Extrinsic innervation of the pelvic organs in the lesser pelvis of human embryos.

Realistic models to understand the developmental appearance of the pelvic nervous system in mammals are scarce. We visualized the development of the inferior hypogastric plexus and its preganglionic connections in human embryos at 4-8 weeks post-fertilization, using Amira 3D reconstruction and Cinema 4D-remodelling software. We defined the embryonic lesser pelvis as the pelvic area caudal to both umbilical arteries and containing the hindgut. Neural crest cells (NCCs) appeared dorsolateral to the median sacral artery near vertebra S1 at ~5 weeks and had extended to vertebra S5 1 day later. Once para-arterial, NCCs either formed sympathetic ganglia or continued to migrate ventrally to the pre-arterial region, where they formed large bilateral inferior hypogastric ganglionic cell clusters (IHGCs). Unlike more cranial pre-aortic plexuses, both IHGCs did not merge because the 'pelvic pouch', a temporary caudal extension of the peritoneal cavity, interposed. Although NCCs in the sacral area started to migrate later, they reached their pre-arterial position simultaneously with the NCCs in the thoracolumbar regions. Accordingly, the superior hypogastric nerve, a caudal extension of the lumbar splanchnic nerves along the superior rectal artery, contacted the IHGCs only 1 day later than the lumbar splanchnic nerves contacted the inferior mesenteric ganglion. The superior hypogastric nerve subsequently splits to become the superior hypogastric plexus. The IHGCs had two additional sources of preganglionic innervation, of which the pelvic splanchnic nerves arrived at ~6.5 weeks and the sacral splanchnic nerves only at ~8 weeks. After all preganglionic connections had formed, separate parts of the inferior hypogastric plexus formed at the bladder neck and distal hindgut.

Careful Dissection of the Distal Ureter Is Highly Important in Nerve-sparing Radical Pelvic Surgery: A 3D Reconstruction and Immunohistochemical Characterization of the Vesical Plexus.

OBJECTIVE: Radical hysterectomy with pelvic lymphadenectomy (RHL) is the preferred treatment for early-stage cervical cancer. Although oncological outcome is good with regard to recurrence and survival rates, it is well known that RHL might result in postoperative bladder impairments due to autonomic nerve disruption. The pelvic autonomic network has been extensively studied, but the anatomy of nerve fibers branching off the inferior hypogastric plexus to innervate the bladder is less known. Besides, the pathogenesis of bladder dysfunction after RHL is multifactorial but remains unclear. We studied the 3-dimensional anatomy and neuroanatomical composition of the vesical plexus and describe implications for RHL. MATERIALS AND METHODS: Six female adult cadaveric pelvises were macroscopically dissected. Additionally, a series of 10 female fetal pelvises (embryonic age, 10-22 weeks) was studied. Paraffin-embedded blocks were transversely sliced in 8-µm sections. (Immuno) histological analysis was performed with hematoxylin and eosin, azan, and antibodies against S-100 (Schwann cells), tyrosine hydroxylase (postganglionic sympathetic fibers), and vasoactive intestinal peptide (postganglionic parasympathetic fibers). The results were 3-dimensionally visualized. RESULTS: The vesical plexus formed a group of nerve fibers branching off the ventral part of the inferior hypogastric plexus to innervate the bladder. In all adult and fetal specimens, the vesical plexus was closely related to the distal ureter and located in both the superficial and deep layers of the vesicouterine ligament. Efferent nerve fibers belonging to the vesical plexus predominantly expressed tyrosine hydroxylase and little vasoactive intestinal peptide. CONCLUSIONS: The vesical plexus is located in both layers of the vesicouterine ligament and has a very close relationship with the distal ureter. Complete mobilization of the ureter in RHL might cause bladder dysfunction due to sympathetic and parasympathetic denervation. Hence, the distal ureter should be regarded as a risk zone in which the vesical plexus can be damaged.

Anatomical basis of the coordination between smooth and striated urethral and anal sphincters: loops of regulation between inferior hypogastric plexus and pudendal nerve. Immuno-histological study with 3D reconstruction.

