* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Evidence of sympathetic ®bers in the male rat pelvic nerve
Survey
Document related concepts
Neural coding wikipedia , lookup
Clinical neurochemistry wikipedia , lookup
Neuropsychopharmacology wikipedia , lookup
Nervous system network models wikipedia , lookup
Central pattern generator wikipedia , lookup
Premovement neuronal activity wikipedia , lookup
Synaptic gating wikipedia , lookup
Axon guidance wikipedia , lookup
Optogenetics wikipedia , lookup
Feature detection (nervous system) wikipedia , lookup
Circumventricular organs wikipedia , lookup
Development of the nervous system wikipedia , lookup
Neuroanatomy wikipedia , lookup
Channelrhodopsin wikipedia , lookup
Neural engineering wikipedia , lookup
Transcript
International Journal of Impotence Research (1997) 9, 179±185 ß 1997 Stockton Press All rights reserved 0955-9930/97 $12.00 Evidence of sympathetic ®bers in the male rat pelvic nerve by Gross anatomy, retrograde labeling and high resolution Autoradiographic study F Giuliano1, P Facchinetti2, J BernabeÂ3, G Benoit1, A Calas2 and O Rampin3 1 Service d'Urologie, HoÃpital de BiceÃtre, 78 du GeÂneÂral Leclerc, 94270 Le Kremlin BiceÃtre, France and 2Institut des Neurosciences, URA CNRS 1488, 7 quai Saint-Bernard, Universite Paris VI, 75252 Paris Cedex, France and 3Laboratoire de Neurobiologie des Fonctions VeÂgeÂtatives, I.N.R.A., 78352 Jouy-en-Josas Cedex, France Several arguments exist in various animal species and man for the presence of a sympathetic component in the pelvic nerve, classically regarded as parasympathetic. We tested this hypothesis in the male rat. Nerve bundles issued from the sacral region of the paravertebral sympathetic chain and reaching the S1 spinal nerve were identi®ed. Neurons in the sacral parasympathetic nucleus of the L6-S1 spinal cord and in the L2-S1 paravertebral sympathetic chain were retrogradely labeled from the pelvic nerve. Radioautography evidenced labeling of unmyelinated ®bers in the pelvic nerve following in vitro incubation with 3H-noradrenaline. A population of sympathetic ®bers issued from the lumbosacral sympathetic chain exists in the pelvic nerve of the male rat. This qualitative study provides a morphological basis to uncover the role of the sympathetic out¯ow present in the pelvic nerve. Keywords: sympathetic ®bers; pelvic nerve; paravertebral sympathetic chain; lumbosacral spinal cord; urogenital tract Introduction The distal colon and the lower urinary and genital tracts receive neural supply from both sympathetic and parasympathetic divisions of the autonomic nervous system. In the rat parasympathetic preganglionic neurons going to the pelvis are located in the parasympathetic nucleus of the sacral spinal cord and their axons run in the pelvic nerve.1;2 Sympathetic preganglionic neurons to the pelvic organs project from the T13-L2 spinal cord to the inferior mesenteric ganglion and further travel distally in the hypogastric nerves. Other sympathetic ®bers travel caudally in the paravertebral sympathetic chain and could reach the pelvic organs either through the pudendal or pelvic nerves.3±6 In humans small trunks issued from the sacral sympathetic chain joining the pelvic nerves have been evidenced.7,8 Human pelvic nerves are very thin and ensheathed within a dense connective tissue. Their dissection is rather dif®cult and requires removal of the connective tissue.9,10 In human pelvic nerves, histochemical evidence of both cholinergic and adrenergic neural ®bers has been provided.11,12 The composition of the pelvic nerves is of clinical importance as the nerves may be lesioned during Correspondence: Dr F Giuliano. Received 5 February 1997; accepted 12 July 1997 surgery of the rectum or the bladder. The anatomical arrangement of the neural supply of the pelvic organs and