Contribution of the cervical sympathetic ganglia to the innervation of the pharyngeal arch arteries and the heart in the chick embryo

In the chick heart, sympathetic innervation is derived from the sympathetic neural crest (trunk neural crest arising from somite level 10–20). Since the trunk neural crest gives rise to sympathetic ganglia of their corresponding level, it suggests that the sympathetic neural crest develops into cervical ganglia 4–14. We therefore tested the hypothesis that, in addition to the first thoracic ganglia, the cervical ganglia might contribute to cardiac innervation as well. Putative sympathetic nerve connections between the cervical ganglia and the heart were demonstrated using the differentiation markers tyrosine hydroxylase and HNK‐1. In addition, heterospecific transplantation (quail to chick) of the cardiac and trunk neural crest was used to study the relation between the sympathetic neural crest and the cervical ganglia. Quail cells were visualized using the quail nuclear antibody QCPN. The results by immunohistochemical study show that the superior and the middle cervical ganglia and possibly the carotid paraganglia contribute to the carotid nerve. This nerve subsequently joins the nodose ganglion of the vagal nerve via which it contributes to nerve fibers in cardiac vagal branches entering the arterial and venous pole of the heart. In addition, the carotid nerve contributes to nerve fibers connected to putative baro‐ and chemoreceptors in and near the wall of pharyngeal arch arteries suggesting a role of the superior and middle cervical ganglia and the paraganglia of the carotid plexus in sensory afferent innervation. The lower cervical ganglia 13 and 14 contribute predominantly to nerve branches entering the venous pole via the anterior cardinal veins. We did not observe a thoracic contribution. Heterospecific transplantation shows that the cervical ganglia 4–14 as well as the carotid paraganglia are derived from the sympathetic neural crest. The cardiac neural crest does not contribute to the neurons of the cervical ganglia. We conclude that the cervical ganglia contribute to cardiac innervation which explains the contribution of the sympathetic neural crest to the innervation of the chick heart. Anat Rec 255:407–419, 1999. © 1999 Wiley‐Liss, Inc.     

Contribution of the cervical sympathetic ganglia to the innervation of the pharyngeal arch arteries and the heart in the chick embryo.

In the chick heart, sympathetic innervation is derived from the sympathetic neural crest (trunk neural crest arising from somite level 10-20). Since the trunk neural crest gives rise to sympathetic ganglia of their corresponding level, it suggests that the sympathetic neural crest develops into cervical ganglia 4-14. We therefore tested the hypothesis that, in addition to the first thoracic ganglia, the cervical ganglia might contribute to cardiac innervation as well. Putative sympathetic nerve connections between the cervical ganglia and the heart were demonstrated using the differentiation markers tyrosine hydroxylase and HNK-1. In addition, heterospecific transplantation (quail to chick) of the cardiac and trunk neural crest was used to study the relation between the sympathetic neural crest and the cervical ganglia. Quail cells were visualized using the quail nuclear antibody QCPN. The results by immunohistochemical study show that the superior and the middle cervical ganglia and possibly the carotid paraganglia contribute to the carotid nerve. This nerve subsequently joins the nodose ganglion of the vagal nerve via which it contributes to nerve fibers in cardiac vagal branches entering the arterial and venous pole of the heart. In addition, the carotid nerve contributes to nerve fibers connected to putative baro- and chemoreceptors in and near the wall of pharyngeal arch arteries suggesting a role of the superior and middle cervical ganglia and the paraganglia of the carotid plexus in sensory afferent innervation. The lower cervical ganglia 13 and 14 contribute predominantly to nerve branches entering the venous pole via the anterior cardinal veins. We did not observe a thoracic contribution. Heterospecific transplantation shows that the cervical ganglia 4-14 as well as the carotid paraganglia are derived from the sympathetic neural crest. The cardiac neural crest does not contribute to the neurons of the cervical ganglia. We conclude that the cervical ganglia contribute to cardiac innervation which explains the contribution of the sympathetic neural crest to the innervation of the chick heart.     