PURPOSE: (1) Describe both nervous pathways to the sphincters, and highlight the anatomical support of their coordination. (2) Obtain a 3D representation of this complex innervation system. METHODS: A computer-assisted anatomical dissection technique was used. Serial histological sections were cut in the pelvis of four female human foetuses (aged 19-32 weeks of gestation). The sections were treated with conventional staining, and with seven different immunostainings. The sections were digitalized and, finally, a 3D representation was built from the corresponding images. RESULTS: Myelinated and sensory fibres were detected at the inferior hypogastric plexus (IHP) level. Our analysis showed that most of the afferent sensory fibres come from the urinary and anal sphincters through the anterior and posterior branches of the IHP respectively. A highly positive nitrergic (anti-NOS1) and sensitive (anti-CGRP) labelling was found in the external layer of the urethral sphincter. The 3D representation allowed describing the two components of the innervation system. A sensory-motor regulation loop was found for both sphincters. CONCLUSION: A 3D description of the components of both nervous pathways to the sphincters has been established. Our findings on the innervation of the sphincters tend to question the classical infra/supra levatorian muscle description. The coordinated work of the internal and external layers of the anal and urethral sphincter is probably mediated by multiple roles regulation.

Optimal plane for nerve sparing total mesorectal excision, immunohistological study and 3D reconstruction: an embryological study.

AIM: Genito-urinary complications are frequent after rectal surgery and are often due to nerve damage. The relationship between the pelvic nerves and surgical planes are unclear. The aim of the study was to determine the relationship between the inferior hypogastric plexus and the fascia of the lateral pelvic wall and between Denonvilliers' fascia and the efferent branches of the inferior hypogastric plexus. METHOD: Computer-assisted anatomical dissection was used. Serial histological sections were made from six human foetuses and a male adult. Sections were stained with haematoxylin and eosin, Masson's trichrome and immunostainings. The sections were then digitalized and reconstructed in three dimensions. RESULTS: The inferior hypogastric plexus was situated in a virtual space between the fascia propria of the rectum and the fascia on the upper surface of the levator ani. During the lateral dissection, the optimal surgical plane is the plane of the fascia propria of the rectum. We located Denonvilliers' fascia in three dimensions. It plays the role of a protective sheet for the neurovascular bundle. The optimal plane for nerve preservation is situated behind Denonvilliers' fascia. CONCLUSION: This study has enabled a clear visualization of the optimal planes to perform total mesorectal excision while ensuring nerve preservation. Three-dimensional visualization clearly helps to bridge the gap between histological examination and the findings of surgery.

Identification of the origin of adrenergic and cholinergic nerve fibers within the superior hypogastric plexus of the human fetus.

Nerve fibers contributing to the superior hypogastric plexus (SHP) and the hypogastric nerves (HN) are currently considered to comprise an adrenergic part of the autonomic nervous system located between vertebrae (T1 and L2), with cholinergic aspects originating from the second to fourth sacral spinal segments (S2, S3 and S4). The aim of this study was to identify the origin and the nature of the nerve fibers within the SHP and the HN, especially the cholinergic fibers, using computer-assisted anatomic dissection (CAAD). Serial histological sections were performed at the level of the lumbar spine and pelvis in five human fetuses between 14 and 30 weeks of gestation. Sections were treated with histological staining [hematoxylin-eosin (HE) and Masson's trichrome (TriM)] and with immunohistochemical methods to detect nerve fibers (anti-S100), adrenergic fibers (anti-TH), cholinergic fibers (anti-VAChT) and nitrergic fibers (anti-nNOS). The sections were then digitalized using a high-resolution scanner and the 3D images were reconstructed using winsurf software. These experiments revealed the coexistence of adrenergic and cholinergic fibers within the SHP and the HNs. One-third of these cholinergic fibers were nitrergic fibers [anti-VACHT (+)/anti-NOS (+)] and potentially pro-erectile, while the others were non-nitrergic [anti-VACHT (+)/anti-NOS (-)]. We found these cholinergic fibers arose from the lumbar nerve roots. This study described the nature of the SHP nerve fibers which gives a better understanding of the urinary and sexual dysfunctions after surgical injuries.

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.

The developmental anatomy of the human superior hypogastric plexus: A morphometrical investigation with clinical and surgical correlations.