distal colon seems to be generally applicable to all mammalian species investigated thus far, including man.6 In the male rat, pelvic neuroanatomy has been previously described and appears simpler when compared to larger species.13±16 In this species the pelvic and hypogastric nerves consist of ®ne bundles which are easily identi®able and both project onto the pelvic plexus, which ®gures as one large ganglion: the major pelvic ganglion. Accordingly the rat appears as a suitable model for anatomical and physiological studies on pelvic neuroanatomy.17±19 The aim of the present study was to demonstrate the mixed composition of the pelvic nerve, both sympathetic and parasympathetic, and to specify the origin of sympathetic ®bers in the pelvic nerve of the male rat. Materials and methods Seventeen adult male Sprague Dawley rats weighing 300±500 g were used in this study. Rats were anesthetized with ketamine (150 mg/kg i.p., ImalgeÁne 500, RhoÃne-MeÂrieux, Lyon, France) and dissections were performed with the aid of a Zeiss dissecting microscope equiped with a Zeiss camera. Topography of the pelvic nerve and its spinal origin Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al 180 and dissection of the lumbosacral sympathetic chain (LSC) were studied in 10 rats via a medial suprapubic incision and a transperitoneal approach. Retrograde labeling studies were performed using horseradish peroxidase in 2 rats (30% in sterile water, HRP type VI, Sigma, Saint-Quentin-Fallavier, France) and Fast Blue in 3 rats (2% in sterile water, FB, Sigma). The left pelvic nerve was dissected free and cut 3 mm proximal to the major pelvic ganglion. Its central cut end was immersed in one or the other neuroanatomical tracer for 30 min, then the pelvic cavity was washed with saline. Abdominal muscles and skin were stitched in separate layers. In 2 rats the pelvic nerve was immersed in HRP after a ligation of the nerve was performed centrally to the application site. Three days following HRP injection and 10 days after FB injection the rats were deeply anesthetized and intracardially perfused with 250 ml of a 0.9% saline solution followed by 1 l of ®xative solution containing 1% paraformaldehyde, 1.25% glutaraldehyde in 0.1 M sodium phosphate buffer (pH 7.4) at room temperature. After perfusion the L1-S4 LSC and the L6-S1 spinal cord were dissected and immersed in a solution of 30% sucrose in phosphate buffer (pH 7.4, 4 C) for 12± 16 h. Twenty micrometers thick longitudinal sections of LSC and transverse sections of the spinal cord were cut on the cryostat and mounted on gelatin-coated slides. HRP activity was revealed with tetramethylbenzidine as the chromogen.20 Following the reaction with hydrogen peroxide, the slices were washed in acetate buffer (10 mM, pH 4.8), counterstained with neutral red, dehydrated and mounted with permount. Sections were examined with a light (HRP) or ¯uorescence (FB) Leitz microscope equiped with an A1 ®lter (Ploempack system). The long axis of densely labeled neurons with clearly de®ned nuclei was measured. Labeled neurons were counted and correction for double counting was made according to Abercrombie.21 In one rat a 3 mm segment of the pelvic nerve was removed before its entry into the major pelvic ganglion. The fragment was immediately incubated in 1075 M 3H-noradrenaline (Amersham, Les Ulis, France) for 20 min at 37 C under constant stirring and oxygenation by air bubbling. The piece was then rinsed in saline for 5 min, ®xed with 4% glutaraldehyde in cacodylate buffer (0.1 M pH 7.4) for 1 h at 4 C and immersed in 2% osmium tetroxide in the same buffer, dehydrated in ethanol and embedded in Epon resin. Semi-thin sections (2 mm) were radioautographied with Ilford KI nuclear emulsion diluted 1:1 in water, exposed 15 days and developed in Microdol Kodak. After staining with