Distribution of different regions of cardiac neural crest in the extrinsic and the intrinsic cardiac nervous system.

In this study we focused upon whether different levels of postotic neural crest as well as the right and left cardiac neural crest show a segmented or mixed distribution in the extrinsic and intrinsic cardiac nervous system. Different parts of the postotic neural crest were labeled by heterospecific replacement of chick neural tube by its quail counterpart. Quail-chick chimeras (n = 21) were immunohistochemically evaluated at stage HH28+, HH29+, and between HH34-37. In another set of embryos, different regions of cardiac neural crest were tagged with a retrovirus containing the LacZ reporter gene and evaluated between HH35-37 (n = 13). The results show a difference in distribution between the right- and left-sided cardiac neural crest cells at the arterial pole and ventral cardiac plexus. In the dorsal cardiac plexus, the right and left cardiac neural crest cells mix. In general, the extrinsic and intrinsic cardiac nerves receive a lower contribution from the right cardiac neural crest compared with the left cardiac neural crest. The right-sided neural crest from the level of somite 1 seeds only the cranial part of the vagal nerve and the ventral cardiac plexus. Furthermore, the results show a nonsegmented overlapping contribution of neural crest originating from S1 to S3 to the Schwann cells of the cranial and recurrent nerves and the intrinsic cardiac plexus. Also the Schwann cells along the distal intestinal part of the vagal nerve are derived exclusively from the cardiac neural crest region. These findings and the smaller contribution of the more cranially emanating cardiac neural crest to the dorsal cardiac plexus compared with more caudal cardiac neural crest levels, suggests an initial segmented distribution of cardiac neural crest cells in the circumpharyngeal region, followed by longitudinal migration along the vagal nerve during later stages.     

Cells migrating from the neural crest contribute to the innervation of the venous pole of the heart

Cells migrating from the neural crest are known to septate the outflow tract of the developing heart, and to contribute to the formation of the arterial valves, their supporting sinuses, the coronary arteries and cardiac neural ganglia. Neural crest cells have also been suggested to contribute to development of the venous pole of the heart, but the extent and fate of such cells remains unclear. In this study, in the mouse, it is shown that cells from the neural crest contribute to the parasympathetic and, to a lesser extent, the sympathetic innervation of the venous pole of the heart. Nerves within the venous pole of the heart are shown to be of mixed origin, with some being derived from the neural crest, while others have an alternative origin, presumably placodal. The neurons innervating the nodal tissue, which can exert chronotropic effects on cardiac conduction, are shown not to be derived from the neural crest. In particular, no evidence was found to support previous suggestions that cells from the neural crest make a direct contribution to the myocardial atrioventricular conduction axis, although a small subset of these cells do co-localize with the developing left bundle branch. We have therefore confirmed that cells from the neural crest migrate to the venous pole of the heart, and that their major role is in the development of the parasympathetic innervation. In addition, in some embryos, a population of cells derived from the neural crest persist in the leaflets of the atrioventricular valves, but their role in subsequent development remains unknown.     

Development of cranial nerves in the chick embryo with special reference to the alterations of cardiac branches after ablation of the cardiac neural crest

Summary Development of cranial nerve branches in the cardiac region was observed in whole-mount specimens which were stained with a monoclonal antibody, E/C8, after the ablation of the cardiac neural crest. In early embryos, nerve trunks of IX and X were lacking or only poorly developed, while the early development of pharyngeal branch primordia was normal. In day 5 embryos, the nerve trunks of IX–X were present in all the embryos, however; extensive communication was observed between X and XII. On day 6 and later, the spiral pattern of superior cardiac branches was disturbed, as were the blood vessels. Furthermore, the distal branches of XII passed within the superficial layer of cardiac outflow mesenchyme. Vagal branches passed within the deeper layer. There was no apparent change in the development of the sinal branch. Using quail — chick chimeras, it was found that the cardiac neural crest cells formed the Schwann cells of XII, and that they were also associated with the hypobranchial muscle primordium, suggesting that the absence of the cardiac neural crest not only disturbs the development of the cardiac outflow septation, but also affects the normal morphogenesis of the hypobranchial musculature and its innervation. Embryologically, the tongue is located close to the cardiac outflow tract, which is the migration pathway of the cardiac neural crest-derived cells.     