The superior hypogastric plexus (SHP) is the part of the autonomic nervous system, which is responsible for the sympathetic innervation of pelvic organs and extrapelvic genitals in humans of both sexes. The SHP also functions as the anatomic pathway for the major part of visceral sensitive fibers originating from pelvic viscera. In this study, the morphology of the SHP was analyzed through anatomical dissections performed both in human adult and fetal cadavers. A computerized morphometrical investigation of the SHP was also performed and the resulting quantitative data statistically assessed. The comparison between fetal and adult SHP revealed that in the male group there was a developmental increase of six times (in height) and of about five times (in width); while in the female group, there was a developmental increase of 3.5 times both in height and width values. In addition, the distance from the superior border of the SHP to the bifurcation of the common iliac arteries presented a developmental increase of about six times in the male group, and about four times in the female group. We propose an original morphological classification with six types, based upon the anatomical arrangement of the nervous fibers in this autonomic plexus.

Development of the human retroperitoneal fasciae.

INTRODUCTION: Although the renal fascia (RF), ureteral sheath, lateroconal fascia (LF) and hypogastric nerve are critical landmarks for retroperitoneal surgery, their laminar relationships require clarification. MATERIALS AND METHODS: Horizontal sections (hematoxylin-eosin staining) of human fetuses at two different developmental stages [9-12 (3 fetuses, crown-rump length, CRL 40-65 mm) and 20-25 weeks of gestation (9 fetuses, CRL 152-220 mm)] were compared. RESULTS: In the early-stage group, the pararenal space had already formed between the posterior RF and the transversalis fascia (TF). The anterior RF extended along the peritoneum and often fused with the latter. In the late-stage group, the posterior RF extended inferomedially toward the anterior aspect of the aorta and inferior vena cava. However, at the level of the renal hilus, the posterior RF was connected with vascular sheaths of the great vessels. The LF was seen developing as a fasciculation of the multilaminar structure in the pararenal space. However, on the posterolateral side of the colon after retroperitoneal fixation, the fusion fascia of the peritoneum could also be identified as LF. CONCLUSIONS: A common sheath for ureters and hypogastric nerves appeared to be likely on the inferior side of the kidney. The LF did not appear to be a primary structure such as the RF, but a result of secondary mechanical stress due to fatty tissue developing earlier along the TF than in the perirenal space. However, the suggested similarity between LF and fusion fascia in the plane occupied was a likely cause for misinterpreting the laminar configurations during surgery.

The uterosacral complex: ligament or neurovascular pathway? Anatomical and histological study of fetuses and adults.

The aim of this study was to define the anatomical relationships of the uterosacral ligament complex (USLC) and to analyze histologically its content. Three fetal and four adult cadavers were used. Anatomical dissections were carried out. Eight fresh biopsies (four fetal and four adult) of the USLC were analyzed histologically and immunohistochemically. Specimens were stained with hematoxylin eosin safran coloration, with anti-nervous cell antibodies (PS 100) and with anti-smooth muscle antibodies (to visualize vessel walls). By removing the visceral pelvic fascia, nervous fibers were found within the USLC forming the hypogastric plexus. Histologically, the USLC contained connective tissue, nervous fibers, sympathetic nodes, vessels, and fatty tissue. No structured ligamentous organization was identified. The uterosacral "ligament" is a "complex" integrating connective tissue as well as nervous and vascular elements. Radical excisions and USLC suspension during pelvic floor reconstructive surgery should be performed with caution in order to preserve pelvic innervation.

Development of the human hypogastric nerve sheath with special reference to the topohistology between the nerve sheath and other prevertebral fascial structures.

