruction correlated with progressive contractile failure and increased amounts of residual urine.11 Several recent studies have reported the effects of distension, bladder filling, and acute and chronic obstruction on bladder blood flow, all using different animal 0090-4295/01/$20.00 PII S0090-4295(01)01142-6 295
models and methods of blood flow measurements.12–14 In general, these studies indicate that bladder distension during filling and resulting from acute and chronic obstruction causes decreased blood flow.12–14 Previous studies in control rabbits showed that catheterization and drainage of the urinary bladder resulted in a significant nitric oxide (NO)-induced increase of blood flow to the bladder within 1 to 2 minutes after drainage.15 It was also shown that intermittent catheterization and drainage of partially obstructed bladders prevented smooth muscle dysfunction.16 We used the animals from our long-term obstruction study to look for alterations in the pattern of blood flow changes after catheterization and drainage. MATERIAL AND METHODS A standardized method was used to create moderate bladder outlet obstruction in 12 male New Zealand White rabbits (⬃3.5 kg) (Milbrook Farms, Amherst, Mass).4,6 The animals were sedated with intramuscular ketamine-xylazine (25 mg/kg ketamine, 10 mg/kg xylazine); surgical anesthesia was maintained with intravenous sodium pentobarbital (25 mg/ kg). Under sterile conditions, the urinary bladder was catheterized and drained with an 8F catheter. The bladder and urethrovesical junction were exposed through a lower abdominal midline incision. After removal of the perivesical fat, a 2-0 silk ligature was tied loosely around the vesical outlet, approximately 1 to 2 cm distal to the insertion of the ureters. The catheter was removed. The bladder was returned to the normal position and the wound closed anatomically. Intramuscular gentamicin (1 mg/kg) and buprenorphine hydrochloride (0.3 mg/kg) were given on postoperative days 1 and 2. All obstructions were performed by the same surgeon. Unobstructed animals served as the controls. Previous studies showed that sham surgery had no effect on blood flow to the bladder muscle or mucosa17; therefore, no sham operations were done in this study. Four different aspects of bladder function after outlet obstruction were examined in this study. 1. Changes in blood flow after catheterization and drainage: after 4 weeks of obstruction, the rabbits were anesthetized as described above, and blood flow was determined by a standardized fluorescent microsphere infusion technique.15,17,18 To avoid any interference with the blood flow measurements, the abdominal wall remained closed and the bladder was not manipulated during the blood flow determinations. The right carotid and femoral arteries were exposed surgically and cannulated with polyethylene 160 tubing (inside diameter 1.14 mm, outside diameter 1.57 mm). The carotid tubing was connected to a pressure transducer and advanced into the left ventricle. Sphere infusion was done by injecting 2.5 ⫻ 106 NuFlow fluorescent microspheres, 15 m in diameter (Interactive Medical Technologies, North Hollywood, Calif) using an infusion pump connected to the left ventricular catheter. Using a withdrawal pump (1.72 mL/min), a reference blood sample was withdrawn from the femoral artery, starting 15 seconds before and ending 60 seconds after sphere infusion. Then, the bladder was catheterized transurethrally with an 8F catheter, drained completely, and the volume of urine recorded. Within 60 to 90 seconds after emptying the bladder, a second blood flow determination procedure was done using a different color of fluorescent microspheres. 2. Time frame in which the blood returned to baseline after drainage: a repetitive blood flow analysis was performed in 4 296
control rabbits before and 1, 5, and 10 minutes after drainage, using four different colors of microspheres in each rabbit, as described above. The systemic arterial blood pressure was monitored throughout all microsphere procedures until the experiment ended. 3. Effects of transurethral catheterization without emptying and drainage by suprapubic puncture on bladder blood flow: to determine whether the change in blood flow was caused by the act of urethral catheterization or by the bladder emptying itself, the bladders of 2 normal control rabbits were emptied by suprapubic puncture. Two other control rabbits were catheterized transurethrally without bladder emptying. The initial and post-catheterization microsphere procedures were performed as described above. 4. Influence of bladder filling on intravesical pressure: to exclude the possibility that the baseline blood flow was artificially increased because of high intravesical pressures, we measured the bladder pressure under natural filling in an additional 14 obstructed rabbits before (by percutaneous puncture) and after surgical opening of the abdominal wall.
BLOOD FLOW ANALYSIS Under pentobarbital anesthesia, the bladder was excised, weighed, and separated into muscle and mucosa. Reference blood and detrusor samples were sent to Interactive Medical Technologies. For microsphere concentration determination, tissue samples were subjected to alkaline digestion, and the fluorescent microspheres from each sample were collected on a filter, resuspended, and quantified using flow cytometry. Microsphere concentrations of tissue samples were compared with those of reference blood samples to determine tissue perfusion; blood flow is expressed as milliliters of blood per gram tissue per minute (mL/g/min). Full-thickness strips from each bladder were used for in vitro contractility studies. The obstructed bladders were separated into compensated and decompensated groups on the basis of their contractile responses to electrical field stimulation.11
STATISTICAL ANALYSIS
All values are reported as the mean ⫾ SEM. Statistical significance was determined by Bonferroni analysis or Student’s t test, as appropriate. P ⬍0.05 was required for statistical significance.
RESULTS Partial outlet obstruction resulted in variable increases in bladder weight and in the volume of urine in the bladder at the time of blood flow measurement. The mean bladder weight in the obstructed group and the unoperated control group was 10.7 ⫾ 2.0 g (range 3.5 to 22.1) and 2.6 ⫾ 0.3 g (range 1.6 to 3.1), respectively (P ⬍0.05). To evaluate a potential relationship between the bladder weight and blood flow, the obstructed rabbits were divided into three different groups according to bladder weight. The degree of neurogenic contractile failure in the in vitro studies correlated highly with the increase in bladder weight. Therefore, we defined bladders weighing less than 5 g (3.1 ⫾ 1.4, n ⫽ 4) as compensated, bladders weighing 5.1 to 15 g (9.9 ⫾ 0.9, n ⫽ 4) as intermediate-decompensated, and bladders weighing greater than 15 g (18.0 ⫾ 1.4, n ⫽ 4) as severely UROLOGY 58 (2), 2001
FIGURE 1. Changes in blood flow to the bladder muscle after drainage. Each bar represents the mean ⫾ SEM of 5 control, 4 compensated, 4 intermediate-decompensated, and 4 decompensated obstructed bladders. Asterisks indicate P ⬍0.05 versus blood flow before drainage; plus signs indicate P ⬍0.05 versus control before drainage.