Peripheral development of the avian vagus nerve with special reference to the morphological innervation of heart and lung

Summary The development of the cardio-pulmonary innervation was studied in whole-mount specimens of chick embryos stained with the anti-neurofilament protein (NFP) antibody. From the morphological point of view, vagal branches could be classified into two categories, i.e., the branchial branches primarily related to the pharnygeal arches, and intestinal arborization derivatives which are associated primarily with the primitive gut. The former consisted of the superior cardiac branch innervating the truncus arteriosus of the heart, and the latter, the sinal branch, pulmonary branches as well as recurrent nerve and the other intestinal branches. The superior cardiac branch at first develops as a pair of branchial branches which passes into the truncus arteriosus at stage 25, and later rotates along the aortic arch 6, thus making an asymmetrical configuration by stage 27. The sinal branch is a medial branch which first develops at stage 24. It arises from the junction of each intestinal arborization in close association with the pulmonary branch.     

Enteric nervous system development: analysis of the selective developmental potentialities of vagal and sacral neural crest cells using quail-chick chimeras

The majority of the enteric nervous system (ENS) is derived from vagal neural crest cells (NCC). For many years, the contribution from a second region of the neuraxis (the sacral neural crest) to the ENS has been less clear, with conflicting reports appearing in the literature. To resolve this longstanding issue, we documented the spatiotemporal migration and differentiation of vagal and sacral-derived NCC within the developing chick embryo using quail-chick grafting and antibody labelling. Results showed that vagal NCC colonised the entire length of the gut in a rostrocaudal direction. The hindgut, the region of the gastrointestinal tract most frequently affected in developmental disorders, was found to be colonised in a complex manner. Vagal NCC initially migrated within the submucosa, internal to the circular muscle layer, before colonising the myenteric plexus region. In contrast, sacral NCC, which colonised the hindgut in a caudorostral direction, were primarily located in the myenteric plexus region from where they subsequently migrated to the submucosa. We also observed that sacral NCC migrated into the hindgut in significant numbers only after vagal-derived cells had colonised the entire length of the gut. This suggested that to participate in ENS formation, sacral cells may require an interaction with vagal-derived cells, or with factors or signalling molecules released by them or their progeny. To investigate this possible inter-relationship, we ablated sections of vagal neural crest (NC) to prevent the rostrocaudal migration of ENS precursors and, thus, create an aganglionic hindgut model. In the same NC ablated animals, quail-chick sacral NC grafts were performed. In the absence of vagal-derived ganglia, sacral NCC migrated and differentiated in an apparently normal manner. Although the numbers of sacral cells within the hindgut was slightly higher in the absence of vagal-derived cells, the increase was not sufficient to compensate for the lack of enteric ganglia. As vagal NCC appear to be more invasive than sacral NCC, since they colonise the entire length of the gut, we investigated the ability of transplanted vagal cells to colonise the hindgut by grafting the vagal NC into the sacral region. We found that when transplanted, vagal cells retained their invasive capacity and migrated into the hindgut in large numbers. Although sacral-derived cells normally contribute a relatively small number of precursors to the post-umbilical gut, many heterotopic vagal cells were found within the hindgut enteric plexuses at much earlier stages of development than normal. Heterotopic grafting of invasive vagal NCC into the sacral neuraxis may, therefore, be a means of rescuing an aganglionic hindgut phenotype.     