Semi-serial sections from the lumbosacral region of nine fetuses (8-25 weeks gestation) were examined to clarify the lumbar prevertebral fascial arrangement. The prevertebral fasciae became evident after 12 weeks of age. After 20 weeks of age, the hypogastric nerve (HGN) was sandwiched by two fascial structures; the ventral fascia which seemed to correspond to the mesorectal fascia, whereas the dorsal fascia corresponded to the presacral fascia. These fasciae or the HGN sheaths extended laterally along the ventral aspects of the great vessels and associated lymph follicles. The ventral fascia is, to some extent, fused with the mesocolon descendens on the left side of the body. Notably, the lateral continuation of these two fasciae also sandwiches the left ureter, but not the right ureter, presumably due to modifications by the left-sided fusion fascia. A hypothetical common sheath for the HGN and ureter (i.e., the ureterohypogastric or vesicohypogastric fascia) might thus be an oversimplification. Before retroperitoneal fixation, the morphology of the peritoneal recess along the mesocolon descendens and mesosigmoid suggested interindividual differences in location, shape, and size. Therefore, in adults the ease of surgical separation of the rectum and left-sided colon from the HGN seems to depend on interindividual differences in the development of the embryonic peritoneal recess. On the caudal side of the second sacral segment, fascial structures were restricted along and around the HGN, pelvic splanchnic nerve, and pelvic plexus. The rectal lateral ligament thus seems to represent a kind of migration fascia formed by mechanical stress.

The contribution of the levator ani nerve and the pudendal nerve to the innervation of the levator ani muscles; a study in human fetuses.

OBJECTIVES: The contributions of the pudendal and levator ani nerves to the innervation of the levator ani muscle (LAM) are disputed. Because of the relatively large size of the nerves in early life, we investigated this issue in human fetuses. METHODS: (Immuno)histochemically stained serial sections of nine human fetuses (9-22 wk of gestation) were investigated. Both the left and right sides of the fetal pelves were studied individually and 3D reconstructions were prepared. RESULTS: The levator ani nerve innervated the LAM in every pelvis, whereas a contribution of the pudendal nerve to the innervation of the LAM could be demonstrated in only 10 pelvic halves (56%). In 10 halves, we observed a communicating nerve branch between the pudendal and levator ani nerves that pierced the pelvic floor between the LAM and the coccygeus muscle. No sex differences were observed, but the innervation pattern did differ between the left and right side of a pelvis. CONCLUSIONS: The LAM often has a dual somatic innervation with the levator ani nerve as its constant and main neuronal supply.

Pelvic plexus contributes ganglion cells to the hindgut enteric nervous system.

The hindgut enteric nervous system (ENS) contains cells originating from vagal and sacral neural crest. In avians, the sacral crest gives rise to the nerve of Remak (NoR) and pelvic plexus. Whereas the NoR has been suggested to serve as the source of sacral crest-derived cells to the gut, the contribution of the pelvic ganglia is unknown. The purpose of this study was to test the hypothesis that the pelvic ganglia contribute ganglion cells to the hindgut ENS. We observed that the quail pelvic plexus develops from neural crest-derived cells that aggregate around the cloaca at embryonic day 5. Using chick-quail tissue recombinations, we found that hindgut grafts did not contain enteric ganglia unless the pelvic plexus was included. Neurofibers extended from the NoR into the intestine, but no ganglion cell contribution from the NoR was identified. These results demonstrate that the pelvic plexus, and not the NoR, serves as the staging area for sacral crest-derived cells to enter the avian hindgut, confirming the evolutionary conservation of this important embryologic process.

Nerve cells and synapses in the human foetal hypogastric nerves.

The hypogastric nerves of a human foetus of 220 mm C-R length (23(rd) week) were investigated with an electron microscope. These nerves were composed mainly of bundles of unmyelinated fibres and single myelinated fibres. Small ganglia and single ganglion cells were observed in the hypogastric nerves. Light and dark cells were found among the nerve cells. The two types of cell differed in the number of ribosomes and the amount rough endoplasmic reticulum. In the period of development investigated protosynapses and mature synapses were observed in the hypogastric nerves.

Development of the myelin sheath of the hypogastric nerves in a human foetus aged 23 weeks.

The formation of the myelin sheath of the human hypogastric nerves was studied by electron microscopy in a foetus of 23 weeks of postovulatory age (220 mm C-R length). In the investigated foetus the hypogastric nerves were mainly composed of bundles of unmyelinated fibres. The myelinated fibres were seen to be at different stages of myelination. Well myelinated fibres had thick compact laminated myelin. The number of myelin lamellae on a single fibre was 22.

Morphometry by computerized three-dimensional reconstruction of the hypogastric plexus of a human fetus.

