Hes1 is required for the development of the superior cervical ganglion of sympathetic trunk and the carotid body

Hes1 gene represses the expression of proneural basic helix–loop–helix (bHLH) factor Mash1, which is essential for the differentiation of the sympathetic ganglia and carotid body glomus cells. The sympathetic ganglia, carotid body, and common carotid artery in Wnt1‐Cre/R26R double transgenic mice were intensely labeled by X‐gal staining, i.e., the neural crest origin. The deficiency of Hes1 caused severe hypoplasia of the superior cervical ganglion (SCG). At embryonic day (E) 17.5–E18.5, the volume of the SCG in Hes1 null mutants was reduced to 26.4% of the value in wild‐type mice. In 4 of 30 cases (13.3%), the common carotid artery derived from the third arch artery was absent in the null mutants, and the carotid body was not formed. When the common carotid artery was retained, the organ grew in the wall of the third arch artery and glomus cell precursors were provided from the SCG in the null mutants as well as in wild‐types. However, the volume of carotid body in the null mutants was only 52.5% of the value in wild‐types at E17.5–E18.5. These results suggest that Hes1 plays a critical role in regulating the development of neural crest derivatives in the mouse cervical region. Developmental Dynamics 241:1289–1300, 2012. © 2012 Wiley Periodicals, Inc.     

Comparative embryology of the carotid body

Vertebrate carotid bodies and related structures (branchial arch oxygen chemoreceptors in fishes, carotid labyrinth in amphibians, chemoreceptors in the wall of the common carotid and its branches in birds) develop in embryos when neural crest cells, blood vessels, and nerve fibers from sympathetic and cranial nerve ganglia invade mesenchymal primordia in the wall of the 3rd branchial arch. This review focuses on literature published since the 1970s investigating similarities and differences in the embryological development of 3rd arch oxygen chemoreceptors, especially between mammals and birds, but also considering reptiles, amphibians and fishes.     

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.     

Specific localization of manserin peptide in the rat carotid body

The carotid body, located at the bifurcation of the common carotid artery, is a small sensory organ that detects changes in oxygen concentration and plays a vital role in controlling respiration. Although several molecules, such as neurotransmitters and neuropeptides, are involved in the regulation of the respiratory system, their detailed mechanisms have not been established yet. This study identifies that the presence of manserin, a neuropeptide, in the carotid body may play a crucial role in regulating respiration.The carotid bodies of adult Wistar rats were perfused with paraformaldehyde, and the frozen sections were subjected to immunohistochemical analyses. The carotid body comprises two distinct types of cells, neuron-like glomus cells and glial-like sustentacular cells. We used specific antibodies to distinguish the specific location of manserin in the carotid body, which included a tyrosine hydroxylase-positive antibody for glomus cells and an S100 protein antibody for sustentacular cells. Immunofluorescence analysis revealed that while tiny, round signals were exclusively observed in the cytoplasm of glomus cells, no signals were observed in sustentacular cells.Because manserin is believed to be secreted from precursor proteins by the endoproteolytic processing of a large precursor protein called secretogranin II, manserin secretion systems may exist in the carotid body, and thus, behave as potential regulators of respiration in the carotid body.     

Bilateral carotid body tumor: the role of fine-needle aspiration biopsy in the preoperative diagnosis

Carotid body (CB) is a round to ovoid or flattened structure situated within the adventitia of the common carotid artery bifurcation on both sides of the neck. CB contains two basic types of cells: chief cells (or glomus type 1) and sustentacular cells (glomus type 2). Carotid body tumor (CBT) or paraganglioma arises from the chief cells of the carotid body. The diagnosis of CBT is typically made with radiological studies. Fine needle aspiration biopsy (FNAB) is seldom requested for this purpose due to rare but dreadful reported complications such as hemorrhage and damage to the carotid artery. In this report we discuss the cytological findings of a malignant CBT diagnosed by FNAB in a 22 year-old female.     