decompensated. Complete information can be found in Schro¨der et al.11 1. Changes in blood flow after catheterization and drainage: the blood flow to the smooth muscle of the controls was 0.08 ⫾ 0.01 mL/g/min, similar to that of the compensated obstructed bladders (0.08 ⫾ 0.02 mL/g/min). The blood flow to the smooth muscle of the intermediate-decompensated bladders was reduced significantly from that of the controls (0.04 ⫾ 0.01 mL/g/min). The blood flow to the severely decompensated smooth muscle was reduced even further (0.03 ⫾ 0.01 mL/g/ min) (Fig. 1). The blood flow to the mucosa of the obstructed bladders was unchanged compared with the controls (Table I). Depending on the degree of compensation, a variable increase in blood flow after catheterization was found. Similar to the controls, the obstructed bladders weighing less than 5 g had a 2.3-fold increase in blood flow after drainage of the bladder. The bladders weighing greater than 5 g had a 4.6-fold increase in blood flow after drainage (Figs. 1 and 2). The blood flow to the mucosa followed similar patterns, but without reaching a significant difference in most cases (Table I). 2. Time frame in which the blood returned to baseline after drainage—repetitive blood flow measurements: repetitive measurement of blood flow 1, 5, and 10 minutes after drainage showed that the blood flow increased within the first minute after drainage and returned to near baseline values within 5 minutes (Fig. 3). The systemic blood pressure remained stable and was not affected by the repetitive microsphere injections. 3. Effects of transurethral catheterization withUROLOGY 58 (2), 2001
out emptying and drainage by suprapubic puncture on bladder blood flow: in the four control animals in which we determined the effects of suprapubic puncture (n ⫽ 2) versus transurethral catheterization (n ⫽ 2), we found an increase in the blood flow to the bladder after drainage by suprapubic puncture (0.05 ⫾ 0.01 mL/g/min before drainage and 0.47 ⫾ 0.23 mL/g/min after drainage). Transurethral catheterization without drainage caused no changes (0.05 ⫾ 0.002 mL/g/min before and 0.07 ⫾ 0.02 mL/g/min after insertion and removal of the catheter). 4. Influence of bladder filling on intravesical pressure: in 14 obstructed rabbits (mean bladder weight 11.67 ⫾ 0.74 g), we measured the bladder pressure under natural filling before (by percutaneous puncture) and after surgical opening of the abdominal wall. The mean bladder volume was 160 ⫾ 20 mL. Intravesical pressure with the abdominal wall closed was 9.0 ⫾ 0.9 cm H2O and with the wall open was 5.5 ⫾ 0.5 cm H2O. Bladders containing large urine volumes had intravesical pressures about 5 cm H2O higher than those with low urine volumes when the abdominal wall was closed (Fig. 4). There was no influence of urine volume on bladder pressure once the abdominal wall was opened. COMMENT Drainage of the bladder causes a significant increase in blood flow to smooth muscle and mucosa in both the obstructed bladders and control animals. The increase of blood flow to the muscle caused by catheterization and drainage of the bladder was 2.5-fold in the control and compensated groups (less than 5 g bladder weight) but significantly higher (4.6-fold) in the decompensated groups. The more severe the functional decompensation was, according to the contractile response in vitro and the bladder weight, the lower were the baseline perfusion levels.11 This suggests that the increase of blood flow after drainage is a compensatory mechanism, responding to decreased baseline perfusion. The functionally severely decompensated bladders with weights greater than 15 g, although increasing their initial blood flow 4.5fold, did not reach the same blood flow values as the controls and less decompensated bladders. This could be caused by the severe damage of the bladder muscle, which would also affect the function of the blood vessels. It was shown in several other studies using Doppler flowmetry that bladder contractions (and thereby increased intravesical pressure) caused a reversible decrease of blood flow.7,14,19 –21 An increase in bladder perfusion after drainage was also demonstrated.14 A similar situation occurs in the 297
TABLE I. Blood flow to the mucosa before and after drainage Blood Flow to Mucosa (mL/g/min) Group Controls ⬍5 g 5.1–15 g ⬎15 g
Before Drainage 0.226 0.331 0.101 0.203
⫾ ⫾ ⫾ ⫾
0.041 0.101 0.101 0.019
After Drainage 0.621 1.070 1.291 1.088
⫾ ⫾ ⫾ ⫾
0.175 0.193 0.271* 0.413
Increase After Drainage (%) 222 222 327 323
⫾ ⫾ ⫾ ⫾
45 66 55 40
Data presented as the mean ⫾ SEM from 5 control, 4 compensated (⬍5 g), 4 intermediate-decompensated (5.1–15 g), and 4 decompensated (⬎15 g) obstructed bladders. * P ⬍0.05 vs. blood flow before drainage.
FIGURE 2. Changes in blood flow to the bladder muscle (expressed as percentage). Each bar represents the mean ⫾ SEM of 5 control, 4 compensated, 4 intermediate-decompensated, and 4 decompensated bladders.
FIGURE 3. Repetitive blood flow determination after drainage in 4 individual animals. The points represent the blood flow values obtained for each control rabbit at the different times. The bar represents the average period during which the second blood flow determination was done for the experiments shown in Figures 2 and 3.
heart. High pressure within the cardiac muscle during systole compresses the coronary vessels, decreasing blood flow. The blood flow then increases rapidly above baseline at the beginning of diastole and decreases slightly during diastole.22 298
FIGURE 4. Scatter plot and linear regression analysis of intravesical pressure relative to intravesical volume measured with intact and opened abdominal wall. Each pair of points represents the intravesical pressure for an individual rabbit before (black circles) and after (white circles) the opening of the abdominal wall.
