Ibrahim Karam

Division of Autonomic Nerves Within the Neurovascular Bundles Distally into Corpora Cavernosa and Corpus Spongiosum Components: Immunohistochemical Confirmation with Three-Dimensional Reconstruction

BACKGROUND Detailed knowledge of the distribution and distal course of periprostatic nerves is essential to improve functional outcomes (erection and continence) after radical prostatectomy (RP). OBJECTIVE To describe the location of nerve fibres within neurovascular bundles (NVBs) and around the prostate by three-dimensional (3D) computer-assisted anatomic dissection (CAAD) in human foetuses and adult cadavers. DESIGN, SETTING, AND PARTICIPANTS Serial transverse sections of the pelvic portion were performed in seven human male foetuses and four male adult cadavers. Sections were treated by histologic coloration and neuronal immunolabelling of S100 protein. 3D pelvic reconstruction was achieved with digitised serial sections and WinSurf software. MEASUREMENTS We evaluated the distribution of nerve fibres within the NVB qualitatively. The distribution of periprostatic nerves was also evaluated quantitatively in the adult specimens. RESULTS AND LIMITATIONS Periprostatic nerve fibres were dispersed around the prostate on all sides with a significant percentage of these fibres present in the anterior and anterolateral sectors. At the prostate apex and the urethral levels, the NVBs have two divisions: cavernous nerves (CNs) and corpus spongiosum nerves (CSNs). The CNs were a continuation of the anterior and anterolateral fibres around the apex of the prostate, travelling towards the corpora cavernosa. The CSNs were a continuation of the posterolateral NVBs, and they eventually reached the corpus spongiosum. The limitations of this study were the small number of specimens available and the lack of functional information. CONCLUSIONS The anterolateral position of CNs at the apex of the prostate and the autonomic innervation towards the corpus spongiosum via CSNs indicate possible ways to minimise the effect of prostate surgery on sexual function. The ideal dissection plane should probably include the preservation of the anterolateral tissues and fascias to avoid CN lesions. Anatomic knowledge gained from CAAD pertains directly to proper surgical technique and subsequent recovery of erectile function after RP.

Coexistence of adrenergic and cholinergic nerves in the inferior hypogastric plexus: anatomical and immunohistochemical study with 3D reconstruction in human male fetus

Classic anatomical methods have failed to determine the precise location, origin and nature of nerve fibres in the inferior hypogastric plexus (IHP). The purpose of this study was to identify the location and nature (adrenergic and/or cholinergic) of IHP nerve fibres and to provide a three‐dimensional (3D) representation of pelvic nerves and their relationship to other anatomical structures. Serial transverse sections of the pelvic portion of two human male fetuses (16 and 17 weeks’ gestation) were studied histologically and immunohistochemically, digitized and reconstructed three‐dimensionally. 3D reconstruction allowed a ‘computer‐assisted dissection’, identifying the precise location and distribution of the pelvic nerve elements. Proximal (supra‐levator) and distal (infra‐levator) communications between the pudendal nerve and IHP were observed. By determining the nature of the nerve fibres using immunostaining, we were able to demonstrate that the hypogastric nerves and pelvic splanchnic nerves, which are classically considered purely sympathetic and parasympathetic, respectively, contain both adrenergic and cholinergic nerve fibres. The pelvic autonomic nervous system is more complex than previously thought, as adrenergic and cholinergic fibres were found to co‐exist in both ‘sympathetic’ and ‘parasympathetic’ nerves. This study is the first step to a 3D cartography of neurotransmitter distribution which could help in the selection of molecules to be used in the treatment of incontinence, erectile dysfunction and ejaculatory disorders.

The structure and innervation of the male urethra: histological and immunohistochemical studies with three-dimensional reconstruction.

The structure of the striated urethral sphincter, the so‐called rhabdosphincter, remains the subject of controversy. There are two main concepts regarding its structure: either it is a part of the urogenital diaphragm, or it extends from the base of the bladder up to the urogenital diaphragm and is an integral part of the urethra. It is also uncertain whether it possesses a somatic innervation or a mixed innervation (i.e. autonomic and somatic). The purpose of this study was to show the precise location of the nerves running to the urethra, and to try to determine their exact nature. Histology and immunohistochemistry were performed in the external urethral sphincter of ten male fetuses (114–342 mm crown–rump length, or between 14 and 40 weeks of gestation). A three‐dimensional (3D) reconstruction of the urethral structure and its innervation was made from serial sections. The 3D reconstruction of the same section levels with different strains allowed us to identify the precise structure of the muscle layers (smooth and striated muscle fibres) and the nature of the nerve elements (myelinated and unmyelinated), their distributions and their relationship to the urethral wall, the prostate and the seminal vesicles. Histological and immunohistochemical 3D reconstruction of the anatomical elements of the urethral sphincter helps us to understand the 3D arrangement of the sphincter muscle layers. It also provides a better understanding of the origin and nature of the nerve elements that play a role in urinary continence.