Distribution of antigen epitopes shared by nerves and the myocardium of the embryonic chick heart using different neuronal markers

We examined which neuronal elements and nonneuronal tissues in the embryonic myocardium are stained with antibodies traditionally used for staining nerve tissue. Furthermore, we studied whether nonneuronal myocardial staining was confined to regions determining initial nerve entry points and development of cardiac ganglia. The third focus was whether nerves preferentially distribute in regions of the conduction system. Different neuronal markers were used such as the HNK-1 antibody against neural crest and nerve tissue, Tyrosine Hydroxylase antibody (TH) against putative sympathetic nerve tissue, anti-GFAP against glia cells, antibodies against phosphorylated neurofilaments DO170, RMO270, 3A10, and RT97, and finally the antibody Snap25 against a synaptic protein. Chick embryonic hearts between stage HH25-44 where immunohistochemically evaluated. Transient HNK-1 staining in the basal region of the heart coincided with ingrowing vagal branches and crest-derived neuronal precursor cells seeding the region of the atrioventricular sulcus, suggesting a role for HNK-1 in the homing of the parasympathetic plexus. Transient TH staining was confined to regions of the atrial myocardium coincident with the localization of the few early TH-positive nerve fibers before stage HH40, whereas the second wave of TH-positive nerve fibers at HH42 was mainly localized around myocardial coronary arteries. This transient myocardial TH staining might be involved in early emergence of the catecholaminergic phenotype, while coronary arteries or blood borne factors might be involved in later differentiation. Some myocardial expression, not related with initial nerve ingrowth, using Snap25, TH, HNK-1, DO170, and RMO270 was confined to regions of the ventricular conduction system. HNK-1 is the only marker staining the region of the putative sinoatrial node. Just before hatching nerve fibers, including TH-positive nerve fibers, are uniformly distributed throughout the myocardium, without being specifically confined to regions containing the conduction system or coronary arteries.     

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.     

Migration and distribution of circumpharyngeal crest cells in the chick embryo. Formation of the circumpharyngeal ridge and E/C8+ crest cells in the vertebrate head region.

The cardiac neural crest is located in a transitional area on the neuraxis between trunk and cephalic regions and gives rise to both the dorsolateral and ventrolateral crest cell populations. Around stage 18 of chick development, a mass of E/C8+ cells surrounds the postotic pharyngeal arches and forms a crescent-shaped arch, termed the circumpharyngeal ridge. Using immunohistochemistry and quail-chick chimeras, it was determined that the E/C8+ cell mass located in the circumpharyngeal ridge derives from the dorsolateral component of the cardiac neural crest. The ventrolateral cell population of the cardiac crest is located more medially and shows long-persistent HNK-1 immunoreactivity dorsolateral to the foregut. The crest cells that populate the gut arise from the caudal portion of the circumpharyngeal crest and are always located caudal to the caudal-most pharyngeal ectomesenchyme. Circumpharyngeal crest cells continuously populate the pharyngeal arch ectomesenchyme and enteric nervous system on the lateral side of the foregut wall, as well as the hypoglossal pathway which develops within the ventral portion of the circumpharyngeal ridge. E/C8 and HNK-1 immunoreactivity are associated with the cells migrating via the dorsolateral (circumpharyngeal) and ventrolateral pathways, respectively, with one exception: there is a population of putative crest cells along the proximal course of the vagal intestinal branch that shows both immunoreactivities around stage 20. DiI labeling of the cells in the circumpharyngeal ridge suggests that the cells are contributed from the circumpharyngeal ridge to this population. Thus, the distribution of the circumpharyngeal crest cells and their derivatives coincides with the peripheral branch distribution of the cranial nerves IX, X, and XII, whose development is selectively affected in the absence of the cardiac neural crest, the source of the circumpharyngeal crest.     