Bladder contractions against an obstructed bladder neck reduced the blood flow significantly more than contractions against a nonobstructed bladder neck.7,14,20,21 When emptying against increased outflow resistance, the voiding phase is prolonged and, depending on the degree of decompensation, the poor force of detrusor contractions may further increase the workload on the bladder. Overall, an increased metabolic demand is expected. In most vascular beds, the blood flow is regulated by coordinated interactions between local myogenic tone, flow-induced dilation, and the washout of vasodilator metabolites. After metabolite-induced dilation, enhanced flow and stress on the vascular endothelium will increase the production and release of NO, which in turn further improves perfusion.23 In a recent study in control rabbits, the basal bladder smooth muscle blood flow was unaffected by treatment with the NO synthase (NOS) inhibitor, N-nitro-L-arginine methyl ester (L-NAME), and increases in blood flow after catheterization were prevented by NOS inhibition.15 This may explain the blood flow responses obtained in this report in the obstructed bladders, in which an increased UROLOGY 58 (2), 2001
metabolic demand is expected to exist. Further implicating a role for NO in the obstructed bladder, the expression of the inducible form of NOS, iNOS, was enhanced in obstructed bladders in wild-type mice.24 Mice incapable of producing iNOS failed to respond to obstruction with the expected increases in mass and functional changes observed in the wild type.25 Therefore, increased NO expression was suggested to be a mechanism by which the bladder responds to overcome or improve the increased metabolic demand during obstruction-induced ischemia. After acute urinary retention, an increase in the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH)-diaphorase staining (an indirect marker for NOS) in intramural ganglia has been described for the guineapig bladder.26 Possible neuromodulatory functions for NO at a ganglionic level in regulation of vascular tone may be considered. The repetitive determination of blood flow during the first 10 minutes after drainage revealed that the increase of blood flow after drainage is temporary and that the blood flow returns to near baseline values within minutes. This indicates that the blood flow measured before drainage can in fact be considered the baseline value and the increase in blood flow stimulated by drainage is an acute phenomenon, unrelated to the intravesical pressure. Multiple injections of different colored microspheres have proved to be a reliable method of regional blood flow determinations.27–29 Nonetheless, it is a limitation of our study that the microsphere technique, although very accurate, does not allow a continuous measurement of perfusion over a longer period. To exclude irritation of the bladder or urethra as a reason for the altered blood flow, two control animals were catheterized transurethrally without drainage. The blood flow was determined before and after the manipulation, which had no effect on the perfusion of bladder muscle or mucosa. In contrast, drainage of the bladder by suprapubic puncture caused a significant increase in blood flow to the bladder muscle and mucosa. This shows that bladder emptying is the reason for the alterations in blood flow in this study, not catheter manipulation. One might assume that the decreased blood flow before drainage is due to high intravesical pressure, caused by large amounts of residual urine in the functionally decompensated bladders, and the increase of blood flow afterward results from relief of the pressure. This explanation has already been excluded by the finding that the blood flow returns to near baseline values within 5 minutes after complete drainage. Nonetheless, the intravesical pressure is of great importance when blood flow is investigated. In the 14 additional obstructed rabbits, the intravesical pressure increased slightly at UROLOGY 58 (2), 2001
higher urine volumes, but even the highest amount of residual urine (approximately 300 mL) caused intravesical pressure values less than 15 cm H2O. Bladders containing large urine volumes had intravesical pressures about 5 cm H2O higher than those with low urine volumes when the abdominal wall was closed. This difference was not statistically significant and was less than the normal micturition pressure of about 25 cm H2O in male rabbits. After opening the abdominal wall, the pressure in the bladders containing large volumes of urine (greater than 200 mL) decreased to 5 to 6 cm H2O. This suggests that the high intravesical pressures observed in large-volume bladders are the result of compression by the abdominal contents. These findings also support the concept that the decreased blood flow observed in the decompensated bladders in this study was not caused by an increase in intravesical pressure. Nonetheless, residual urine does seem to play a role in the process of decompensation. A previous study showed that intermittent catheterization and complete drainage (every 8 hours) after outlet obstruction, beginning immediately after creation of the obstruction, significantly reduced the degree of bladder hypertrophy and the development of contractile dysfunction.30 Although the effect of partial drainage of the bladder was not investigated in this study, our results show that complete drainage has a beneficial effect compared with obstruction with untreated residual urine. It has been hypothesized that the phenomenon of the severe decrease of blood flow during micturition against partial outlet obstruction, although transient, is an important cause of damage to the bladder muscle.7,14,20,21 The increase of perfusion after the emptying of the bladder may be a compensatory mechanism. The finding that this effect is found in functionally impaired bladders supports this assumption and suggests an important therapeutic intervention. Whether partial drainage of the bladder leads to the same effect will be investigated during future studies. It is still controversial whether the obstructioninduced decrease of bladder muscle blood flow leads to contractile failure or if it is a consequence. Nonetheless, the alterations in basal blood flow after outlet obstruction, which correlate with in vitro and in vivo dysfunction, as well as changes in the response to drainage, show that bladder blood flow plays a major role in the process of decompensation. CONCLUSIONS The enhanced increase of blood flow after drainage of the bladder seems to be a compensating response to the overall decreased blood flow to the 299
