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EVIDENCE FOR INVOLVEMENT OF THE SUBCOERULEUS NUCLEUS AND NUCLEUS RAPHE MAGNUS IN URINE STORAGE AND PENILE ERECTION IN DECEREBRATE RATS
0022-5347/98/1596-2172$03.00/0
Vol. 159,2172-2176, June 1998 Printed in U.S.A.
THE JOURNAL OF UROLOGY
copyright 8 1998 by AMERICAN UROLOGICAL ASSOCIATION, bc.
EVIDENCE FOR INVOLVEMENT OF THE SUBCOERULEUS NUCLEUS AND NUCLEUS W H E MAGNUS IN URINE STORAGE AND PENILE ERECTION IN DECEREBRATE RATS KIM10 SUGAYA,* YOSHIHIDE OGAWA, TADASHI HATANO, W Z O K O Y W , TOMONORI MIYAZATO AND MASAMI ODA From the Department of Urology, Faculty of Medicine, Uniuersity of the Ryukyus, Okinawa, Japan
ABSTRACT
Purpose: Micturition and male sexual activity require the lower urinary tract to function. During the sexual act, micturition must be inhibited and urine stored in the bladder. We studied the role of the brainstem in relation to both micturitiodurine storage and penile erection in rats. Materials and Methods: Wire electrodes were placed on the dorsal nerve of the penis and microelectrodes for stimulation were introduced into the brainstem in decerebrate male rats. Electrical stimulation was used to locate optimally responding sites by monitoring the isovolumetric intravesical pressure and intracavernous pressure. Results: Electrical stimulation of the dorsal nerve of the penis, the subcoeruleus nucleus in the rostral pons, and the nucleus raphe magnus in the caudal pons increased intracavernous pressure, but inhibited rhythmic bladder contractions. Electrical stimulation of Barrington's nucleus (the pontine micturition center in the rat) in the rostral pons induced bladder contraction. Stimulation of the pontine reticular formation did not increase intracavernous pressure. Acute transection of the thoracic spinal cord eliminated rhythmic bladder contractions, but gave rise t o sporadic increments of intracavernous pressure. Conclusions: This electrophysiologicalstudy demonstrated that the subcoeruleus nucleus and nucleus raphe magnus are involved in both urine storage and penile erection, and that their physiological functions are reciprocally controlled; so that erection leads t o inhibition of micturition. KEYWORDS:urine storage, penile erection, subcoeruleus nucleus, nucleus raphe magnus, decerebrate rat
The lower urinary tract plays an important role in micturition, urine storage and sexual activity in men. When the sexual act is occurring, micturition must be inhibited and urine stored in the bladder. Therefore, we speculated that some neural mechanism may be involved in the control of both urine storage and penile erection. In some animals, certain regions of the brainstem are associated with micturition and urine storage.'-' Electrical stimulation of the nucleus locus coeruleus alpha has been reported to elicit micturition in cats' and dogs.' Stimulation of the laterodorsal tegmental nucleus, periaqueductal gray or lateral parabrachial nucleus evokes bladder contraction in r a t s 3 In addition, stimulation of the nucleus locus subcoeruleus? nucleus reticularis pontis ~ r a l i s or ~ .nucleus ~ raphe magnus7 in cats, and the nucleus locus subcoeruleus in dogs' has been reported to inhibit bladder contraction. We employed electrical stimulation in decerebrate rats to locate several brainstem regions controlling both micturitionlurine storage and penile erection. MATERIALS AND METHODS
Twenty male Sprague-Dawley rats (250 to 390 gm.) were intubated and anesthetized with halothane inhalation (2%). The bilateral internal cervical arteries were ligated and the bilateral ureters were divided with the distal ends ligated. The bladder neck was also closed and a catheter (PE-50) was placed into the bladder from the dome to monitor the isovolu-
metric intravesical pressure. The penile skin was removed and 2 wire electrodes (diameter 50 pm.) for electrical stimulation were placed directly onto the bilateral dorsal nerves of the penis. A needle (25 G) was inserted into the corpus cavernosum of the penis to record the intracavernous pressure. Following closure of the abdomen and the neck wound, the animal was fixed on a stereotaxic instrument. Anesthesia was discontinued following decerebration a t the precollicular-postmammillary level. The bladder was filled with physiological saline t o either under or over the threshold volume inducing isovolumetric rhythmic bladder contractions. The intracavernous pressure was measured with continuous infusion of physiological saline (2.4 ml./hr.) via the inserted needle to prevent it clogging. As hypotension due to hemorrhage after decerebration might cause penile erection to fail despite stimulation of the dorsal nerve of the penis,' the nerve was electrically stimulated for a preliminary test prior t o initiation of brainstem stimulation. After confirmation of an increase in intracavernous pressure with electrical stimulation (10 to 20 V, 20 pulsek, pulse duration 0.2 ms, train duration 20 to 40 s) of the dorsal nerve of the penis, a stimulating microelectrode (10 t o 50 50 pulsek, pulse duration 0.2 ms, train duration 5 to 40 s) was introduced stereotaxically into the brainstem (Bregma -8.5 to 12.0 mm., LR 0 to 2.0 mm., H 4.0 to 0.5 mm.), and the sites giving an optimal response to stimulation were located by monitoring both intracavernous and intravesical pressure. The stimulating microelectrode was a monopolar glass electrode filled with Wood's metal (tip diameter 10 to 20 pm.). At the completion of brainstem stimulation, the site was marked by electrocoagulation (DC current, 50 p,A for 30 s). Finally, the first thoracic spinal cord was transected under transient
Accepted for publication December 9, 1997. * Requests for reprints: Department of Urology, Faculty of Medicine, University of the Ryukyus, 207 Uehara, Nishihara, Okinawa 903-0215, Japan. Supported by Narishige Neuroscience Research Foundation. 2172
a,
BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION halothane anesthesia t o examine the effect of the whole brainstem on penile erection. Under deep halothane anesthesia, the brainstem was removed, fmed in 10%formalin for ten days, cut into 50-pm. sections, and stained with neutral red. The location of the lesion was identified with reference to representative sections of the stereotaxic atlas.''