The histological study of the plexus hypogastricus inferior (hypogastric plexus) of a human fetus does not permit the direct appreciation of its spatial configuration and its complicated relations. Developments in the field of computer science and three-dimensional (3D) reconstruction from serial histological sections have allowed a precise description of its morphometry and relations. The histological sections which were used came from the "Rouvière" collection of the Institute of Anatomy in Paris. A personal computer (IMAC) system of image analysis with reconstruction software was used. Serial pelvic histological sections were directly digitized from the slides. Image treatment and reconstruction were done with manual methods. The 3D reconstruction of the hypogastric plexus, the nerves, the pelvic skeleton and viscera were done. The hypogastric plexus and its topographic relations with the other organs were visualized and studied in three dimensions, and its morphometry was studied. The direct acquisition of the images from the slides allowed excellent high-quality digital images to be obtained. However, manual processing for the reconstruction was time-consuming. At first, the reconstruction of the various pelvic structures was done separately for each organ. Then the structures were visualized all together. Thus, the hypogastric plexus could be examined under various incidences with each organ. The virtual images obtained show new details of the topographic relations and improve knowledge of the precise innervation of the pelvic organs.

[Anatomic and clinical evidence of intrapelvic pudendal nerve and its relation with striated sphincter of the urethra]. / Evidencias anatómicas y clínicas sobre el pudendo intrapélvico y su relación con el esfínter estriado de la uretra.

OBJECTIVE: To demonstrate that somatic innervation of the urethrae striate sphincter is intrapelvic and not through an internal pudendal nerve, an extrapelvic nerve in its entire path. To study the relationship of the pelvic plexus with genito-urinary organs and its surgical implications. METHODS: 6 embryos and 2 fetuses, sliced and stained with techniques suitable for nervous structures were studied. Sequential observation of the pelvic plexus structures and the internal pudendal nerve was carried out using light microscopy. Three-dimensional reconstruction of the two fetuses was performed to study the relationships of these structures with the genito-urinary organs. RESULTS: A nervous branch was identified in the 19, 25 and 30 mm long specimens that started at the internal pudendal nerve and joined the hypogastric ganglion close to the site of pelvic nerves binding. Hypogastric ganglion efferent branches penetrating the striate sphincter after a short descendent run were seen in the 30 and 39 mm embryos. CONCLUSIONS: Our findings confirm that the urethrae striate sphincter received the autonomous and somatic innervation from the pelvic plexus and, therefore, is susceptible to damage during cancer related surgery of the pelvic organs. An improved knowledge of these structures and the use of nerve preservation surgical techniques can reduce the incidence of post-operative incontinence in this type of surgery.

In vivo transplantation of mammalian neural crest cells into chick hosts reveals a new autonomic sublineage restriction.

The study of mammalian neural crest development has been limited by the lack of an accessible system for in vivo transplantation of these cells. We have developed a novel transplantation system to study lineage restriction in the rodent neural crest. Migratory rat neural crest cells (NCCs), transplanted into chicken embryos, can differentiate into sensory, sympathetic, and parasympathetic neurons, as shown by the expression of neuronal subtype-specific and pan-neuronal markers, as well as into Schwann cells and satellite glia. In contrast, an immunopurified population of enteric neural precursors (ENPs) from the fetal gut can also generate neurons in all of these ganglia, but only expresses appropriate neuronal subtype markers in Remak's and associated pelvic parasympathetic ganglia. ENPs also appear restricted in the kinds of glia they can generate in comparison to NCCs. Thus ENPs have parasympathetic and presumably enteric capacities, but not sympathetic or sensory capacities. These results identify a new autonomic lineage restriction in the neural crest, and suggest that this restriction preceeds the choice between neuronal and glial fates.

Origin and topography of the pelvic nerves in human embryos and fetuses.

The pelvic plexus is observed in human embryos at stage 15 and during the last two embryonic weeks the secondary plexuses are formed. The pelvic nerves are found at stage 15 and in embryos they arise from anterior rami of the sacral nerves S2-S4. In fetuses the pelvic nerves originate from the anterior rami S1-S5. Number of pelvic nerves from one anterior ramus varies from 1 to 3.