bladder smooth muscle after outlet obstruction. From these results, we suggest that chronically increased amounts of residual urine, which would prevent this effect, could be the reason for the progressive damage of the smooth muscle. ACKNOWLEDGMENT. To Catherine Whitbeck and Patrick Horan for their technical assistance and to Penelope A. Longhurst for constructive criticism and support. REFERENCES 1. Kuo HC: Clinical prostate score for diagnosis of bladder outlet obstruction by prostate measurements and uroflowmetry. Urology 54: 90 –96, 1999. 2. Sonke GS, Heskes T, Verbeek AL, et al: Prediction of bladder outlet obstruction in men with lower urinary tract symptoms using artificial neural networks. J Urol 163: 300 – 305, 2000. 3. Kato K, Monson FC, Longhurst PA, et al: The functional effects of long-term outlet obstruction on the rabbit urinary bladder. J Urol 143: 600 – 606, 1990. 4. Buttyan R, Chen MW, and Levin RM: Animal models of bladder outlet obstruction and molecular insights into the basis for the development of bladder dysfunction. Eur Urol 32(suppl 1): 32–39, 1997. 5. Levin RM, Haugaard N, Mogavero L, et al: Biochemical evaluation of obstructive bladder dysfunction in men secondary to BPH: a preliminary report. Urology 53: 446 – 450, 1999. 6. Levin RM, Brading AF, Mills IW, et al: Experimental models of bladder obstruction, in Lepor H (Ed): Prostatic Disease. Philadelphia, WB Saunders, 2000, pp 169 –196. 7. Greenland JE, Hvistendahl JJ, Andersen H, et al: The effect of bladder outlet obstruction on tissue oxygen tension and blood flow in the pig bladder. BJU Int 85: 1109 –1114, 2000. 8. Lin AT, Chen MT, Yang CH, et al: Blood flow of the urinary bladder: effects of outlet obstruction and correlation with bioenergetic metabolism. Neurourol Urodyn 14: 285– 292, 1995. 9. Nielsen KK: Blood flow rate and total blood flow related to length density and total length of blood vessels in mini-pig urinary bladder after chronic outflow obstruction and after recovery from obstruction. Neurourol Urodyn 14: 177–186, 1995. 10. Tong-Long Lin A, Chen KK, Yang CH, et al: Recovery of microvascular blood perfusion and energy metabolism of the obstructed rabbit urinary bladder after relieving outlet obstruction. Eur Urol 34: 448 – 453, 1998. 11. Schro¨der A, Chichester P, Kogan BA, et al: Effect of chronic bladder outlet obstruction on the blood flow of the rabbit urinary bladder. J Urol 165: 640 – 646, 2001. 12. Finkbeiner A, and Lapides J: Effect of distension on blood flow in dogs urinary bladder. Invest Urol 12: 210 –212, 1974. 13. Nemeth CJ, Khan RM, Kirchner P, et al: Changes in canine bladder perfusion with distension. Invest Urol 15: 149 –150, 1977. 14. Azadzoi KM, Pontari M, Vlachiotis J, et al: Canine bladder blood flow and oxygenation: changes induced by filling,
300
contraction and outlet obstruction. J Urol 155: 1459 –1465, 1996. 15. Lieb J, Kogan BA, Das AK, et al: The effect of urine volume and nitric oxide on basal bladder blood flow: response to catheterization and drainage. Neurourol Urodyn 20: 115– 124, 2001. 16. Ohnishi N, Horan P, Levin SS, et al: Intermittent catheterization limits rabbit bladder dysfunction in response to partial outlet obstruction. J Urol 163: 292–295, 2000. 17. Lieb J, Chichester P, Kogan BA, et al: Rabbit urinary bladder blood flow changes during the initial stage of partial outlet obstruction. J Urol 164: 1390 –1397, 2000. 18. Shabsigh A, Hayek OH, Weiner D, et al: Acute increase in blood flow to the rat bladder subsequent to partial bladder outlet obstruction. Neurourol Urodyn 19: 195–206, 2000. 19. Brading AF, Greenland JE, Mills IW, et al: Blood supply to the bladder during filling. Scand J Urol Nephrol Suppl 201: 25–31, 1999. 20. Greenland JE, and Brading AF: Urinary bladder blood flow changes during the micturition cycle in a conscious pig model. J Urol 156: 1858 –1861, 1996. 21. Greenland JE, and Brading AF: The effect of bladder outflow obstruction on detrusor blood flow changes during the voiding cycle in conscious pigs. J Urol 165: 245–248, 2001. 22. Guyton AC: The coronary circulation and ischemic heart disease, in Guyton AC (Ed): A Textbook of Medical Physiology. Philadelphia, WB Saunders, 1986, pp 295–304. 23. Busse R, and Fleming I: Nitric oxide and the regulation of the vascular tone, in Mayer B (Ed): Handbook of Experimental Pharmacology, Vol 143. Berlin, Springer-Verlag, 2000, pp 179 –198. 24. Lemack GE, Zimmern PE, Vazquez D, et al: Altered response to partial bladder outlet obstruction in mice lacking inducible nitric oxide synthase. J Urol 163: 1981–1987, 2000. 25. Lemack GE, Burkhard F, Zimmern PE, et al: Physiologic sequelae of partial infravesical obstruction in the mouse: role of inducible nitric oxide synthase. J Urol 161: 1015–1022, 1999. 26. Zhou Y, and Ling EA: Increased NADPH-diaphorase reactivity in bladder afferent pathways following urethral obstruction in guinea pigs. J Peripher Nerv Syst 2: 333–342, 1997. 27. Walter B, Bauer R, Gaser E, et al: Validation of the multiple colored microsphere technique for regional blood flow measurements in newborn piglets. Basic Res Cardiol 92: 191–200, 1997. 28. Kobayashi N, Kobayashi K, Kouno K, et al: Effects of intra-atrial injection of colored microspheres on systemic hemodynamics and regional blood flow in rats. Am J Physiol 266: H1910 –H1917, 1994. 29. Jasper MS, McDermott P, Gann DS, et al: Measurement of blood flow to the adrenal capsule, cortex and medulla in dogs after hemorrhage by fluorescent microspheres. J Auton Nerv Syst 30: 159 –167, 1990. 30. Ohnishi N, Liu SP, Horan P, et al: Effect of repetitive stimulation on the contractile response of rabbit urinary bladder subjected to in vitro hypoxia or in vitro ischemia followed by reoxygenation. Pharmacology 57: 139 –147, 1998.
UROLOGY 58 (2), 2001
INCREASED BLOOD FLOW AFTER CATHETERIZATION AND DRAINAGE IN THE CHRONICALLY OBSTRUCTED RABBIT URINARY BLADDER ¨ DER, BARRY A. KOGAN, JEREMY LIEB, ANNETTE SCHRO
AND
ROBERT M. LEVIN
ABSTRACT Objectives. To determine the effect of drainage on rabbit bladder blood flow after 4 weeks of partial outlet obstruction. Previous studies have shown that catheterization and drainage of the urinary bladder in control rabbits resulted in a significant nitric oxide-induced increase of blood flow to the bladder. It was also shown that 4 weeks’ partial outlet obstruction caused a significant decrease in blood flow to the bladder. Methods. Male New Zealand White rabbits underwent partial outlet obstruction by standard methods. After 4 weeks, the blood flow to the bladder muscle and mucosa was determined by a microsphere technique. Within 1 to 2 minutes after transurethral catheterization and complete drainage of the bladder, the blood flow was again determined. Unobstructed animals served as controls. Four other control animals underwent a repetitive blood flow study during 10 minutes to determine the time frame of blood flow changes after drainage. Blood flow was also measured in 2 control rabbits after transurethral catheterization without drainage and in 2 control rabbits after drainage by suprapubic puncture. To exclude the possibility that increased intravesical pressure alters the blood flow measurements, the relationship between the intravesical volume and the bladder pressure was examined in the obstructed rabbits. Results. After drainage of the bladder, the blood flow to the bladder muscle increased 4.5-fold in the decompensated obstructed group (bladder weights greater than 15 g) and 2.5-fold in the compensated animals (bladder weights less than 5 g) and control animals. Blood flow to the mucosa followed the same pattern but without reaching significance. Blood flow returned to near baseline values within 5 minutes. Catheterization without drainage did not alter the blood flow. In contrast, drainage by puncture increased the blood flow significantly. Higher intravesical volumes increased the intravesical pressure slightly, but after opening the abdominal fascia, the intravesical pressure did not change with increasing volumes. Conclusions. Although the previously shown decreased blood flow to the bladder smooth muscle may be an etiologic factor in bladder contractile dysfunction secondary to partial outlet obstruction, the bladder does have the ability to increase the blood flow after drainage. This ability could be a compensatory and possibly protective mechanism after outlet obstruction. UROLOGY 58: 295–300, 2001. © 2001, Elsevier Science Inc.