2173
mH20
801
RESULTS
Filling the bladder with physiological saline caused isovolumetric rhythmic bladder contractions (amplitude 35 to 100 cm. water) in all decerebrate rats. With these bladder ICP contractions, the intracavernous pressure either increased slightly (n = 12,515 cm. water, fig. 1) or did not change (n = 8). Electrical stimulation of the dorsal nerve of the penis increased the intracavernous pressure (amplitude 30 to 80 cm. water) after a delay of 7 to 12 seconds and inhibited rhythmic bladder contractions in 10 rats (fig. 2). The inhibition of bladder contraction persisted for about 1 to 2 minutes after electrical stimulation was discontinued. Electrical stimulation of the dorsal nerve of the penis did not increase intracavernous pressure in the remaining 10 rats, but inhibited bladder contraction for 1 to 2 minutes. We were able to locate brainstem sites that optimally elic- cmH2O @-PMC ited a n increase of intracavernous or intravesical pressure with the bladder filled to under the threshold volume evoking rhythmic contractions. During stimulation from the caudal I midbrain (Bregma - 8.5 mm.) to the rostral medulla (Bregma IVp - 12.0 mm.), we identified one site in the rostral pontine dorsolateral area that evoked bladder contraction, as well as two sites that increased intracavernous pressure in the ros- m H 2 O tral pontine lateral area and caudal pontine (or rostral medullary) ventral raphe area in the 10 rats in which electrical stimulation of the dorsal nerve of the penis caused an increase of intracavernous pressure. In the remaining 10 rats, 0 in which electrical stimulation of the dorsal nerve of the lmin penis failed to increase the intracavernous pressure, no KP brainstem sites influencing the intracavernous pressure or FIG.3. Bladder contraction and small synchronous increase of intravesical pressure were identified. Electrical stimulation intracavernouspressure induced by electrical stimulation of pontine of the pontomedullary reticular formation did not increase micturition center (ES-PMC)comes onding to Bamngton's nucleus in decerebrate rat. With bladder tiyled to below threshold volume the intracavernous pressure in any of the rats. Electrical stimulation at the presumed site of Barrington's inducing rhythmic contractions, electrical Stimulation induced bladder contraction (60 em. water) after delay of 2 seconds. Intracavernnucleus in the rostral pontine dorsolateral area induced blad- ous pressure increased slightly (12 cm. water) along with bladder der contraction (40 t o 75 cm. water) after a delay of a few contraction. seconds (fig. 3). During the contraction, intracavernous pressure increased slightly (515 cm. water). Electrical stimula- after a delay of a few seconds (fig. 4), but there was no change tion did not induce bladder contraction upon probing the area of intravesical pressure. Electrical stimulation did not induce over 0.25 mm. in any direction from the optimal site for an increase of intracavernous pressure upon probing the area inducing contraction. Electrical stimulation of sites ventrolateral and caudal to Barrington's nucleus in the rostral pontine lateral area inE-SLC creased the intracavernous pressure (60 to 80 cm. water)
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FIG. 1. Rhythmic bladder contractions and synchronous increase of intracavernous pressure in decerebraterat. Small rhythmic waves (amplitude:5 to 7 cm. water, duration: 52 seconds) of intracavernous
pressure were mechanically induced by infusion pump, because pressure was measured with continuous infusion of physiolo 'cal saline into penis. When bladder was filled with physiological sxne. rhythmic contractions (amplitude: 62 cm. water) developed and mtracavernous pressure increased (amplitude: 12 cm. water) with these Contractions.
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FIG. 4. Increase of intracavernouspressure with electrical stimulation of subcoeruleus nucleus (ES-SLC) in decerebrate rat. With bladder filled to below threshold volume inducing rhythmic contractions, electrical stimulation of nucleus induced increase of intracavernous pressure (68 cm. water) after delay of 2 seconds. Despite increase of intracavernous pressure, intravesical pressure did not change. Intracavernous pressure decreased on completion of electrical stimulation.