Chemoreceptor properties of glomus tissue found in the carotid region of the cat

1. ;Miniglomera' appearing as small masses of tissue with ample vascularization were found around the common carotid artery of the cat. Physiological, gross anatomical and electron microscopic studies were conducted on these tissues.2. The chemosensory function of each ;miniglomus' was evident from the behaviour of the afferent nerve fibres supplying the tissue: afferent responses became more active during asphyxia, when the blood flow through the tissue was reduced or blocked and when cyanide or ACh were applied. The afferent impulses became more infrequent during hyperventilation.3. Sensory frequency response curves constructed against percentage of inhaled O(2) showed that the impulses of single units increased in frequency with lowering of O(2) content of the inhaled gas.4. These miniglomera are innervated by afferent fibres emerging from the nodose ganglion; sometimes these fibres are contained in the aortic or common carotid baroreceptor nerves, but sometimes they emerge as independent nerves. None of the miniglomera are supplied by branches of the sinus nerve.5. The fine structure of the miniglomus is similar to that of the carotid body. The tissue contains two types of cells: glomus cells which contain dense cored granules, and sustentacular cells whose fine processes enclose the former. Membrane densifications occur where glomus cells lie adjacent to one another or where they are contacted by nerve terminals. Nerve fibres are common in the miniglomus but they contact glomus cells less frequently than in the carotid body.     

Chemoreceptor A-fibres in the human carotid body contain tyrosine hydroxylase and neurofilament immunoreactivity.

Previous retrograde tracing studies on rat and guinea-pig showed a projection of sensory tyrosine hydroxylase-immunoreactive neurons to the region of the carotid bifurcation via the carotid sinus nerve. In the present study, focussing on the sensory innervation of the human carotid body, antisera to tyrosine hydroxylase and other catecholamine synthesizing enzymes were applied for an immunohistochemical investigation of carotid bodies obtained at autopsy. In addition, an array of antisera directed to non-enzyme antigens known to be present in viscero-afferent neurons were incorporated in the study. The glomic lobules consisting of glomus cells and sustentacular cells contained a variable number of enzyme-immunoreactive glomus cells. Arteries were supplied by nerve fibres displaying the full phenotype of sympathetic noradrenergic axons, i.e. immunoreactivity to tyrosine hydroxylase, aromatic-L-amino-acid-decarboxylase and dopamine-beta-hydroxylase. The glomic lobules, however, were densely innervated by tyrosine hydroxylase-immunoreactive axons lacking immunoreactivity to aromatic-L-amino-acid-decarboxylase and dopamine-beta-hydroxylase. These fibres reacted with neurofilament 160kD-antibody but were devoid of immunoreactivity to all neuropeptides tested (calcitonin gene-related peptide, somatostatin, substance P). Ultrastructurally, tyrosine hydroxylase/neurofilament 160kD-immunoreactive axons gave rise to large axonal swellings filled with mitochondria and vesicles, and established extensive contacts to glomus cells. Nerve bundles surrounded by a perineural sheath contained both myelinated (2.0-2.8 microns in diameter) and unmyelinated (0.14-3.0 microns) tyrosine hydroxylase-immunoreactive axons. Most of the unmyelinated immunoreactive axons were running singularly within a Schwann cell-sheath. Judged from the pattern of immunoreactivities as well as their preterminal and terminal ultrastructure, tyrosine hydroxylase-immunoreactive axons innervating glomus cells are of sensory origin. Although final proof by retrograde tracing cannot be presented in man, this conclusion is supported by experimental evidence in laboratory animals. The myelinated immunoreactive axons correspond to chemoreceptor A-fibres whereas the classification of the large unmyelinated immunoreactive axons has yet to be established. The lack of immunoreactivity to the dopamine-synthesizing enzyme, aromatic-L-amino-acid-decarboxylase, in this fibre type does not support the view of dopamine being the primary transmitter of chemoreceptor afferents.     