C
hronic bladder outlet obstruction (eg, resulting from benign prostatic hyperplasia) causes several changes in bladder physiology in humans and animal models.1– 6 The reason why the status of These projects were supported by grants from the Department of Veterans Affairs and National Institutes of Health grants RO1DK26508, RO1-DK53965, and RO1-DK47949. From the Department of Urology, Johannes-Gutenberg-University, Mainz, Germany; Albany Medical College, Albany College of Pharmacy, and Stratton Veterans Affairs Medical Center, Albany, New York Dr. Schro¨der is currently at the Department of Clinical Pharmacology, Lund University Hospital, Lund, Sweden. Reprint requests: Robert M. Levin, Ph.D., Albany College of Pharmacy, 106 New Scotland Avenue, Albany, NY 12208 Submitted: November 27, 2000, accepted (with revisions): March 23, 2001 © 2001, ELSEVIER SCIENCE INC. ALL RIGHTS RESERVED
decompensated bladder function is reversible in some individuals and irreversible in others remains unknown. Previous studies by other groups have shown that long-term partial obstruction of the urinary bladder leads to a significant decrease in blood flow to the bladder muscle,7–10 and this phenomenon is thought to be one reason for the damage to the bladder caused by outlet obstruction. We obtained similar results in a study of long-term outlet obstruction on blood flow to the rabbit bladder.11 The decrease in blood flow after obstruction correlated with progressive contractile failure and increased amounts of residual urine.11 Several recent studies have reported the effects of distension, bladder filling, and acute and chronic obstruction on bladder blood flow, all using different animal 0090-4295/01/$20.00 PII S0090-4295(01)01142-6 295
models and methods of blood flow measurements.12–14 In general, these studies indicate that bladder distension during filling and resulting from acute and chronic obstruction causes decreased blood flow.12–14 Previous studies in control rabbits showed that catheterization and drainage of the urinary bladder resulted in a significant nitric oxide (NO)-induced increase of blood flow to the bladder within 1 to 2 minutes after drainage.15 It was also shown that intermittent catheterization and drainage of partially obstructed bladders prevented smooth muscle dysfunction.16 We used the animals from our long-term obstruction study to look for alterations in the pattern of blood flow changes after catheterization and drainage. MATERIAL AND METHODS A standardized method was used to create moderate bladder outlet obstruction in 12 male New Zealand White rabbits (⬃3.5 kg) (Milbrook Farms, Amherst, Mass).4,6 The animals were sedated with intramuscular ketamine-xylazine (25 mg/kg ketamine, 10 mg/kg xylazine); surgical anesthesia was maintained with intravenous sodium pentobarbital (25 mg/ kg). Under sterile conditions, the urinary bladder was catheterized and drained with an 8F catheter. The bladder and urethrovesical junction were exposed through a lower abdominal midline incision. After removal of the perivesical fat, a 2-0 silk ligature was tied loosely around the vesical outlet, approximately 1 to 2 cm distal to the insertion of the ureters. The catheter was removed. The bladder was returned to the normal position and the wound closed anatomically. Intramuscular gentamicin (1 mg/kg) and buprenorphine hydrochloride (0.3 mg/kg) were given on postoperative days 1 and 2. All obstructions were performed by the same surgeon. Unobstructed animals served as the controls. Previous studies showed that sham surgery had no effect on blood flow to the bladder muscle or mucosa17; therefore, no sham operations were done in this study. Four different aspects of bladder function after outlet obstruction were examined in this study. 1. Changes in blood flow after catheterization and drainage: after 4 weeks of obstruction, the rabbits were anesthetized as described above, and blood flow was determined by a standardized fluorescent microsphere infusion technique.15,17,18 To avoid any interference with the blood flow measurements, the abdominal wall remained closed and the bladder was not manipulated during the blood flow determinations. The right carotid and femoral arteries were exposed surgically and cannulated with polyethylene 160 tubing (inside diameter 1.14 mm, outside diameter 1.57 mm). The carotid tubing was connected to a pressure transducer and advanced into the left ventricle. Sphere infusion was done by injecting 2.5 ⫻ 106 NuFlow fluorescent microspheres, 15 m in diameter (Interactive Medical Technologies, North Hollywood, Calif) using an infusion pump connected to the left ventricular catheter. Using a withdrawal pump (1.72 mL/min), a reference blood sample was withdrawn from the femoral artery, starting 15 seconds before and ending 60 seconds after sphere infusion. Then, the bladder was catheterized transurethrally with an 8F catheter, drained completely, and the volume of urine recorded. Within 60 to 90 seconds after emptying the bladder, a second blood flow determination procedure was done using a different color of fluorescent microspheres. 2. Time frame in which the blood returned to baseline after drainage: a repetitive blood flow analysis was performed in 4 296
control rabbits before and 1, 5, and 10 minutes after drainage, using four different colors of microspheres in each rabbit, as described above. The systemic arterial blood pressure was monitored throughout all microsphere procedures until the experiment ended. 3. Effects of transurethral catheterization without emptying and drainage by suprapubic puncture on bladder blood flow: to determine whether the change in blood flow was caused by the act of urethral catheterization or by the bladder emptying itself, the bladders of 2 normal control rabbits were emptied by suprapubic puncture. Two other control rabbits were catheterized transurethrally without bladder emptying. The initial and post-catheterization microsphere procedures were performed as described above. 4. Influence of bladder filling on intravesical pressure: to exclude the possibility that the baseline blood flow was artificially increased because of high intravesical pressures, we measured the bladder pressure under natural filling in an additional 14 obstructed rabbits before (by percutaneous puncture) and after surgical opening of the abdominal wall.