Tridimensional Computer-Assisted Anatomic Dissection of Posterolateral Prostatic Neurovascular Bundles

BACKGROUND Detailed knowledge of nerve distribution in the neurovascular bundle (NVB) is essential to preserve sexual function after prostatic surgery. OBJECTIVE To identify the location as well as the type (adrenergic, cholinergic, and sensory) of nerve fibres within the NVB and to provide a three-dimensional (3D) representation of their structural relationship in the human male foetus. DESIGN, SETTING, AND PARTICIPANTS Serial transverse sections were performed every 150-200 microm in the pelvic portion of six human male foetuses (15-20 wk of gestation). Sections were treated with histologic and immunohistochemical methods (hematin-eosin-safran, Luxol Fast Blue, immunolabelling of protein S100, vesicular acetylcholine transporter, tyrosine hydroxylase, calcitonin gene-related peptide, and substance P). The 3D pelvic reconstruction was obtained from digitised serial sections using WinSurf software. MEASUREMENTS NVB nerve location and type were evaluated qualitatively. RESULTS AND LIMITATIONS The 3D reconstruction allowed precise identification of pelvic organ innervation. Nerve fibres derived from the inferior hypogastric plexus followed two courses: posterior and lateral, providing cholinergic, adrenergic, and sensory innervation to seminal vesicles, vas deferens, prostate, and urethral sphincter. Cavernous nerve fibres did not strictly follow the NVB course; they were distributed at several levels, in a fanlike formation. The main limitations of this study were the limited number of specimens available due to legal restriction and the time-consuming nature of the manually performed stages in the method. CONCLUSIONS The distribution of nerve fibres within the posterolateral prostatic NVB and the existence of mixed innervation in the posterior and lateral fibre courses at the level of the prostate and seminal vesicles give us an insight into how to minimise effects on sexual function during prostatic surgery. The 3D computer-assisted anatomic dissection represents an original method of applying anatomic knowledge to surgical technique to improve nerve preservation and decrease postoperative sexual complications.

Autonomic-somatic communications in the human pelvis: computer-assisted anatomic dissection in male and female fetuses

Sphincter continence and sexual function require co‐ordinated activity of autonomic and somatic neural pathways, which communicate at several levels in the human pelvis. However, classical dissection approaches are only of limited value for the determination and examination of thin nerve fibres belonging to autonomic supralevator and somatic infralevator pathways. In this study, we aimed to identify the location and nature of communications between these two pathways by combining specific neuronal immunohistochemical staining and three‐dimensional reconstruction imaging. We studied 14 normal human fetal pelvic specimens (seven male and seven female, 15–31 weeks’ gestation) by three‐dimensional computer‐assisted anatomic dissection (CAAD) with neural, nitrergic and myelin sheath markers. We determined the precise location and distribution of both the supra‐ and infralevator neural pathways, for which we provide a three‐dimensional presentation. We found that the two pathways crossed each other distally in an X‐shaped area in two spatial planes. They yielded dual innervation to five targets: the anal sphincter, levator ani muscles, urethral sphincter, corpus spongiosum and perineal muscles, and corpora cavernosa. The two pathways communicated at three levels: proximal supralevator, intermediary intralevator and distal infralevator. The dorsal penis/clitoris nerve (DN) had segmental nitrergic activity. The proximal DN was nNOS‐negative, whereas the distal DN was nNOS‐positive. Distal communication was found to involve interaction of the autonomic nitrergic cavernous nerves with somatic nitrergic branches of the DN, with nitrergic activity carried in the distal part of the nerve. In conclusion, the pelvic structures responsible for sphincter continence and sexual function receive dual innervation from the autonomic supralevator and the somatic infralevator pathways. These two pathways displayed proximal, intermediate and distal communication. The distal communication between the CN and branches of the DN extended nitrergic activity to the distal part of the cavernous bodies in fetuses of both sexes. These structures are important for erectile function, and care should therefore be taken to conserve this communication during reconstructive surgery.