toluidine blue the sections were mounted with permount and observed with light microscopy. Ultrathin transversal sections (50 nm) were cut from this block and mounted on celloidin coated glass slides, then contrasted with uranyl acetate and lead citrate. A thin layer of carbon was vaporized on the slides which were dipped into Ilford L4 nuclear track emulsion diluted 1:4 in water. After 1 month exposure, the autoradiographs were developed in Microdol Kodak. CelloõÈdin membranes were detached on water and mounted on 22-P-mesh grids. After thinning of the celloidin ®lm during 2.5 min in isoamylacetate, the autoradiographs were examined with a Philips EM 300 electron microscope. Results Gross anatomy The pelvic nerve originated from two joined bundles arising at the fusion of the L6-S1 spinal nerves proximal to the sacral plexus. The pelvic nerve travelled with a branch of the inferior vesical artery and ran dorsally to the internal iliac vein. The major pelvic ganglion was closely apposed by the pelvic fascia on the lateral side of the dorsal prostatic lobe anterior to the lateral wall of the rectum (Figure 1) and was naturally masked to the ventral approach by the distal vas deferens. The ganglion had an estimated surface of 3 mm2. The LSC laid on the anterior surface of the vertebral column, embedded in the psoas muscle, in the retroperitoneal space dorsal to the abdominal aorta and to the caudal vena cava. The LSC consisted in two parallel trunks, sometimes fusioned, along which several buldges occurred. Thin rami connected the trunks to the vertebrae. Further experiments con®rmed that buldges contained neuronal cell bodies and therefore represented LSC Figure 1 Schematic representation of a lateral view of the major pelvic ganglion of the male rat. Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al ganglia. LSC gave off lumbar splanchnic nerves which joined the intermesenteric nerve lying on the ventral surface of the aorta. Interindividual variation was present in the number and location of fusions between the two sympathetic trunks between L3 and L6 LSC ganglia. Caudally to the aortic bifurcation, the LSC was deeply located and twisted around the middle caudal artery, rendering dif®cult the identi®cation of sacral sympathetic ganglia in gross dissection. In 3 rats the LSC was carefully separated from the middle caudal artery. Two of these rats displayed a ®ne nerve bundle passing through psoas major and iliacus muscles. This bundle was given off by the cephalad part of the sacral segment of the sympathetic chain and joined the S1 spinal nerve (Figure 2). FB applied to the proximal cut end of the left pelvic nerve retrogradely labeled 124 5 neurons in the L6 segment and 85 28 neurons in the S1 segment of the spinal cord homolaterally to the injected nerve. In the transverse plane they formed a dense collection of closely packed cells in the lateral part of lamina VII (Figure 3). Labeled neurons were triangular or oval-shaped and their mean diameter was 19 2 mm. The L2-S1 LSC ganglia contained 182 21 neurons retrogradely labeled with FB (Figures 4 and 5a) and 106 7 neurons retrogradely labeled with HRP (Figure 5b). The majority of labeled neurons was present in the L3-L5 LSC ganglia (Figure 6). Retrogradely labeled neurons were oval-shaped, had a mean diameter of 12 2 mm and were homogeneously distributed in a ganglion. Following ligation of the pelvic nerve central to the site where HRP was applied respectively 12 and 15 neurons were labeled in the L2-S1 LSC. Semi-thin transversal sections of the pelvic nerve evidenced presence of several fascicles, some of which displayed silver grain accumulations. The majority of axons was unmyelinated, contained mitochondria, neurotubules and pro®les of endoplasmic reticulum, and was ensheathed with collagen ®bers. These axons appeared to be