BLOOD FLOW ANALYSIS Under pentobarbital anesthesia, the bladder was excised, weighed, and separated into muscle and mucosa. Reference blood and detrusor samples were sent to Interactive Medical Technologies. For microsphere concentration determination, tissue samples were subjected to alkaline digestion, and the fluorescent microspheres from each sample were collected on a filter, resuspended, and quantified using flow cytometry. Microsphere concentrations of tissue samples were compared with those of reference blood samples to determine tissue perfusion; blood flow is expressed as milliliters of blood per gram tissue per minute (mL/g/min). Full-thickness strips from each bladder were used for in vitro contractility studies. The obstructed bladders were separated into compensated and decompensated groups on the basis of their contractile responses to electrical field stimulation.11
STATISTICAL ANALYSIS
All values are reported as the mean ⫾ SEM. Statistical significance was determined by Bonferroni analysis or Student’s t test, as appropriate. P ⬍0.05 was required for statistical significance.
RESULTS Partial outlet obstruction resulted in variable increases in bladder weight and in the volume of urine in the bladder at the time of blood flow measurement. The mean bladder weight in the obstructed group and the unoperated control group was 10.7 ⫾ 2.0 g (range 3.5 to 22.1) and 2.6 ⫾ 0.3 g (range 1.6 to 3.1), respectively (P ⬍0.05). To evaluate a potential relationship between the bladder weight and blood flow, the obstructed rabbits were divided into three different groups according to bladder weight. The degree of neurogenic contractile failure in the in vitro studies correlated highly with the increase in bladder weight. Therefore, we defined bladders weighing less than 5 g (3.1 ⫾ 1.4, n ⫽ 4) as compensated, bladders weighing 5.1 to 15 g (9.9 ⫾ 0.9, n ⫽ 4) as intermediate-decompensated, and bladders weighing greater than 15 g (18.0 ⫾ 1.4, n ⫽ 4) as severely UROLOGY 58 (2), 2001
FIGURE 1. Changes in blood flow to the bladder muscle after drainage. Each bar represents the mean ⫾ SEM of 5 control, 4 compensated, 4 intermediate-decompensated, and 4 decompensated obstructed bladders. Asterisks indicate P ⬍0.05 versus blood flow before drainage; plus signs indicate P ⬍0.05 versus control before drainage.
decompensated. Complete information can be found in Schro¨der et al.11 1. Changes in blood flow after catheterization and drainage: the blood flow to the smooth muscle of the controls was 0.08 ⫾ 0.01 mL/g/min, similar to that of the compensated obstructed bladders (0.08 ⫾ 0.02 mL/g/min). The blood flow to the smooth muscle of the intermediate-decompensated bladders was reduced significantly from that of the controls (0.04 ⫾ 0.01 mL/g/min). The blood flow to the severely decompensated smooth muscle was reduced even further (0.03 ⫾ 0.01 mL/g/ min) (Fig. 1). The blood flow to the mucosa of the obstructed bladders was unchanged compared with the controls (Table I). Depending on the degree of compensation, a variable increase in blood flow after catheterization was found. Similar to the controls, the obstructed bladders weighing less than 5 g had a 2.3-fold increase in blood flow after drainage of the bladder. The bladders weighing greater than 5 g had a 4.6-fold increase in blood flow after drainage (Figs. 1 and 2). The blood flow to the mucosa followed similar patterns, but without reaching a significant difference in most cases (Table I). 2. Time frame in which the blood returned to baseline after drainage—repetitive blood flow measurements: repetitive measurement of blood flow 1, 5, and 10 minutes after drainage showed that the blood flow increased within the first minute after drainage and returned to near baseline values within 5 minutes (Fig. 3). The systemic blood pressure remained stable and was not affected by the repetitive microsphere injections. 3. Effects of transurethral catheterization withUROLOGY 58 (2), 2001
out emptying and drainage by suprapubic puncture on bladder blood flow: in the four control animals in which we determined the effects of suprapubic puncture (n ⫽ 2) versus transurethral catheterization (n ⫽ 2), we found an increase in the blood flow to the bladder after drainage by suprapubic puncture (0.05 ⫾ 0.01 mL/g/min before drainage and 0.47 ⫾ 0.23 mL/g/min after drainage). Transurethral catheterization without drainage caused no changes (0.05 ⫾ 0.002 mL/g/min before and 0.07 ⫾ 0.02 mL/g/min after insertion and removal of the catheter). 4. Influence of bladder filling on intravesical pressure: in 14 obstructed rabbits (mean bladder weight 11.67 ⫾ 0.74 g), we measured the bladder pressure under natural filling before (by percutaneous puncture) and after surgical opening of the abdominal wall. The mean bladder volume was 160 ⫾ 20 mL. Intravesical pressure with the abdominal wall closed was 9.0 ⫾ 0.9 cm H2O and with the wall open was 5.5 ⫾ 0.5 cm H2O. Bladders containing large urine volumes had intravesical pressures about 5 cm H2O higher than those with low urine volumes when the abdominal wall was closed (Fig. 4). There was no influence of urine volume on bladder pressure once the abdominal wall was opened. COMMENT Drainage of the bladder causes a significant increase in blood flow to smooth muscle and mucosa in both the obstructed bladders and control animals. The increase of blood flow to the muscle caused by catheterization and drainage of the bladder was 2.5-fold in the control and compensated groups (less than 5 g bladder weight) but significantly higher (4.6-fold) in the decompensated groups. The more severe the functional decompensation was, according to the contractile response in vitro and the bladder weight, the lower were the baseline perfusion levels.11 This suggests that the increase of blood flow after drainage is a compensatory mechanism, responding to decreased baseline perfusion. The functionally severely decompensated bladders with weights greater than 15 g, although increasing their initial blood flow 4.5fold, did not reach the same blood flow values as the controls and less decompensated bladders. This could be caused by the severe damage of the bladder muscle, which would also affect the function of the blood vessels. It was shown in several other studies using Doppler flowmetry that bladder contractions (and thereby increased intravesical pressure) caused a reversible decrease of blood flow.7,14,19 –21 An increase in bladder perfusion after drainage was also demonstrated.14 A similar situation occurs in the 297
TABLE I. Blood flow to the mucosa before and after drainage Blood Flow to Mucosa (mL/g/min) Group Controls ⬍5 g 5.1–15 g ⬎15 g
Before Drainage 0.226 0.331 0.101 0.203
⫾ ⫾ ⫾ ⫾
0.041 0.101 0.101 0.019
After Drainage 0.621 1.070 1.291 1.088
⫾ ⫾ ⫾ ⫾
0.175 0.193 0.271* 0.413
Increase After Drainage (%) 222 222 327 323
⫾ ⫾ ⫾ ⫾
45 66 55 40
Data presented as the mean ⫾ SEM from 5 control, 4 compensated (⬍5 g), 4 intermediate-decompensated (5.1–15 g), and 4 decompensated (⬎15 g) obstructed bladders. * P ⬍0.05 vs. blood flow before drainage.