The Carotid Sinus Nerve—Structure,Function, and Clinical Implications

Interest has been renewed in the anatomy and physiology of the carotid sinus nerve (CSN) and its targets (carotid sinus and carotid body, CB), due to recent proposals of surgical procedures for a series of common pathologies, such as carotid sinus syndrome, hypertension, heart failure, and insulin resistance. The CSN originates from the glossopharyngeal nerve soon after its appearance from the jugular foramen. It shows frequent communications with the sympathetic trunk (usually at the level of the superior cervical ganglion) and the vagal nerve (main trunk, pharyngeal branches, or superior laryngeal nerve). It courses on the anterior aspect of the internal carotid artery to reach the carotid sinus, CB, and/or intercarotid plexus. In the carotid sinus, type I (dynamic) carotid baroreceptors have larger myelinated A-fibers; type II (tonic) baroreceptors show smaller A- and unmyelinated C-fibers. In the CB, afferent fibers are mainly stimulated by acetylcholine and ATP, released by type I cells. The neurons are located in the petrosal ganglion, and centripetal fibers project on to the solitary tract nucleus: chemosensory inputs to the commissural subnucleus, and baroreceptor inputs to the commissural, medial, dorsomedial, and dorsolateral subnuclei. The baroreceptor component of the CSN elicits sympatho-inhibition and the chemoreceptor component stimulates sympatho-activation. Thus, in refractory hypertension and heart failure (characterized by increased sympathetic activity), baroreceptor electrical stimulation, and CB removal have been proposed. Instead, denervation of the carotid sinus has been proposed for the “carotid sinus syndrome.” Anat Rec, 302:575–587, 2019. © 2018 Wiley Periodicals, Inc.     

CO-induced K(+) currents in rat glomus cells are insensitive to light unlike carotid body neural discharge and Vo(O(2))

The hypothesis that the light sensitive properties of CO-induced chemosensory nerve (CSN) discharge and oxygen consumption of the carotid body (CB) were shared by the pre-synaptic glomus cells was tested. The light effect on K(+) currents were measured before and during perfusion of the isolated rat glomus cells with high P(CO) of 550 Torr during nomoxia (P(O(2)approximately equal 100 Torr) at extra-cellular pH 7.0 and intracellular pH 6.8 with HEPES buffer. CO increased the K(+) currents with a left ward shift of the reversal potential, which showed no light effect. Thus the K(+) permeability of the glomus cell membrane were not shared by the light-sensitive CSN discharge of the CB and oxygen consumption in the presence of high P(CO.)     

A comparison of the blood supply of the carotid body in rats and rabbits

Light-microscopic findings regarding the arterial blood supply of the carotid bodies in 145 Wistar rats, in 175 spontaneously hypertensive rats (SHR) and in 14 rabbits were compared. In rats, only one glomic artery was found in each of 637 carotid bifurcations. In 2 bifurcations 2 carotid body arteries supplied the glomus caroticum, the glomic artery being of the muscular type. At the origin of the carotid body artery almost regularly intraarterial cushions were observed in rats. They were also demonstrable at the origin of first or second order branches of the glomic artery. Concerning the origin of the carotid body artery as well as presence and form of the intraarterial cushions some differences between the 2 strains of rats studied were detected. In rabbits the glomic artery was of the elastic type. In 2 out of 28 carotid bifurcations 2 carotid body arteries were found. In all other cases the glomus caroticum was supplied by one artery only. The glomic artery did not terminate in the glomus caroticum in rats as well as in rabbits. Many similarities between the light-microscopic picture of the internal carotid artery and the carotid body artery were observed in rabbits. Therefore a baroreceptor function of the glomic artery seems to be possible.     

ATP causes glomus cell [Ca2+]c increase without corresponding increases in CSN activity