in close contact with myelinated ones. Clear vesicles were sparse and no varicosities or terminals were identi®ed on examined grids. Positive radioautographic reactions were noticed upon unmyelinated ®bers, closely apposed with unmarked ones (Figure 7a and b). Figure 2 Gross anatomical preparation of sacral sympathetic trunks as viewed from an anterior approach in the male rat. The rectum has been removed. In A: photograph of dissected sacral sympathetic chain and ®rst sacral spinal nerve is displayed. B: Diagrammatic representation of nerves identi®ed on photograph in A. Scale bar: 1 mm. 181 Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al 182 Figure 4 Segmental distribution of left sympathetic trunk postganglionic neurons projecting in the left pelvic nerve. These neurons were retrogradely labeled with Fast Blue. Data are expressed as mean sem (n 3). Figure 3 (a) Photomicrograph of a transverse section of the spinal cord at the level of S1 (20 mm thick, magni®cation: 6160). Neurons were retrogradely labeled with Fast Blue. Cell bodies are located in the sacral parasympathetic nucleus corresponding to the intermediolateral part of the gray matter (IML). Calibration: 50 mm. (b) At a higher magni®cation ( 6680) labeled neurons appeared triangular or oval-shaped. Calibration: 10 mm. Discussion The present study provides evidence that the pelvic nerve of the rat contains sympathetic postganglionic and parasympathetic preganglionic ®bers. We found that the pelvic nerve was issued from L6 and S1 spinal nerves, in agreement with previous studies conducted in male and female rats.2,19,22,23 Labeled neurons present in the intermediolateral column of the L6-S1 spinal cord con®rm that the cell bodies of parasympathetic neurons are located in the sacral parasympathetic nucleus.1,2 In humans, parasympathetic ®bers in the pelvic nerve arise in the S2-S4 sacral spinal nerves.7,8 In the cat, parasympathetic neurons projecting to the pelvic nerves are located in the S1-S3 spinal cord.24 Thus, it is likely that L6S1 levels of the rat spinal cord correspond to the S2S4 spinal levels in humans and to the S1-S3 levels in the cat. Individual variations in the arrangement of paravertebral sympathetic ganglia were found at the lumbar level in the rat. Fusion of both lumbar paravertebral sympathetic trunks often occurred but was not constant and locations of cross-connections differed, con®rming a previous description of anatomical organization of the paravertebral sympathetic chain of the rat.25 The sacral part of the sympathetic chain was dif®cult to individualize due to its close apposition to the median caudal artery. Small nerve ®laments joining the sacral sympathetic chain and the S1 spinal nerve may represent a contribution of the LSC either to the pelvic or to the pudendal nerve. The latter takes its origin in the sacral plexus formed by the L6-S1 spinal nerve trunk and the lumbosacral trunk composed of the L3-L5 spinal nerves.26 In cats and dogs connections between LSC and the pelvic nerve have previously been evidenced.27,28 In humans nerve bundles mainly issued from the fourth sacral sympathetic ganglion and joining the pelvic plexus have also been described.8,9 In human embryos the pelvic plexus receives neural ®bers from the sacral sympathetic trunk and its ganglia by rami connecting the sympathetic chain and the sacral nerves.29 Thus in various species including humans anatomical pathways joining the sacral sympathetic chain and the pelvic nerves or the pelvic plexus exist. LSC neurons retrogradely labeled from the pelvic nerve con®rm these anatomical ®ndings. The differ- Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al 183 Figure 6 Diagrammatic representation of the segmental distribution of postganglionic neurons in the lumbosacral sympathetic ganglia which were labeled with