FIGURE 2. Changes in blood flow to the bladder muscle (expressed as percentage). Each bar represents the mean ⫾ SEM of 5 control, 4 compensated, 4 intermediate-decompensated, and 4 decompensated bladders.
FIGURE 3. Repetitive blood flow determination after drainage in 4 individual animals. The points represent the blood flow values obtained for each control rabbit at the different times. The bar represents the average period during which the second blood flow determination was done for the experiments shown in Figures 2 and 3.
heart. High pressure within the cardiac muscle during systole compresses the coronary vessels, decreasing blood flow. The blood flow then increases rapidly above baseline at the beginning of diastole and decreases slightly during diastole.22 298
FIGURE 4. Scatter plot and linear regression analysis of intravesical pressure relative to intravesical volume measured with intact and opened abdominal wall. Each pair of points represents the intravesical pressure for an individual rabbit before (black circles) and after (white circles) the opening of the abdominal wall.
Bladder contractions against an obstructed bladder neck reduced the blood flow significantly more than contractions against a nonobstructed bladder neck.7,14,20,21 When emptying against increased outflow resistance, the voiding phase is prolonged and, depending on the degree of decompensation, the poor force of detrusor contractions may further increase the workload on the bladder. Overall, an increased metabolic demand is expected. In most vascular beds, the blood flow is regulated by coordinated interactions between local myogenic tone, flow-induced dilation, and the washout of vasodilator metabolites. After metabolite-induced dilation, enhanced flow and stress on the vascular endothelium will increase the production and release of NO, which in turn further improves perfusion.23 In a recent study in control rabbits, the basal bladder smooth muscle blood flow was unaffected by treatment with the NO synthase (NOS) inhibitor, N-nitro-L-arginine methyl ester (L-NAME), and increases in blood flow after catheterization were prevented by NOS inhibition.15 This may explain the blood flow responses obtained in this report in the obstructed bladders, in which an increased UROLOGY 58 (2), 2001
metabolic demand is expected to exist. Further implicating a role for NO in the obstructed bladder, the expression of the inducible form of NOS, iNOS, was enhanced in obstructed bladders in wild-type mice.24 Mice incapable of producing iNOS failed to respond to obstruction with the expected increases in mass and functional changes observed in the wild type.25 Therefore, increased NO expression was suggested to be a mechanism by which the bladder responds to overcome or improve the increased metabolic demand during obstruction-induced ischemia. After acute urinary retention, an increase in the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH)-diaphorase staining (an indirect marker for NOS) in intramural ganglia has been described for the guineapig bladder.26 Possible neuromodulatory functions for NO at a ganglionic level in regulation of vascular tone may be considered. The repetitive determination of blood flow during the first 10 minutes after drainage revealed that the increase of blood flow after drainage is temporary and that the blood flow returns to near baseline values within minutes. This indicates that the blood flow measured before drainage can in fact be considered the baseline value and the increase in blood flow stimulated by drainage is an acute phenomenon, unrelated to the intravesical pressure. Multiple injections of different colored microspheres have proved to be a reliable method of regional blood flow determinations.27–29 Nonetheless, it is a limitation of our study that the microsphere technique, although very accurate, does not allow a continuous measurement of perfusion over a longer period. To exclude irritation of the bladder or urethra as a reason for the altered blood flow, two control animals were catheterized transurethrally without drainage. The blood flow was determined before and after the manipulation, which had no effect on the perfusion of bladder muscle or mucosa. In contrast, drainage of the bladder by suprapubic puncture caused a significant increase in blood flow to the bladder muscle and mucosa. This shows that bladder emptying is the reason for the alterations in blood flow in this study, not catheter manipulation. One might assume that the decreased blood flow before drainage is due to high intravesical pressure, caused by large amounts of residual urine in the functionally decompensated bladders, and the increase of blood flow afterward results from relief of the pressure. This explanation has already been excluded by the finding that the blood flow returns to near baseline values within 5 minutes after complete drainage. Nonetheless, the intravesical pressure is of great importance when blood flow is investigated. In the 14 additional obstructed rabbits, the intravesical pressure increased slightly at UROLOGY 58 (2), 2001
higher urine volumes, but even the highest amount of residual urine (approximately 300 mL) caused intravesical pressure values less than 15 cm H2O. Bladders containing large urine volumes had intravesical pressures about 5 cm H2O higher than those with low urine volumes when the abdominal wall was closed. This difference was not statistically significant and was less than the normal micturition pressure of about 25 cm H2O in male rabbits. After opening the abdominal wall, the pressure in the bladders containing large volumes of urine (greater than 200 mL) decreased to 5 to 6 cm H2O. This suggests that the high intravesical pressures observed in large-volume bladders are the result of compression by the abdominal contents. These findings also support the concept that the decreased blood flow observed in the decompensated bladders in this study was not caused by an increase in intravesical pressure. Nonetheless, residual urine does seem to play a role in the process of decompensation. A previous study showed that intermittent catheterization and complete drainage (every 8 hours) after outlet obstruction, beginning immediately after creation of the obstruction, significantly reduced the degree of bladder hypertrophy and the development of contractile dysfunction.30 Although the effect of partial drainage of the bladder was not investigated in this study, our results show that complete drainage has a beneficial effect compared with obstruction with untreated residual urine. It has been hypothesized that the phenomenon of the severe decrease of blood flow during micturition against partial outlet obstruction, although transient, is an important cause of damage to the bladder muscle.7,14,20,21 The increase of perfusion after the emptying of the bladder may be a compensatory mechanism. The finding that this effect is found in functionally impaired bladders supports this assumption and suggests an important therapeutic intervention. Whether partial drainage of the bladder leads to the same effect will be investigated during future studies. It is still controversial whether the obstructioninduced decrease of bladder muscle blood flow leads to contractile failure or if it is a consequence. Nonetheless, the alterations in basal blood flow after outlet obstruction, which correlate with in vitro and in vivo dysfunction, as well as changes in the response to drainage, show that bladder blood flow plays a major role in the process of decompensation. CONCLUSIONS The enhanced increase of blood flow after drainage of the bladder seems to be a compensating response to the overall decreased blood flow to the 299