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BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION
(0.25to 0.5 mm.) in any direction from the optimal site. Electrical stimulation of the midline extending from the caudal pons to the rostral medulla also increased the intracavernous pressure after a delay of a few seconds. The midline (raphe) sites increasing intracavernous pressure spread over 2 to 3 mm. rostrocaudally and dorsoventrally, but not laterally. Among these raphe sites, the optimally responsive area included the nucleus raphe magnus in the caudal pons. Electrical stimulation of the optimal site in the nucleus induced an increase of intracavernous pressure to 65 to 80 cm. water. When the bladder was filled above the threshold volume evoking rhythmic contractions, electrical stimulation of the rostral pontine lateral region or caudal pontine ventral raphe region increased the intracavernous pressure and inhibited rhythmic bladder contractions (fig. 5). Acute transection of the thoracic spinal cord at T1 (n = 5) under transient halothane anesthesia abolished rhythmic bladder contractions, even when the bladder was filled, confirming areflexia of the bladder, but sporadic undefined intracavernous pressure waves (525 cm. water) were documented (fig. 6).under these conditions, electrical stimulation of the dorsal nerve of the penis also increased intracavernous pressure (35 to 45 cm. water). The optimal sites for inducing bladder contraction or for increasing the intracavernous pressure were electrically coagulated: the electrical lesions were located in Barrington's nucleus or its vicinity (n = lo), the subcoeruleus nucleus or its vicinity in the rostral pons (n = 101, and the ventral part of the nucleus raphe magnus in the caudal pons (n = 10) (fig. 7). DISCUSSION
As transection of the spinal cord induced an increase of intracavernous pressure, the brainstem seemed to have an overall inhibitory effect on penile erection. Electrical stimulation of the subcoeruleus nucleus in the rostral pons or the nucleus raphe magnus in the caudal pons inhibited bladder contraction and increased the intracavernous pressure. Therefore, the rostral pontine lateral region and the caudal pontine ventral raphe region may influence both urine storage and penile erection. Blood pressure increases with bladder contraction in anesthetized rats." It has been reported that the blood pressure influences the increase of intracavernous pressure when electrical stimulation is applied to the dorsal nerve of the penkg Thus, hypotension might be related to the failure of electrical stimulation of the dorsal nerve or the brainstem to change the intracavernous pressure in 10 of 20 rats. The slight increase of intracavernous pressure synchronized with rhythmic bladder contractions or concomitant with bladder contraction induced by electrical stimulation of Barrington's
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FIG.5. Electrical stimulation of nucleus raphe magnus (ES-RM) inhibited rhythmic bladder contractions and increased intracavernous pressure in decerebrate rat. With bladder filled to above threshold volume, electrical stimulation of nucleus induced increase of intracavernous pressure (70 cm. water) after delay of 2 seconds. Intracavernous pressure decreased again a t 37 seconds after beginning of stimulation. Rhythmic bladder contractions (interval: 4.2 to 4.5 minutes) were inhibited by electrical stimulation of nucleus, and new bladder contraction developed a t 4.5 minutes after completion of stimulation,
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FIG.6. Changes of intravesical and intracavernous pressure with electrical stimulation of dorsal nerve of penis (ES-DNP) before and after transection of thoracic spinal cord in decerebrate rat. A, before cord transection, electrical stimulation of dorsal nerve of penis increased intracavernous pressure (64 to 67 cm. water) after delay of 7 seconds with bladder filled to below threshold volume. Additional infusion of physiological saline into bladder induced rhythmic contractions. Transection of thoracic spinal cord a t T1 was performed under transient halothane anesthesia. B, ten minutes after cord transection, rhythmic bladder contractions did not develop, but sporadic undefined intracavernous pressure waves (525 cm. water) were recorded. Electrical stimulation of dorsal nerve of penis also increased intracavernous pressure (38 to 45 cm. water) in this spinal rat.
nucleus (the pontine micturition center in the rat". 13)that was noted in this study might have been associated with a transient increase in blood pressure. Barrington's nucleus is located between the laterodorsal tegmental nucleus and the locus coeruleus according to the rat brain maps in the atlas of Swanson," and it has been included in the laterodorsal tegmental nucleus14 which was previously called the pontine micturition center of the rat. Electrical stimulation of the dorsal nerve of the penis inhibited rhythmic bladder contractions with or without an increase of the intracavernous pressure, suggesting that stimulation of afferent fibers of the dorsal nerve inhibited the output of the vesical branch of the pelvic nerves at the spinal cord.g Transection of the thoracic spinal cord eliminated bladder contraction but induced sporadic undefined intracavernous pressure waves, suggesting that the penile erection center is located in the lumbosacral spinal cord from which the pelvic and pudendal nerves originate,15 and that the brainstem projects a tonic descending inhibitory influence on the spinal erection reflex. Regarding descending inhibition from the brainstem, the rostral pole of the paragigantocellular reticular nucleus in the rostral medulla has been reported to project to the lumbosacral spinal cord and inhibit the coitus reflex in rats.16 Electrical stimulation of the region extending from the ventrolateral area to the pontine micturition center inhibits bladder contraction in rats,3 cats4 and dogs: and facilitates external urethral sphincter muscle activity in cats4 and dogs8 The subcoeruleus nucleus is included in this area and its role in affecting penile erection has not previously been identified. Our study showed that electrical stimulation of the subcoeruleus nucleus or its vicinity not only inhibited bladder contraction but also increased intracavernous pressure
BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION
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this situation, the external urethral sphincter muscle might be activated to promote urine storage. When pseudorabies virus, which infects neurons transsynaptically, was injected into the bladder,'". l 3 ~ r e t h r a , 'or ~ penis" i n rats, many virus-labeled neurons were found in Barrington's nucleus and the nucleus raphe magnus, and there were also some labeled neurons in the subcoeruleus nucleus. The nucleus raphe magnus also receives neuronal terminals from t h e lateral pontine area, including the nucleus locus subcoeruleus, i n cats."' Therefore, neurons in the rostra1 pontine lateral region and the caudal pontine ventral raphe region may project to the lumbosacral spinal cord, which innervates the bladder, urethra, and penis via t h e pelvic and pudendal nerves, and may be part of a n integrated neural mechanism. In decerebrate cats, electrical stimulation or injection of a cholinergic agent into the nucleus reticularis pontis oralis located ventromedial t o the pontine micturition center inhibits bladder Injection of a cholinergic agent into this nucleus also inhibits muscle activity in the extremities and inhibits the external urethral sphincter in decerebrate cats." In the present study, electrical stimulation of the reticular formation did not change the intracavernous pressure. Therefore, not all of the brainstem sites t h a t inhibit bladder contraction have a n effect on penile erection. However, 41 of 45 men with multiple sclerosis and impotence were reported to have bladder hyperreflexia on cystometry,'l" suggesting t h a t the central neural mechanisms for urine storage and penile erection may partially overlap. CONCLUSIONS
The overall effect of the brainstem is to inhibit penile erection. Both t h e subcoeruleus nucleus and the nucleus raphe magnus influence urine storage and penile erection. These regions have integrated neural networks. In addition, the pontine reticular formation influences urine storage but not affect penile erection. Therefore, t h e neural mechanisms for urine storage and penile erection may partially overlap in the brainstem. REFERENCES
FIG. 7. Location of lesions in pons are depicted sites marked as (0) evoked bladder contractions and sites marked as (0)increased intracavernous pressure upon electrical Stimulation in 10 decerebrate rats. Three transverse planes (Bregma -9.5 mm., -9.8 mm. and -10.6 mm.) were modified from Swanson's rat brain maps." B, Barrington's nucleus; GRN, gigantocellular reticular nucleus; ICP, inferior cerebellar peduncle; LC, locus coeruleus; LDT, laterodorsal tegmental nucleus; MEV, mesencephalic nucleus of trigeminal; m, medial vestibular nucleus; PB, parabrachial nucleus; PRNc, pontine reticular nucleus, caudal part; RM, nucleus raphe magnus; RPA, nucleus raphe pallidus; SCP, superior cerebellar peduncle; SLC, subcoeruleus nucleus; SLD, sublaterodorsal nucleus; V4, fourth ventricle; V, motor nucleus of the trigeminal nerve; VII, facial nucleus.
The raphe nuclei which project serotonergic nerve terminals to t h e brain and spinal cord have been reported to be associated with urine s tor age,^ penile erection,15 and muscle activity of the e ~ t r e m i t i e s . ' Electrical ~ stimulation of the nucleus raphe magnus inhibits the micturition reflex i n cats.7 Median and dorsal raphe lesions in the midbrain inhibit penile erection induced by fenfluramine, a serotonergic agonist, i n rats.' Penile erection is elicited by peripheral injection of m-chlorophenyl piperazine, a 5-HT1C agonist, and suppressed by 5-HT1A a n d o r 5-HT2 agonists." Electrical stimulation of the nucleus raphe magnus increases muscle activity of the extremities in decerebrate cats.16 We found t h a t electrical stimulation of this nucleus inhibited bladder contraction and increased t h e intracavernous pressure. In
1. Sugaya, K., Matsuyama, K., Takakusaki, K. and Mori, S.: Electrical and chemical stimulations of the pontine micturition center. Neurosci. Lett., 80: 197, 1987. 2. Nishizawa, 0.. Sugaya, K., Noto, H., Harada, T. and Tsuchida, S.: Pontine micturition center in the dog. J. Urol., 1 4 0 872, 1988. 3. Kruse, M. N., Noto, H., Roppolo, J. R. and de Groat, W. C.:
Pontine control of the urinary bladder and external urethral sphincter in the rat. Brain Res., 532 182, 1990. 4. Matsuzaki, A., Nishizawa, O., Sugaya, K., Noto, H. and Tsuchida, S.: Chemical stimulation of the nucleus locus subcoeruleus functioning for urine storage in the decerebrate cat. Neurourol. Urodynam., 8: 385, 1989. 5. Sugaya, K., Mori, S., Nishizawa, O., Noto, H. and Tsuchida, S.: Chemical stimulation of the pontine micturition center and the nucleus reticularis pontis oralis. Neurourol. Urodynam., 6 143, 1987. 6. Kimura, Y., Ukai,Y., Kimura, K., Sugaya, K. and Nishizawa, 0.:
Inhibitory influence from the nucleus reticularis pontis oralis on the micturition reflex induced by electrical stimulation of the pontine micturition center in cats. Neurosci. Lett., 195: 214, 1995. 7. McMahon, S. B. and Spillane, K.: Brain stem influences on the
parasympathetic supply to the urinary bladder of the cat. Brain Res., 234: 237, 1982. 8. Nishizawa, 0.. Sugaya, K., Noto, H., Harada, T. and Tsuchida, S.: Pontine urine storage center in the doe. Tohoku J. EXD. Med., 1 5 3 77, 1987. 9. Pescatori. E. S.. Calabro, A,, Artibani, W., Papano, F., Triban, C. and Italiano, G.: Electrical stimulation of the dorsal nerve of the penis evokes reflex tonic erections of the penile body and I
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BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION
reflex ejaculatory responses in the spinal rat. J . Urol., 149 627, 1993. 10. Swanson, L.W.:Brain maps: Structure of the rat brain. Amsterdam: Elsevier, 1992. 11. Cheng, C-L., Ma, C-P. and de Groat, W. C.: Effects of capsaicin on micturition and associated reflexes in rats. Am. J . Physiol., 265: R132, 1993. 12. Sugaya, K., Roppolo, J. R., Yoshimura, N., Card, J. P. and de Groat, W. C.: The central neural pathways involved in micturition in the neonatal rat as revealed by the injection of pseudorabies virus into the urinary bladder. Neurosci. Lett., 223: 197,1997. 13. Nadelhaft, I., Vera, P. L., Card, J. P. and Miselis, R. R.: Central nervous system neurons labelled following the injection of pseudorabies virus into the rat urinary bladder. Neurosci. Lett., 143: 271, 1992. 14. Satoh, K., Shimizu, N., Tohyama, M. and Maeda, T.: Localization ofthe micturition reflex center at dorsolateral pontine tegmentum of the rat. Neurosci. Lett., 8: 27, 1978. 15. Maeda, N., Matsuoka, N. and Yamaguchi, I.: Possible involvement of the septo-hippocampal cholinergic and raphehippocampal serotonergic activations in the penile erection induced by fenfluramine in rats. Brain Res., 652 181, 1994. 16. Marson, L. and McKenna, K. E.: The identification of a brain-
stem site controlling spinal sexual reflexes in male rats. Brain Res., 5 1 5 303, 1990. 17. Mori, S., Matsuyama, K., Kohyama, J., Kobayashi, Y. and Takakusaki, K.: Neuronal constituents of postural and locomotor control systems and their interactions in cats. Brain Dev. Suppl., 1 4 S109,1992. 18. Berendsen, H. H.G., Jenck, F. and Broekkamp, C. L. E.: Involvement of 5-HT 1C receptors in drug-induced penile erection in rats. Psychopharmacology, 101: 57, 1990. 19. Vizzard, M. A,, Erickson, V. L., Card, J. P., Roppolo, J. R. and de Groat, W. C.: Transneuronal labeling of neurons in the adult rat brain and spinal cord after injection of pseudorabies virus into the urethra. J. Comp. Neurol., 3 5 5 629,1995. 20. Marson, L., Platt, K. B. and McKenna, K. E.: Central nervous system innervation of the penis as revealed by the transneuronal transport of pseudorabies virus. Neurosci., 5 5 263,1993. 21. Holstege, G.: Anatomical evidence for a strong ventral parabrachial projection to nucleus raphe magnus and adjacent tegmental field. Brain Res., 447: 154, 1988. 22. Betts, C. D.,Jones, S. J., Fowler, C. G. and Fowler, C. J.:Erectile dysfunction in multiple sclerosis. Associated neurological and neurophysiological deficits, and treatment of the condition. Brain, 117: 1303,1994.
Vol. 159,2172-2176, June 1998 Printed in U.S.A.
THE JOURNAL OF UROLOGY
copyright 8 1998 by AMERICAN UROLOGICAL ASSOCIATION, bc.
EVIDENCE FOR INVOLVEMENT OF THE SUBCOERULEUS NUCLEUS AND NUCLEUS W H E MAGNUS IN URINE STORAGE AND PENILE ERECTION IN DECEREBRATE RATS KIM10 SUGAYA,* YOSHIHIDE OGAWA, TADASHI HATANO, W Z O K O Y W , TOMONORI MIYAZATO AND MASAMI ODA From the Department of Urology, Faculty of Medicine, Uniuersity of the Ryukyus, Okinawa, Japan
ABSTRACT
Purpose: Micturition and male sexual activity require the lower urinary tract to function. During the sexual act, micturition must be inhibited and urine stored in the bladder. We studied the role of the brainstem in relation to both micturitiodurine storage and penile erection in rats. Materials and Methods: Wire electrodes were placed on the dorsal nerve of the penis and microelectrodes for stimulation were introduced into the brainstem in decerebrate male rats. Electrical stimulation was used to locate optimally responding sites by monitoring the isovolumetric intravesical pressure and intracavernous pressure. Results: Electrical stimulation of the dorsal nerve of the penis, the subcoeruleus nucleus in the rostral pons, and the nucleus raphe magnus in the caudal pons increased intracavernous pressure, but inhibited rhythmic bladder contractions. Electrical stimulation of Barrington's nucleus (the pontine micturition center in the rat) in the rostral pons induced bladder contraction. Stimulation of the pontine reticular formation did not increase intracavernous pressure. Acute transection of the thoracic spinal cord eliminated rhythmic bladder contractions, but gave rise t o sporadic increments of intracavernous pressure. Conclusions: This electrophysiologicalstudy demonstrated that the subcoeruleus nucleus and nucleus raphe magnus are involved in both urine storage and penile erection, and that their physiological functions are reciprocally controlled; so that erection leads t o inhibition of micturition. KEYWORDS:urine storage, penile erection, subcoeruleus nucleus, nucleus raphe magnus, decerebrate rat
The lower urinary tract plays an important role in micturition, urine storage and sexual activity in men. When the sexual act is occurring, micturition must be inhibited and urine stored in the bladder. Therefore, we speculated that some neural mechanism may be involved in the control of both urine storage and penile erection. In some animals, certain regions of the brainstem are associated with micturition and urine storage.'-' Electrical stimulation of the nucleus locus coeruleus alpha has been reported to elicit micturition in cats' and dogs.' Stimulation of the laterodorsal tegmental nucleus, periaqueductal gray or lateral parabrachial nucleus evokes bladder contraction in r a t s 3 In addition, stimulation of the nucleus locus subcoeruleus? nucleus reticularis pontis ~ r a l i s or ~ .nucleus ~ raphe magnus7 in cats, and the nucleus locus subcoeruleus in dogs' has been reported to inhibit bladder contraction. We employed electrical stimulation in decerebrate rats to locate several brainstem regions controlling both micturitionlurine storage and penile erection. MATERIALS AND METHODS
Twenty male Sprague-Dawley rats (250 to 390 gm.) were intubated and anesthetized with halothane inhalation (2%). The bilateral internal cervical arteries were ligated and the bilateral ureters were divided with the distal ends ligated. The bladder neck was also closed and a catheter (PE-50) was placed into the bladder from the dome to monitor the isovolu-