horseradish peroxidase applied to the left pelvic nerve in two rats (A and B). Figure 5 (a) Photomicrograph of neurons in the sympathetic ganglion L5 which were labeled with Fast Blue applied to the pelvic nerve. Calibration: 10 mm. (b) Photomicrograph of neurons in the sympathetic ganglion L5 which were labeled with horseradish peroxidase (arrows) applied to the pelvic nerve. Calibration: 10 mm. ence in the nature of the retrograde tracers used, HRP and FB respectively, may explain the differences between the number of labeled neurons in the LSC in each experience. Postganglionic sympathetic cell bodies which axons travel in the pelvic nerve are widely distributed in the L2-S1 sympathetic ganglia and are mainly located in the L3-L5 ganglia. In the female rat the sympathetic innervation of the bladder is conveyed in equal parts by the hypogastric and pelvic nerves.30 Postganglionic neurons destined to the bladder are numerous in the L6 LSC ganglion and sparse in the L1-L5 ganglia. In the male rat, retrograde tracing from the cavernous nerve labeled about 100 neurons in the paravertebral sympathetic chain mainly at the L5-L6 levels.31 In the present study LSC neurons which send axons in the pelvic nerve are more numerous and more widely distributed. It may be partly explained by the fact that the pelvic nerve contains neurons destined to different pelvic viscera. Bilateral location of retrogradely labeled neurons occurred in the Figure 7 (a) Radioautography of a cross section of the pelvic nerve in an adult male rat after the nerve was incubated in vitro in 3 H-noradrenaline. Magni®cation: 69000; calibration: 1 mm. (b) Detail from a: positive radioautographic reaction upon one fascicle of unmyelinated axons. Magni®cation: 630000; calibration 300 nm. Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al 184 LSC. It is therefore possible that postganglionic axons in the LSC cross over to the opposite side. In our study the number of retrogradely labeled neurons in the sacral parasympathetic nucleus and in the LSC was comparable. Extended lesion of the LSC or ventral rhizotomy (T12-S2) decreased by 25% the number of ®bers in the rat pelvic nerve31 . To our knowledge there is no clear demonstration of a loss of neural ®bers in the pelvic nerve following lesion of the lumbosacral spinal cord in the rat. However retrograde labelling from the pelvic nerve in this species suggests that the lumbosacral spinal cord contribute a contingent of ®bers in the pelvic nerve. It is also unclear whether preganglionic parasympathetic neurons may give off several axons. This fact would lead to an overestimation of the contribution of sacral preganglionic neurons following lesion of the lumbosacral spinal cord. The cat's pelvic nerve contains twice as many postganglionic sympathetic ®bers as preganglionic parasympathetic ®bers.24 In the rhesus monkey 40% of the pelvic nerve's efferent ®bers originate in the LSC ganglia.32 The ultrastructural study of the rat's pelvic nerve revealed the presence of several fascicles with few myelinated and many unmyelinated axons, con®rming a report of 80% unmyelinated axons in the rat pelvic nerve.31 The noradrenaline uptake ability of some unmyelinated axons supports the view that the pelvic nerve contains catecholaminergic ®bers which are very likely postganglionic sympathetic axons.33 In human pelvic nerves, catecholaminergic ®bers have been identi®ed using formaldehyde ¯uorescence technique.11 Since the pelvic plexus provides innervation to the pelvic organs, neurons arising in the paravertebral sympathetic trunks and conveyed by the pelvic nerve may be destined to various pelvic viscera. The cat urinary bladder receives sympathetic ®bers issued from the LSC.34 In the female rat, sympathetic innervation to the bladder also arises from the pelvic nerves.30 Nevertheless