Migration and distribution of circumpharyngeal crest cells in the chick embryo

The cardiac neural crest is located in a transitional area on the neuraxis between trunk and cephalic regions and gives rise to both the dorsolateral and ventrolateral crest cell populations. Around stage 18 of chick development, a mass of E/C8⁺ cells surrounds the postotic pharyngeal arches and forms a crescent-shaped arch, termed the circumpharyngeal ridge. Using immunohistochemistry and quail-chick chimeras, it was determined that the E/C8⁺ cell mass located in the circumpharyngeal ridge derives from the dorsolateral component of the cardiac neural crest. The ventrolateral cell population of the cardiac crest is located more medially and shows long-persistent HNK-1 immunoreactivity dorsolateral to the foregut. The crest cells that populate the gut arise from the caudal portion of the circumpharyngeal crest and are always located caudal to the caudalmost pharyngeal ectomesenchyme. Circumpharyngeal crest cells continuously populate the pharyngeal arch ectomesenchyme and enteric nervous system on the lateral side of the foregut wall, as well as the hypoglossal pathway which develops within the ventral portion of the circumpharyngeal ridge. E/C8 and HNK-1 immunoreactivity are associated with the cells migrating via the dorsolateral (circumpharyngeal) and ventrolateral pathways, respectively, with one exception: there is a population of putative crest cells along the proximal course of the vagal intestinal branch that shows both immunoreactivities around stage 20. Dil labeling of the cells in the circumpharyngeal ridge suggests that the cells are contributed from the circumpharyngeal ridge to this population. Thus, the distribution of the circumpharyngeal crest cells and their derivatives coincides with the peripheral branch distribution of the cranial nerves IX, X, and XII, whose development is selectively affected in the absence of the cardiac neural crest, the source of the circumpharyngeal crest.© Willey-Liss, Inc.     

人膈肌内神经分支分布

目的：探讨人膈的神经支配和肌内神经分支分布特点。方法：改良Sihler’s肌内神经分支染色法。结果：(1)一侧膈神经入肌后一般分为3—4支(3支型4例,4支型2例),1支(前支)向前内侧走行,支配胸部,1～2支(前外侧支)向外侧走行,支配肋部,最后1支(后支)最粗大,向后下走行,分为后外侧支和后脚支,分别支配膈中心腱外侧叶后外侧的肋部和腰部,各级神经的分支在肌束中部密集排列成神经丛。(2)6例标本均未发现左侧或右侧膈神经越过中线至对侧。(3)2例带肋间肌的标本肉眼未见有肋间神经分支进入膈。结论：(1)左、右膈神经分布于膈,未见左右侧膈神经重叠支配和优势支配。(2)膈神经的终末分支在肌束中部密集排列形成似“肾形”的神经丛带。     

Vagal neural crest cell migratory behavior: a transition between the cranial and trunk crest

Migration and differentiation of cranial neural crest cells are largely controlled by environmental cues, whereas pathfinding at the trunk level is dictated by cell-autonomous molecular changes owing to early specification of the premigratory crest. Here, we investigated the migration and patterning of vagal neural crest cells. We show that (1) vagal neural crest cells exhibit some developmental bias, and (2) they take separate pathways to the heart and to the gut. Together these observations suggest that prior specification dictates initial pathway choice. However, when we challenged the vagal neural crest cells with different migratory environments, we observed that the behavior of the anterior vagal neural crest cells (somite-level 1-3) exhibit considerable migratory plasticity, whereas the posterior vagal neural crest cells (somite-level 5-7) are more restricted in their behavior. We conclude that the vagal neural crest is a transitional population that has evolved between the head and the trunk.     

The Contribution of the Left Phrenic Nerve to Innervation of the Esophagogastric Junction

The contribution of the left phrenic nerve to innervation of the esophagogastric junction. The esophagogastric junction is part of the barrier preventing gastroesophageal reflux. We have investigated the contribution of the phrenic nerves to innervation of the esophagogastric junction in humans and piglets by dissecting 30 embalmed human specimens and 14 piglets. Samples were microdissected and nerves were stained and examined by light and electron microscopy. In 76.6% of the human specimens, the left phrenic nerve participated in the innervation of the esophagogastric junction by forming a neural network together with the celiac plexus (46.6%) or by sending off a distinct phrenic branch, which joined the anterior vagal trunk (20%). Distinct left phrenic branches were always accompanied by small branches of the left inferior phrenic artery. In 10% there were indirect connections with a distinct phrenic nerve branch joining the celiac ganglion, from which celiac plexus branches to the esophagogastric junction emerged. Morphological examination of phrenic branches revealed strong similarities to autonomic celiac plexus branches. There was no contribution of the left phrenic nerve or accompanying arteries from the caudal phrenic artery in any of the piglets. The right phrenic nerve made no contribution in any of the human or piglet samples. We conclude that the left phrenic nerve in humans contributes to the innervation of the esophagogastric junction by providing ancillary autonomic nerve fibers. Experimental studies of the innervation in pigs should consider that neither of the phrenic nerves was found to contribute. Clin. Anat. 33:265–274, 2020. © 2019 Wiley Periodicals, Inc.     