metric intravesical pressure. The penile skin was removed and 2 wire electrodes (diameter 50 pm.) for electrical stimulation were placed directly onto the bilateral dorsal nerves of the penis. A needle (25 G) was inserted into the corpus cavernosum of the penis to record the intracavernous pressure. Following closure of the abdomen and the neck wound, the animal was fixed on a stereotaxic instrument. Anesthesia was discontinued following decerebration a t the precollicular-postmammillary level. The bladder was filled with physiological saline t o either under or over the threshold volume inducing isovolumetric rhythmic bladder contractions. The intracavernous pressure was measured with continuous infusion of physiological saline (2.4 ml./hr.) via the inserted needle to prevent it clogging. As hypotension due to hemorrhage after decerebration might cause penile erection to fail despite stimulation of the dorsal nerve of the penis,' the nerve was electrically stimulated for a preliminary test prior t o initiation of brainstem stimulation. After confirmation of an increase in intracavernous pressure with electrical stimulation (10 to 20 V, 20 pulsek, pulse duration 0.2 ms, train duration 20 to 40 s) of the dorsal nerve of the penis, a stimulating microelectrode (10 t o 50 50 pulsek, pulse duration 0.2 ms, train duration 5 to 40 s) was introduced stereotaxically into the brainstem (Bregma -8.5 to 12.0 mm., LR 0 to 2.0 mm., H 4.0 to 0.5 mm.), and the sites giving an optimal response to stimulation were located by monitoring both intracavernous and intravesical pressure. The stimulating microelectrode was a monopolar glass electrode filled with Wood's metal (tip diameter 10 to 20 pm.). At the completion of brainstem stimulation, the site was marked by electrocoagulation (DC current, 50 p,A for 30 s). Finally, the first thoracic spinal cord was transected under transient
Accepted for publication December 9, 1997. * Requests for reprints: Department of Urology, Faculty of Medicine, University of the Ryukyus, 207 Uehara, Nishihara, Okinawa 903-0215, Japan. Supported by Narishige Neuroscience Research Foundation. 2172
a,
BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION halothane anesthesia t o examine the effect of the whole brainstem on penile erection. Under deep halothane anesthesia, the brainstem was removed, fmed in 10%formalin for ten days, cut into 50-pm. sections, and stained with neutral red. The location of the lesion was identified with reference to representative sections of the stereotaxic atlas.''
2173
mH20
801
RESULTS
Filling the bladder with physiological saline caused isovolumetric rhythmic bladder contractions (amplitude 35 to 100 cm. water) in all decerebrate rats. With these bladder ICP contractions, the intracavernous pressure either increased slightly (n = 12,515 cm. water, fig. 1) or did not change (n = 8). Electrical stimulation of the dorsal nerve of the penis increased the intracavernous pressure (amplitude 30 to 80 cm. water) after a delay of 7 to 12 seconds and inhibited rhythmic bladder contractions in 10 rats (fig. 2). The inhibition of bladder contraction persisted for about 1 to 2 minutes after electrical stimulation was discontinued. Electrical stimulation of the dorsal nerve of the penis did not increase intracavernous pressure in the remaining 10 rats, but inhibited bladder contraction for 1 to 2 minutes. We were able to locate brainstem sites that optimally elic- cmH2O @-PMC ited a n increase of intracavernous or intravesical pressure with the bladder filled to under the threshold volume evoking rhythmic contractions. During stimulation from the caudal I midbrain (Bregma - 8.5 mm.) to the rostral medulla (Bregma IVp - 12.0 mm.), we identified one site in the rostral pontine dorsolateral area that evoked bladder contraction, as well as two sites that increased intracavernous pressure in the ros- m H 2 O tral pontine lateral area and caudal pontine (or rostral medullary) ventral raphe area in the 10 rats in which electrical stimulation of the dorsal nerve of the penis caused an increase of intracavernous pressure. In the remaining 10 rats, 0 in which electrical stimulation of the dorsal nerve of the lmin penis failed to increase the intracavernous pressure, no KP brainstem sites influencing the intracavernous pressure or FIG.3. Bladder contraction and small synchronous increase of intravesical pressure were identified. Electrical stimulation intracavernouspressure induced by electrical stimulation of pontine of the pontomedullary reticular formation did not increase micturition center (ES-PMC)comes onding to Bamngton's nucleus in decerebrate rat. With bladder tiyled to below threshold volume the intracavernous pressure in any of the rats. Electrical stimulation at the presumed site of Barrington's inducing rhythmic contractions, electrical Stimulation induced bladder contraction (60 em. water) after delay of 2 seconds. Intracavernnucleus in the rostral pontine dorsolateral area induced blad- ous pressure increased slightly (12 cm. water) along with bladder der contraction (40 t o 75 cm. water) after a delay of a few contraction. seconds (fig. 3). During the contraction, intracavernous pressure increased slightly (515 cm. water). Electrical stimula- after a delay of a few seconds (fig. 4), but there was no change tion did not induce bladder contraction upon probing the area of intravesical pressure. Electrical stimulation did not induce over 0.25 mm. in any direction from the optimal site for an increase of intracavernous pressure upon probing the area inducing contraction. Electrical stimulation of sites ventrolateral and caudal to Barrington's nucleus in the rostral pontine lateral area inE-SLC creased the intracavernous pressure (60 to 80 cm. water)
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FIG. 1. Rhythmic bladder contractions and synchronous increase of intracavernous pressure in decerebraterat. Small rhythmic waves (amplitude:5 to 7 cm. water, duration: 52 seconds) of intracavernous
pressure were mechanically induced by infusion pump, because pressure was measured with continuous infusion of physiolo 'cal saline into penis. When bladder was filled with physiological sxne. rhythmic contractions (amplitude: 62 cm. water) developed and mtracavernous pressure increased (amplitude: 12 cm. water) with these Contractions.