Computer-assisted anatomic dissection (CAAD): evolution, methodology and application in intra-pelvic innervation study

ObjectiveClassic anatomical methods have limitations in micro determination of nerve fibre location. Furthermore, the precise detection of the nerve fibres nature is not possible by means of dissection. The combination of immunohistochemistry and three-dimensional reconstruction could be used to resolve these limitations of morphological sciences. Our aim is to describe the evolution of computer-assisted anatomic dissection (CAAD), which is an original method applied to study the distribution of intra-pelvic nerves in anatomic research.Materials and methodsSerial transverse sectioning of the pelvic region in rabbit, human fetus, infant and adult cadaver was performed. Sections were immuno-histochemically stained and digitized with a high optical resolution scanner. Photoshop 7 software was used in regrouping of the adult cadaver sections then a tri-dimensional reconstruction was achieved using WinSurf software.ResultsThe 3D reconstruction of the immuno-histochemically stained histologic sections of the pelvis allowed for precise structural identification of the prostate and its innervations (in fetus, infant and adult). In addition, we reconstructed the entire intra-pelvic organs with accurate demonstration of the location of both adrenergic and cholinergic pathways. Moreover, we performed a virtual dissection of each of the pelvic structures with description of the exact location of the inferior hypogastric plexus, as well as the nature and the distribution of its fibres.ConclusionThe CAAD is an original method in anatomic research, which illustrates the fact that descriptive anatomy is still a dynamic science. This method allows for a 3D presentation of the intra-organic innervation, the nature of the nerve fibres, and the distribution of receptors and their neurotransmitters. This technique improves the understanding of the complex anatomic regions such as the pelvis from both surgical and educational point of view.

Effect of a Local Delivery of Triiodothyronine (T3) Within Neuroregenerative Guide on Recovery of Erectile Function in a Rat-Model of Cavernous Nerve Injury

INTRODUCTION A promoting effect of thyroid hormones has been established on the maturation of central and peripheral nervous systems. However, effects on autonomic nerves have never been experimentally investigated. AIM To assess the effect of a local treatment combining silicone guides and local administration of Triiodothyronine (T3) on the erectile function and the histological neuroregeneration of crushed cavernous nerves (CNs) in rats. METHODS Forty-five rats were divided into five equal groups: SHAM surgery, guide without crush, crush, crush + guide, crush + guide + T3. All surgical procedures were bilateral. CNs were crushed with microvascular bulldog clamp of 100 g/cm(2). A silicone guide was placed around the nerves. The guides were filled with T3 neuroregenerative solution. MAIN OUTCOME MEASURES Erectile function was assessed 10 weeks post-operatively. Intra-cavernous pressure (ICP) and mean arterial pressure (MAP) were monitored during electrical stimulation of CNs at various frequencies. The main outcome was hardness of erection defined as DeltaICP/MAP. Fluorescent immunohistochemical analysis of CNs was performed to assess regeneration of nerves morphologically. RESULTS Electrophysiological data showed increased recovery of erectile function in the group with guide + T3 neuroregenerative solution compared with the empty guide. Immunohistochemical analysis of cavernous nerves demonstrated in morphology that regenerated axons were straighter in nerves with guide and more regular if guides had been filled with T3. CONCLUSION The use of guides prevented axonal sprouting, facilitated functional neuroregeneration and enabled a local delivery of thyroid hormones. Triiodothyronine improved neuroregeneration and recovery of erectile function after a nerve-sparing-like injury in a rat model.

Musculature et innervation de l’urètre masculin : étude histologique et immuno-histochimique avec reconstruction tridimensionnelle

But La structure du rhabdosphincter reste un sujet controverse ; deux concepts sont trouves dans la litterature : soit il fait partie du diaphragme urogenital et il se situe en position caudale par rapport a la prostate, soit il s’etend de la base de la vessie jusqu’au diaphragme urogenital. Possede-t-il une innervation somatique ou une innervation mixte ? Materiels et methodes Une etude microscopique histologique et immuno-histochimique (colorations speciales : trichrome Masson et Luxol Fast Blue, immuno-marquage neuronal par proteine S100 et immuno-marquage musculaire de l’actine lisse) a ete realisee sur dix fœtus de sexe masculin. Une reconstruction tridimensionnelle de l’uretre et de son innervation a ete realisee, a partir d’une serie des coupes bidimensionnelles. Resultats L’etude microscopique a permis la mise en evidence d’une innervation double (somatique pour la musculature striee et autonome pour la musculature lisse). La reconstruction tridimensionnelle a permis de representer les fibres nerveuses myelinisees et non myelinisees ; leur distribution et leurs rapports avec la paroi uretrale, la prostate et les vesicules seminales. Conclusion L’etude de la structure propre de l’uretre masculin et de son innervation, permet de mieux comprendre les mecanismes de la continence et preserver les nerfs du sphincter et ceux de l’erection en limitant la dissection a la face externe des vesicules seminales et en sectionnant les vaisseaux des ailerons a leur contact. La conservation de la prostate caudale (la coque prostatique) et des vesicules seminales permet de preserver la continence, il evite de sectionner les bandelettes neurovasculaires et permet de preserver les erections.