The hypothesis that an increase in intracellular calcium [Ca(2+)](c) in carotid body (CB) glomus cells will cause enhanced afferent carotid sinus nerve (CSN) activities was tested in the rat CB in-vitro with the use of extracellular ATP. ATP caused a dose dependent [Ca(2+)](c) increase in identified glomus cells. A major part of total [Ca(2+)](c) increase (2/3) was due to the [Ca(2+)] influx. The rest of [Ca(2+)](c) increase (1/3) was due to the release of [Ca(2+)] from the endoplasmic reticulum (ER) [Ca(2+)] stores, and it was inhibited by the pretreatment of cells with cyclopiazonic acid (CPA), an intracellular Ca(2+)-ATPase blocker. Suramin, a purinergic P(2) receptor membrane blocker, blocked [Ca(2+)] influx due to ATP in the presence of extracellular [Ca(2+)]. Perfusion with 5 and 10 microM ATP stimulated CSN activities in both normoxia (Nx) and hypoxia (Hx). Above that level, 100 microM ATP induced slight initial stimulation in CSN activities which were subsided subsequently in Nx and partly diminished in Hx, while 500 microM ATP completely inhibited CSN activities in Nx and Hx after a slight initial stimulation. Electrophysiological measurements of the glomus cell membrane potential in the presence of ATP (100 microM) during Nx indicated cellular enhanced outward K(+) current and hyperpolarization, suggesting potential mechanism for the inhibition of CSN activities. Thus, ATP dependent linear increases in [Ca(2+)](c) did not give rise to a corresponding increase in CSN activities, contravening the normally expected increase in CSN activities following [Ca(2+)](c) rise.     

The spatial relationship between type I glomus cells and arteriolar myocytes in the mouse carotid body

The structural relationship between type I glomus cells and the vascular smooth muscle was investigated by electron microscopy in the mouse carotid body. A close apposition (<0.1 μm) between the glomus parenchyma and the neighbouring arterioles was regularly present. Profiles of type I glomus cells were found to be exposed to the vascular smooth muscle without any supporting cell investment. In circumscribed areas of these profiles, type I glomus cells and the vascular smooth muscle cells made contact by fusion of their basal laminae. These glomus-cell-myocyte junctions structurally resemble vascular neuromuscular junctions of sympathetic nerve terminals. In addition to the occurrence of such glomus cell-myocyte contacts, myoendothelial junctions also appeared frequently. On the basis of these observations, it is suggested that type I glomus cells play a role in the regulation of the vascular tone in the carotid body and that a physiological interaction between the endothelial cells, the vascular smooth muscle cells and the type I glomus cells exists.     

Fine structure of the carotid body of the midterm human fetus

Summary The human fetal carotid body was studied using both histochemical and electron microscopic methods. The glomus cells of a mid term fetal carotid body evidently contain catecholamines. This was demonstrated both by formaldehyder-induced fluorescence of the cells and by the presence of typical dense-cored vesicles (diameter 1430–3200 Å) in the cytoplasm of the chief cells. The glomus cells were densely innervated and the synapses found on their surface were probably cholinergic in type, containing agranular synaptic vesicles measuring 400–700 Å in diameter with a few dense-cored vesicles measuring 900 to 1300 Å. Synapses were not found in any other cell type within the glomus caroticum. The prominent feature of the glomus cell cytoplasm was the presence of the dense-cored vesicles. The density of the vesicular core varied only slightly from cell to cell. There were no perceptible differences in vesicular size between the different cells. The glomus cells were mostly surrounded by the processes of the sustentacular cells, which usually also surrounded the capillary walls. No glomus cells were ever found in direct contact with the capillary wall. The capillaries were wide and very numerous over the restricted area of the organ. They formed sinusoidal loops, probably anastomosing with each other. Finally, the features of the fine structure are discussed, correlating the present findings with our knowledge about the adult functional carotic body.     

The districution of biogenic amines in the carotid bifurcation region

1. The distribution of phenolic amines in the carotid body and of adrenergic nerves in the arterial walls of the carotid bifurcation region was investigated histochemically using the phenolic amine-formaldehyde gas condensation reaction, with supplementary electron microscopical examination. Studies were conducted on normal rabbits and on rabbits after superior cervical ganglionectomy.2. The glomus cells of the carotid body contain phenolic amines. The cellular amine persists after long-term sympathetic ganglionectomy.3. The blood vessels supplying the carotid body receive an adrenergic innervation which disappears following superior cervical ganglionectomy.4. The walls of the carotid sinus contain adrenergic nerve fibres derived from the superior cervical ganglion.5. There is no penetration of the arterial tunica media by vasomotor fibres in any part of the carotid bifurcation region.6. Ultimate terminations of adrenergic nerves upon smooth muscle occur across the medio-adventitial border of the common, internal and external carotid arterial walls but only in relation to smooth-muscle cells in the adventitia of the carotid sinus wall.     