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FIG. 4. Increase of intracavernouspressure with electrical stimulation of subcoeruleus nucleus (ES-SLC) in decerebrate rat. With bladder filled to below threshold volume inducing rhythmic contractions, electrical stimulation of nucleus induced increase of intracavernous pressure (68 cm. water) after delay of 2 seconds. Despite increase of intracavernous pressure, intravesical pressure did not change. Intracavernous pressure decreased on completion of electrical stimulation.
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BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION
(0.25to 0.5 mm.) in any direction from the optimal site. Electrical stimulation of the midline extending from the caudal pons to the rostral medulla also increased the intracavernous pressure after a delay of a few seconds. The midline (raphe) sites increasing intracavernous pressure spread over 2 to 3 mm. rostrocaudally and dorsoventrally, but not laterally. Among these raphe sites, the optimally responsive area included the nucleus raphe magnus in the caudal pons. Electrical stimulation of the optimal site in the nucleus induced an increase of intracavernous pressure to 65 to 80 cm. water. When the bladder was filled above the threshold volume evoking rhythmic contractions, electrical stimulation of the rostral pontine lateral region or caudal pontine ventral raphe region increased the intracavernous pressure and inhibited rhythmic bladder contractions (fig. 5). Acute transection of the thoracic spinal cord at T1 (n = 5) under transient halothane anesthesia abolished rhythmic bladder contractions, even when the bladder was filled, confirming areflexia of the bladder, but sporadic undefined intracavernous pressure waves (525 cm. water) were documented (fig. 6).under these conditions, electrical stimulation of the dorsal nerve of the penis also increased intracavernous pressure (35 to 45 cm. water). The optimal sites for inducing bladder contraction or for increasing the intracavernous pressure were electrically coagulated: the electrical lesions were located in Barrington's nucleus or its vicinity (n = lo), the subcoeruleus nucleus or its vicinity in the rostral pons (n = 101, and the ventral part of the nucleus raphe magnus in the caudal pons (n = 10) (fig. 7). DISCUSSION
As transection of the spinal cord induced an increase of intracavernous pressure, the brainstem seemed to have an overall inhibitory effect on penile erection. Electrical stimulation of the subcoeruleus nucleus in the rostral pons or the nucleus raphe magnus in the caudal pons inhibited bladder contraction and increased the intracavernous pressure. Therefore, the rostral pontine lateral region and the caudal pontine ventral raphe region may influence both urine storage and penile erection. Blood pressure increases with bladder contraction in anesthetized rats." It has been reported that the blood pressure influences the increase of intracavernous pressure when electrical stimulation is applied to the dorsal nerve of the penkg Thus, hypotension might be related to the failure of electrical stimulation of the dorsal nerve or the brainstem to change the intracavernous pressure in 10 of 20 rats. The slight increase of intracavernous pressure synchronized with rhythmic bladder contractions or concomitant with bladder contraction induced by electrical stimulation of Barrington's
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FIG.5. Electrical stimulation of nucleus raphe magnus (ES-RM) inhibited rhythmic bladder contractions and increased intracavernous pressure in decerebrate rat. With bladder filled to above threshold volume, electrical stimulation of nucleus induced increase of intracavernous pressure (70 cm. water) after delay of 2 seconds. Intracavernous pressure decreased again a t 37 seconds after beginning of stimulation. Rhythmic bladder contractions (interval: 4.2 to 4.5 minutes) were inhibited by electrical stimulation of nucleus, and new bladder contraction developed a t 4.5 minutes after completion of stimulation,
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FIG.6. Changes of intravesical and intracavernous pressure with electrical stimulation of dorsal nerve of penis (ES-DNP) before and after transection of thoracic spinal cord in decerebrate rat. A, before cord transection, electrical stimulation of dorsal nerve of penis increased intracavernous pressure (64 to 67 cm. water) after delay of 7 seconds with bladder filled to below threshold volume. Additional infusion of physiological saline into bladder induced rhythmic contractions. Transection of thoracic spinal cord a t T1 was performed under transient halothane anesthesia. B, ten minutes after cord transection, rhythmic bladder contractions did not develop, but sporadic undefined intracavernous pressure waves (525 cm. water) were recorded. Electrical stimulation of dorsal nerve of penis also increased intracavernous pressure (38 to 45 cm. water) in this spinal rat.