the physiological role of the sympathetic ®bers in the pelvic nerves remains unknown. In dogs, cats, rabbits and rats electrical stimulation of the lower part of the sympathetic chain elicits subsidence of erection.34ÿ39 Sympathetic ®bers issued from the lumbar sympathetic chain may reach their targets either through the pelvic or the pudendal nerves. In the rat electrical stimulation of the L4-L5 LSC elicits a response on the dorsal nerve of the penis.40 There is no evidence for a role of sympathetic ®bers in the pelvic nerve in the control of erection. These ®bers may contribute to the control of micturition or secretion and motility of sex accessory glands.34 Recent data suggest that sympathetic ®bers in the pelvic nerve and sympathetic chain provide input into the testes.41 Furthermore some labelling of neurons in the lumbosacral sympathetic chain from the penis has been demonstrated.42 In conclusion in the rat the pelvic ganglion receives sympathetic inputs from both the hypogastric nerve and the paravertebral sympathetic chain. Fibers issued from the latter run in the pelvic nerve. Acknowledgements Pr Jacques Taxi and Mrs Sylvette Gougis are gratefully acknowledged for their help in the present experiment. References 1 Hancock MB, Peveto CA. Preganglionic neurons in the sacral spinal cord of the rat: an HRP study. Neurosci Lett 1979; 11: 1±5. 2 Nadelhaft I, Booth AM. The location and morphology of preganglionic neurons and the distribution of visceral afferents from the rat pelvic nerve: a horseradish peroxidase study, J Comp Neurol 1984; 226: 238±245. 3 Hancock MB, Peveto CA. A preganglionic autonomic nucleus in the dorsal gray commissure of the lumbar spinal cord of the rat. J Comp Neurol 1979; 183: 65±72. 4 Dail WG, Trujillo D, DeLaRosa D, Walton G. Autonomic innervation of reproductive organs: analysis of the neurons whose axons project in the main penile nerve in the pelvic plexus of the rat, Anat Rec 1989; 224: 94±101. 5 Nadelhaft I, McKenna KE. Sympathetic afferent and preganglionic neurons labelled by horseradish peroxidase applied to the hypogastric nerve and the sacral sympathetic chain of the rat. Soc Neurosci Abstr 1985; 11: 764. 6 JaÈnig W, McLachlan EM. Organization of lumbar sympathetic out¯ow to distal colon and pelvic organs. Physiol Rev 1987; 67: 1332±1404. 7 De Groat WC, Steers WD. Neuroanatomy and neurophysiology of penile erection. In: Contemporary Management of Impotence and Infertility. Tanagho EA, Lue TF & McClure RD eds., Williams & Wilkins: Baltimore, 1988, pp 3±27. 8 Benoit G, Delmas V, Gillot C, Jardin A. The anatomy of erection. Surg Radiol Anat 1987; 9: 263±272. 9 Latarjet A, Bonnet P. Le plexus hypogastrique chez l'homme. Lyon Chir 1913; 9: 221. 10 Cordier P, Couloma A. Les nerfs eÂrecteurs. Bull Assoc Anat (Paris) 1933; 143: 199. 11 Benoit G et al. Identi®cation histologique des affeÂrences du plexus pelvien. Prog Urol 1991; 1: 132±138. 12 Fritsch H. Topography of the pelvic autonomic nerves in human fetuses between 21±29 weeks of gestation. Anat Embryol 1989; 180: 57±64. 13 Langworthy OR. Innervation of the pelvic organs of the rat. Invest Urol 1965; 2: 491±511. 14 Foroglou C, Winckler G. CaracteÂristiques du plexus hypogastrique infeÂrieur (pelvien) chez le rat. Bull Ass Anat 1973; 57: 853±866. 15 Purinton PT, Fletcher TF, Bradley WE. Gross and light microscopic features of the pelvic plexus in the rat. Anat Rec 1973; 175: 697±706. 16 Tanaka S, Zukeran C, Nakagawa S, Nakao T. A macroscopical study of the somatic and visceral nerves innervating the male rat urogenital organs. Acta Anat Nippon 1981; 56: 400±414. 17 Steers WD, Mallory B, DeGroat WC. Electrophysiological study of neural activity in penile nerve of the rat. Am J Physiol 1988; 254: R989±R1000. 