The innervation of the umbilical vein in human embryos and fetuses

In human embryos and fetuses, a small bundle of nerve fibers from the anterior and posterior vagal trunks descends between the layers of the hepatogastric ligament. These fibers pass to the region of the junction of the umbilical vein with the ductus venosus. At this junction, there is a slight thickening of the muscular wall. Nerve fibers pass to this junction and the proximal portion of the umbilical vein. Fibers from the posterior vagal trunk follow the left gastric artery to the celiac plexus. Fibers from this plexus follow the hepatic artery into the lesser omentum and along the portal vein to the liver. Continuing along the left branch of the portal vein, fibers reach the proximal portion of the umbilical vein and its junction with the ductus venosus. Ganglion cells were observed along the course of vagus nerve fibers to the umbilical vein. In embryos these cells were observed on the lower end of the anterior vagal trunk near the attachment of the upper end of the lesser omentum to the lower end of the esophagus. In older fetuses they were found in a small ganglion in the connective tissue surrouding the distal end of the ductus venosus.     

The hyaloid vascular system of the pig

Summary The hyaloid vascular system of the pig was studied from 4 weeks of gestation until 2 weeks after birth by means of semithin sections and vascular corrosion casts. The vascular tunic of the lens is supplied by the posterior lens branches of the hyaloid artery (at the posterior lens pole), by the intermediate lens branches of the proper hyaloid arteries (at the lens equator) and by the anterior lens branches of the radial iridial arteries (at the anterior lens pole). Venous drainage takes place via the venous lens branches which leave the lens anteriorly and drain into the radial iridial veins. Regression of the vascular tunic of the lens occurs during the second half of fetal life and is nearly completed in the first postnatal days. The involution first affects the proper hyaloid arteries and their intermediate lens branches. Subsequently, the posterior lens branches regress, whereas the anterior lens branches in the pupillary membrane disappear in the perinatal period only.Abbreviations 1 Hyaloid artery - 2 proper hyaloid arteries - 3 annular plexus - 4 posterior lens branches - 5 intermediate lens branches - 6 major arterial circle of the iris - 7 radial iridial arteries - 8 anterior lens branches - 9 venous lens branches - 10 radial iridial veins - 11 retinal blood vessel - C Cornea - CP ciliary processes - I iris - L lens - ON optic nerve - PM pupillary membrane - R retina - VTL vascular tunic of the lens. White bar in Figs. 4–16=1 mm     

The nerve branches to the external anal sphincter: the macroscopic supply and microscopic structure

The study was performed using 45 pelvic half section specimens (41 fetal ones and four adults). The macroscopic dissection followed the nerve branches from their spinal roots up to the external anal sphincter. Three nerve branches were found: the anterior ramus arising from the external perineal nerve, the inferior rectal nerve and an independent posterior branch. The anterior and the inferior rectal nerve branches always emerged from the pudendal plexus. The posterior branch arising either from S4 or from the inferior rectal nerve was only found in (31%) of our cases. Five anatomical distributions are described, percentages of every type notified. The fibre content of these nerve bundle branches was evaluated through histological sections using Heidenhain's azan stain and Luxol fast blue. The branches consisted of 2,896 to 2,137 fibres, 20% of them being unmyelinated and 80% containing various myelinated fibres. The nomenclature of these nerve branches has to be debated. The terms of anterior, middle and posterior anal nerves seem more suitable.