nucleus (the pontine micturition center in the rat". 13)that was noted in this study might have been associated with a transient increase in blood pressure. Barrington's nucleus is located between the laterodorsal tegmental nucleus and the locus coeruleus according to the rat brain maps in the atlas of Swanson," and it has been included in the laterodorsal tegmental nucleus14 which was previously called the pontine micturition center of the rat. Electrical stimulation of the dorsal nerve of the penis inhibited rhythmic bladder contractions with or without an increase of the intracavernous pressure, suggesting that stimulation of afferent fibers of the dorsal nerve inhibited the output of the vesical branch of the pelvic nerves at the spinal cord.g Transection of the thoracic spinal cord eliminated bladder contraction but induced sporadic undefined intracavernous pressure waves, suggesting that the penile erection center is located in the lumbosacral spinal cord from which the pelvic and pudendal nerves originate,15 and that the brainstem projects a tonic descending inhibitory influence on the spinal erection reflex. Regarding descending inhibition from the brainstem, the rostral pole of the paragigantocellular reticular nucleus in the rostral medulla has been reported to project to the lumbosacral spinal cord and inhibit the coitus reflex in rats.16 Electrical stimulation of the region extending from the ventrolateral area to the pontine micturition center inhibits bladder contraction in rats,3 cats4 and dogs: and facilitates external urethral sphincter muscle activity in cats4 and dogs8 The subcoeruleus nucleus is included in this area and its role in affecting penile erection has not previously been identified. Our study showed that electrical stimulation of the subcoeruleus nucleus or its vicinity not only inhibited bladder contraction but also increased intracavernous pressure
BRAINSTEM NEURAL MECHANISMS OF URINE STORAGE AND PENILE ERECTION
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this situation, the external urethral sphincter muscle might be activated to promote urine storage. When pseudorabies virus, which infects neurons transsynaptically, was injected into the bladder,'". l 3 ~ r e t h r a , 'or ~ penis" i n rats, many virus-labeled neurons were found in Barrington's nucleus and the nucleus raphe magnus, and there were also some labeled neurons in the subcoeruleus nucleus. The nucleus raphe magnus also receives neuronal terminals from t h e lateral pontine area, including the nucleus locus subcoeruleus, i n cats."' Therefore, neurons in the rostra1 pontine lateral region and the caudal pontine ventral raphe region may project to the lumbosacral spinal cord, which innervates the bladder, urethra, and penis via t h e pelvic and pudendal nerves, and may be part of a n integrated neural mechanism. In decerebrate cats, electrical stimulation or injection of a cholinergic agent into the nucleus reticularis pontis oralis located ventromedial t o the pontine micturition center inhibits bladder Injection of a cholinergic agent into this nucleus also inhibits muscle activity in the extremities and inhibits the external urethral sphincter in decerebrate cats." In the present study, electrical stimulation of the reticular formation did not change the intracavernous pressure. Therefore, not all of the brainstem sites t h a t inhibit bladder contraction have a n effect on penile erection. However, 41 of 45 men with multiple sclerosis and impotence were reported to have bladder hyperreflexia on cystometry,'l" suggesting t h a t the central neural mechanisms for urine storage and penile erection may partially overlap. CONCLUSIONS
The overall effect of the brainstem is to inhibit penile erection. Both t h e subcoeruleus nucleus and the nucleus raphe magnus influence urine storage and penile erection. These regions have integrated neural networks. In addition, the pontine reticular formation influences urine storage but not affect penile erection. Therefore, t h e neural mechanisms for urine storage and penile erection may partially overlap in the brainstem. REFERENCES
FIG. 7. Location of lesions in pons are depicted sites marked as (0) evoked bladder contractions and sites marked as (0)increased intracavernous pressure upon electrical Stimulation in 10 decerebrate rats. Three transverse planes (Bregma -9.5 mm., -9.8 mm. and -10.6 mm.) were modified from Swanson's rat brain maps." B, Barrington's nucleus; GRN, gigantocellular reticular nucleus; ICP, inferior cerebellar peduncle; LC, locus coeruleus; LDT, laterodorsal tegmental nucleus; MEV, mesencephalic nucleus of trigeminal; m, medial vestibular nucleus; PB, parabrachial nucleus; PRNc, pontine reticular nucleus, caudal part; RM, nucleus raphe magnus; RPA, nucleus raphe pallidus; SCP, superior cerebellar peduncle; SLC, subcoeruleus nucleus; SLD, sublaterodorsal nucleus; V4, fourth ventricle; V, motor nucleus of the trigeminal nerve; VII, facial nucleus.
The raphe nuclei which project serotonergic nerve terminals to t h e brain and spinal cord have been reported to be associated with urine s tor age,^ penile erection,15 and muscle activity of the e ~ t r e m i t i e s . ' Electrical ~ stimulation of the nucleus raphe magnus inhibits the micturition reflex i n cats.7 Median and dorsal raphe lesions in the midbrain inhibit penile erection induced by fenfluramine, a serotonergic agonist, i n rats.' Penile erection is elicited by peripheral injection of m-chlorophenyl piperazine, a 5-HT1C agonist, and suppressed by 5-HT1A a n d o r 5-HT2 agonists." Electrical stimulation of the nucleus raphe magnus increases muscle activity of the extremities in decerebrate cats.16 We found t h a t electrical stimulation of this nucleus inhibited bladder contraction and increased t h e intracavernous pressure. In
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