18 Mallory B, Steers WD, DeGroat WC. Electrophysiological study of micturition re¯exes in rats. Am J Physiol 1989; 257: R410±R421. Evidence of sympathetic ®bers in the male rat pelvic nerve F Giuliano et al 19 Quinlan DM et al. The rat as a model for the study of penile erection. J Urol 1989; 141: 656±661. 20 Mesulam MM. Tetramethylbenzidine for horseradish peroxidase neurohistochemistry. A non-carcinogenic blue reactionproduct with superior sensitivity visualizing neural afferents and efferents. J Histochem Cytochem 1978; 26: 106±117. 21 Abercrombie M. Estimation of nuclear population from microtome sections. Anat Rec 1946; 94: 239±247. 22 Greene EC. Anatomy of the Rat. Hafner Publishing Co: New York, 1959. 23 Baljet B, Drukker J. The extrinsic innervation of the pelvic organs in the female rat. Acta Anat 1980; 107: 241±267. 24 Nadelhaft I, DeGroat WC, Morgan C. Location and morphology of parasympathetic preganglionic neurons in the spinal cord of the cat revealed by retrograde axonal transport of horseradish peroxidase. J Comp Neurol 1980; 193: 265±281. 25 Baron R, JaÈnig W, Kollmann W. Sympathetic and afferent somata projecting in hindlimb nerves and the anatomical organization of the lumbar sympathetic nervous system of the rat. J Comp Neurol 1988; 275: 460±468. 26 Mc Kenna KE, Nadelhaft I. The organization of the pudendal nerve in the male and female rat. J Comp Neurol 1986; 248: 532±549. 27 Kuo DC, Hisamitsu T, DeGroat WC. A sympathetic projection from sacral paravertebral ganglia to the pelvic nerve and to postganglionic nerves on the surface of the urinary bladder and large intestine of the cat. J Comp Neurol 1984; 226: 76±86. 28 Kihara K et al. Lumbosacral sympathetic trunk as a compensatory pathway for seminal emission after bilateral hypogastric nerve transections in the dog. J Urol 1991; 145: 640±643. 29 Pearson AA, Sauter RW. Nerve contributions to the pelvic plexus and the umbilical cord. Am J Anat 1970; 128: 495±498. 30 Vera PL, Nadelhaft I. Afferent and sympathetic innervation of the dome and the base of the urinary bladder of the female rat. Brain Res Bull 1992; 29: 651±658. 31 Hulseboch CE, Coggeshall RE. An analysis of the axon populations in the nerves of the pelvic viscera in the rat. J Comp Neurol 1982; 211: 1±10. 32 Schnitzlein HN, Hoffman HH, Tucker CC, Quigley MB. The pelvic splanchnic nerves of the male rhesus monkey. J Comp Neurol 1960; 114: 51±65. 33 Taxi J, Droz B. Etude de l'incorporation de noradreÂnaline tritieÂe (NA3H) et de 5-hydroxytryptophane 3H (5.HTP.3H) dans l'eÂpiphyse et le ganglion cervical supeÂrieur. Comptesrendus Acad Sci (Paris) 1966; 263: 1326±1329. 34 Langley JN, Anderson HK. The innervation of the pelvic and adjoining viscera. J Physiol (London) 1895; 19: 71±130. 35 Semans JH, Langworthy OR. Observations on the neurophysiology of sexual functions in the male cat. J Urol 1938; 40: 836±846. 36 SjoÈstrand NO, Klinge EK. Principal mechanisms controlling penile retraction and protrusion in rabbits. Acta Physiol Scand 1979; 106: 199±214. 37 JuÈnemann KP et al. Neurophysiological aspects of penile erection: the role of the sympathetic nervous system. Brit J Urol 1989; 64: 84±92. 38 Diederichs W, Stief CG, Lue TF, Tanagho EA. Sympathetic inhibition of papaverine induced erection. J Urol 1991; 146: 195±198. 39 Giuliano F, Rampin O, Bernabe J, Rousseau JP. Neural control of penile erection in the rat. J Autonom Nerv Sys 1995; 55: 36± 44. 40 Giuliano F, Rampin O, Jardin A, Rousseau JP. Electrophysiological study of relations between the dorsal nerve of the penis and the lumbar sympathetic chain in the rat. J Urol 1993; 150: 1960±1964. 41 Rauchenwald M, Steers WD, Desjardins C. Efferent innervation of the rat testis. Biol Reprod 1995; 52: 1136±1143. 42 Vanhatalo S, Klinge E, SjoÈstrand NO, Soinila S. Nitric oxidesynthesizing neurons originating at several different levels innervate the rat penis. Neuroscience 1996; 75: 891±899. 185