bladder smooth muscle after outlet obstruction. From these results, we suggest that chronically increased amounts of residual urine, which would prevent this effect, could be the reason for the progressive damage of the smooth muscle. ACKNOWLEDGMENT. To Catherine Whitbeck and Patrick Horan for their technical assistance and to Penelope A. Longhurst for constructive criticism and support. REFERENCES 1. Kuo HC: Clinical prostate score for diagnosis of bladder outlet obstruction by prostate measurements and uroflowmetry. Urology 54: 90 –96, 1999. 2. Sonke GS, Heskes T, Verbeek AL, et al: Prediction of bladder outlet obstruction in men with lower urinary tract symptoms using artificial neural networks. J Urol 163: 300 – 305, 2000. 3. Kato K, Monson FC, Longhurst PA, et al: The functional effects of long-term outlet obstruction on the rabbit urinary bladder. J Urol 143: 600 – 606, 1990. 4. Buttyan R, Chen MW, and Levin RM: Animal models of bladder outlet obstruction and molecular insights into the basis for the development of bladder dysfunction. Eur Urol 32(suppl 1): 32–39, 1997. 5. Levin RM, Haugaard N, Mogavero L, et al: Biochemical evaluation of obstructive bladder dysfunction in men secondary to BPH: a preliminary report. Urology 53: 446 – 450, 1999. 6. Levin RM, Brading AF, Mills IW, et al: Experimental models of bladder obstruction, in Lepor H (Ed): Prostatic Disease. Philadelphia, WB Saunders, 2000, pp 169 –196. 7. Greenland JE, Hvistendahl JJ, Andersen H, et al: The effect of bladder outlet obstruction on tissue oxygen tension and blood flow in the pig bladder. BJU Int 85: 1109 –1114, 2000. 8. Lin AT, Chen MT, Yang CH, et al: Blood flow of the urinary bladder: effects of outlet obstruction and correlation with bioenergetic metabolism. Neurourol Urodyn 14: 285– 292, 1995. 9. Nielsen KK: Blood flow rate and total blood flow related to length density and total length of blood vessels in mini-pig urinary bladder after chronic outflow obstruction and after recovery from obstruction. Neurourol Urodyn 14: 177–186, 1995. 10. Tong-Long Lin A, Chen KK, Yang CH, et al: Recovery of microvascular blood perfusion and energy metabolism of the obstructed rabbit urinary bladder after relieving outlet obstruction. Eur Urol 34: 448 – 453, 1998. 11. Schro¨der A, Chichester P, Kogan BA, et al: Effect of chronic bladder outlet obstruction on the blood flow of the rabbit urinary bladder. J Urol 165: 640 – 646, 2001. 12. Finkbeiner A, and Lapides J: Effect of distension on blood flow in dogs urinary bladder. Invest Urol 12: 210 –212, 1974. 13. Nemeth CJ, Khan RM, Kirchner P, et al: Changes in canine bladder perfusion with distension. Invest Urol 15: 149 –150, 1977. 14. Azadzoi KM, Pontari M, Vlachiotis J, et al: Canine bladder blood flow and oxygenation: changes induced by filling,
300
contraction and outlet obstruction. J Urol 155: 1459 –1465, 1996. 15. Lieb J, Kogan BA, Das AK, et al: The effect of urine volume and nitric oxide on basal bladder blood flow: response to catheterization and drainage. Neurourol Urodyn 20: 115– 124, 2001. 16. Ohnishi N, Horan P, Levin SS, et al: Intermittent catheterization limits rabbit bladder dysfunction in response to partial outlet obstruction. J Urol 163: 292–295, 2000. 17. Lieb J, Chichester P, Kogan BA, et al: Rabbit urinary bladder blood flow changes during the initial stage of partial outlet obstruction. J Urol 164: 1390 –1397, 2000. 18. Shabsigh A, Hayek OH, Weiner D, et al: Acute increase in blood flow to the rat bladder subsequent to partial bladder outlet obstruction. Neurourol Urodyn 19: 195–206, 2000. 19. Brading AF, Greenland JE, Mills IW, et al: Blood supply to the bladder during filling. Scand J Urol Nephrol Suppl 201: 25–31, 1999. 20. Greenland JE, and Brading AF: Urinary bladder blood flow changes during the micturition cycle in a conscious pig model. J Urol 156: 1858 –1861, 1996. 21. Greenland JE, and Brading AF: The effect of bladder outflow obstruction on detrusor blood flow changes during the voiding cycle in conscious pigs. J Urol 165: 245–248, 2001. 22. Guyton AC: The coronary circulation and ischemic heart disease, in Guyton AC (Ed): A Textbook of Medical Physiology. Philadelphia, WB Saunders, 1986, pp 295–304. 23. Busse R, and Fleming I: Nitric oxide and the regulation of the vascular tone, in Mayer B (Ed): Handbook of Experimental Pharmacology, Vol 143. Berlin, Springer-Verlag, 2000, pp 179 –198. 24. Lemack GE, Zimmern PE, Vazquez D, et al: Altered response to partial bladder outlet obstruction in mice lacking inducible nitric oxide synthase. J Urol 163: 1981–1987, 2000. 25. Lemack GE, Burkhard F, Zimmern PE, et al: Physiologic sequelae of partial infravesical obstruction in the mouse: role of inducible nitric oxide synthase. J Urol 161: 1015–1022, 1999. 26. Zhou Y, and Ling EA: Increased NADPH-diaphorase reactivity in bladder afferent pathways following urethral obstruction in guinea pigs. J Peripher Nerv Syst 2: 333–342, 1997. 27. Walter B, Bauer R, Gaser E, et al: Validation of the multiple colored microsphere technique for regional blood flow measurements in newborn piglets. Basic Res Cardiol 92: 191–200, 1997. 28. Kobayashi N, Kobayashi K, Kouno K, et al: Effects of intra-atrial injection of colored microspheres on systemic hemodynamics and regional blood flow in rats. Am J Physiol 266: H1910 –H1917, 1994. 29. Jasper MS, McDermott P, Gann DS, et al: Measurement of blood flow to the adrenal capsule, cortex and medulla in dogs after hemorrhage by fluorescent microspheres. J Auton Nerv Syst 30: 159 –167, 1990. 30. Ohnishi N, Liu SP, Horan P, et al: Effect of repetitive stimulation on the contractile response of rabbit urinary bladder subjected to in vitro hypoxia or in vitro ischemia followed by reoxygenation. Pharmacology 57: 139 –147, 1998.
UROLOGY 58 (2), 2001