The content and localization of catecholamines in the carotid labyrinths and aortic arches of Rana temporaria

1. The catecholamine content of the carotid labyrinth of Rana temporaria was estimated by two different methods, and compared with that of the aortic arch. In both tissues adrenaline was found as the dominant amine, with traces of dopamine and noradrenaline detectable in the labyrinth only.2. Per gram of fresh tissue, the labyrinth usually contained more adrenaline than the aortic arch.3. On microscopic examination, the aortic arch was found to contain nerve fibres which fluoresced after treatment with formaldehyde, whereas within the carotid labyrinth the formaldehyde-induced fluorescence was localized in the cytoplasm of clusters of small cells. This fluorescence resembled that exhibited by glomus cells in the mammalian carotid body.     

A light and electron microscopic study on the embryonic development of the rat carotid body

The embryonic development of the rat carotid body was studied with electron microscopy. In the 11 mm embryo a cell aggregation consisting of undifferentiated cells and unmyelinated nerve fibers appears on the anterior wall of the third branchial artery. Granule-containing cells appear in the 12 mm embryo and continue to increase in number as the cellular aggregation increases in size and becomes separated from the wall of the third branchial artery. Synapse formation and the appearance of fenestrated capillaries occur almost simultaneously at the 17 mm stage. There are two types of synapses, one with membrane densification and vesicles clustered inside the nerve endings, the other with dense material and vesicles inside the granule-containing cells. At the 20 mm stage the undifferentiated cells send enveloping cytoplasmic processes toward adjacent granule-containing cells and the carotid body anlage displays rudimentary lobules.     

Modulatory effects of histamine on cat carotid body chemoreception

Histamine has been proposed to be an excitatory transmitter between the carotid body (CB) chemoreceptor (glomus) cells and petrosal ganglion (PG) neurons. The histamine biosynthetic pathway, its storage and release, as well as the presence of histamine H1, H2 and H3 receptors have been found in the CB. However, there is only indirect evidence showing the presence of histamine in glomus cells, or weather its application produces chemosensory excitation. Thus, we studied the histamine immunocytochemical localization in the cat CB, and the effects of histamine, and H1, H2 and H3 receptor blockers on carotid sinus nerve (CSN) discharge, using CB and PG preparations in vitro. We found histamine immunoreactivity in dense-cored vesicles of glomus cells. Histamine induced dose-dependent increases in CSN discharge in the CB, but not in the PG. The H1-antagonist pyrilamine reduced the CB responses induced by histamine, the H2-antagonists cimetidine and ranitidine had no effect, while the H3-antagonist thioperamide enhanced histamine-induced responses. Present data suggests that histamine plays an excitatory modulatory role in the generation of cat CB chemosensory activity.     

Craniofacial malformations: intrinsic vs extrinsic neural crest cell defects in Treacher Collins and 22q11 deletion syndromes

The craniofacial complex is anatomically the most sophisticated part of the body. It houses all the major sensory organ systems and its origins are synonymous with vertebrate evolution. Of fundamental importance to craniofacial development is a specialized population of stem and progenitor cells, known as the neural crest, which generate the majority of the bone, cartilage, connective and peripheral nerve tissue in the head. Approximately one third of all congenital abnormalities exhibit craniofacial malformations and consequently, most craniofacial anomalies are considered to arise through primary defects in neural crest cell development. Recent advances however, have challenged this classical dogma, underscoring the influence of tissues with which the neural crest cells interact as the primary origin of patterning defects in craniofacial morphogenesis. In this review we discuss these neural crest cell interactions with mesoderm, endoderm and ectoderm in the head in the context of a better understanding of craniofacial malformations such as in Treacher Collins and 22q11 deletion syndromes.