Three dimensional observation of the nerve fibers along the cerebral blood vessels

Three dimensional observation of the nerve fibers along the cerebral blood vessels was investigated by scanning electron microscopy. Electron probe X-ray microanalysis was also performed in the cerebral blood vessels treated with peroxidase-antiperoxidase immunohistochemistry intensified by nickel ammonium sulfate. Nerve fibers (2-8 microns in diameter) formed a plexus on the outer surface of the adventitia. After branching, the nerve fibers penetrated the blood vessel adventitia. Substance P-immunoreactive nerve fibers showed a meshwork pattern in the outer layer of the adventitia, and vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers revealed a spiral running pattern in the inner layer of the adventitia. Taken together with previous studies, these findings suggest that substance P nerve fibers in the cerebral arteries may not be related to arterial dilatation or constriction, but VIP nerve fibers may be vasodilative.     

Calcitonin gene-related peptide (CGRP): perivascular distribution and vasodilatory effects

The distribution of perivascular nerve fibers displaying calcitonin gene-related peptide (CGRP) immunoreactivity and the effect of CGRP on vascular smooth muscle were studied in the guinea-pig. Perivascular CGRP fibers were seen in all vascular beds. Generally, they were more numerous around arteries than veins. Small arteries in the respiratory tract, gastrointestinal tract and genitourinary tract had numerous CGRP fibers. The gastroepiploic artery in particular received a rich supply of such fibers. Coronary blood vessels had a moderate supply of CGRP fibers. In the heart, a moderate number of CGRP fibers was seen running close to myocardial fibers. The atria had a richer supply than the ventricles. Numerous CGRP immunoreactive nerve cell bodies and nerve fibers were seen in sensory (trigeminal, jugular and spinal dorsal root) ganglia. Sequential or double immunostaining with antibodies against substance P and CGRP suggested co-existence of the two peptides in nerve cell bodies in the ganglia and in perivascular fibers. In agreement with previous findings CGRP turned out to be a strong vasodilator in vitro as tested on several blood vessels (e.g. basilar, gastroepiploic and mesenteric arteries). Conceivably, perivascular CGRP/SP fibers have a dual role as regulator of local blood flow and as carrier of sensory information.     

Three-dimensional cytoarchitecture of the media of arteriovenous anastomoses in the rabbit ear

Arteriovenous anastomoses in the rabbit ear were examined with scanning electron microscopy to elucidate the structural differentiation of the media of the shunt. Arterial, intermediate, and venous segments in the shunt and two layers of the media in the intermediate segment were differentiated based on cell shape and cell organization. In the arterial segment, smooth muscle cells were spindle-shaped, either elongated or short, with a few branches, and were arranged circularly or diagonally with respect to the vessel's long axis. There were also stellate muscle cells with radiating processes. In the intermediate segment, the smooth muscle cells of the outer layer of the media were also arranged circularly and resembled the elongated cells in the arterial segments, but they were more irregular in shape and had more processes than those of the arterial segment. The epithelioid cells of the inner layer of the media were oval or polygonal and oriented irregularly with respect to the vessel's long axis, clustering to form longitudinal plicae. The smooth muscle cells of the venous segment were flat with many lateral processes and formed a thin, discontinuous layer. The smooth muscle cells in the arterial segment and those of the outer layer of the intermediate segment exhibited a highly rugged surface texture, indicating their strong contractility; the epithelioid cells and the smooth muscle cells in the venous segment exhibited a generally smooth surface, indicating less contractility. The intermediate segments were supplied with a dense nerve plexus. The intermediate segments, therefore, may be actively involved in the regulation of blood flow under neuronal influence.     

Three dimensional observation of the nerve fibers along the cerebral blood vessels

Summary Three dimensional observation of the nerve fibers along the cerebral blood vessels was investigated by scanning electron microscopy. Electron probe X-ray microanalysis was also performed in the cerebral blood vessels treated with peroxidase-antiperoxidase immunohistochemistry intensified by nickel ammonium sulfate.Nerve fibers (2–8 m in diameter) formed a plexus on the outer surface of the adventitia. After branching, the nerve fibers penetrated the blood vessel adventitia. Substance P-immunoreactive nerve fibers showed a meshwork pattern in the outer layer of the adventitia, and vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers revealed a spiral running pattern in the inner layer of the adventitia. Taken together with previous studies, these findings suggest that substance P nerve fibers in the cerebral arteries may not be related to arterial dilatation or constriction, but VIP nerve fibers may be vasodilative.     

Ultrastructure of venules in the cat brain

Summary Intracerebral venules of the cat were examined to establish criteria for a distinct separation between the venous and arterial system, and to characterize, in greater detail, the mural construction of individual venules. The intracerebral venules were compared with those of other organs. Venules do not have a vascular wall composed clearly of endothelium, media, and adventitia, as is characteristic of arteries and arterioles. The venous endothelium has a similar structure to that of capillaries. The periendothelial cells of the venule differ in shape depending on the vascular diameter. The number of periendothelial cell processes in postcapillary venules increases progressively. Segments in which the basal lamina of the endothelium merges with that of the glia cover a smaller portion of the circumference than in venous capillary loops. In collecting venules, the endothelium is almost completely enveloped by periendothelial cells which have a larger number of filaments. There are no typical smooth muscle cells in the intracerebral venules. The perivascular space becomes wider in collecting venules, contains adventitial cells, phagocytes and a great number of collagen fibers.     

Plasticity of endothelial cells during arterial-venous differentiation in the avian embryo

Remodeling of the primary vascular system of the embryo into arteries and veins has long been thought to depend largely on the influence of hemodynamic forces. This view was recently challenged by the discovery of several molecules specifically expressed by arterial or venous endothelial cells. We here analysed the expression of neuropilin-1 and TIE2, two transmembrane receptors known to play a role in vascular development. In birds, neuropilin-1 was expressed by arterial endothelium and wall cells, but absent from veins. TIE2 was strongly expressed in embryonic veins, but only weakly transcribed in most arteries. To examine whether endothelial cells are committed to an arterial or venous fate once they express these specific receptors, we constructed quail-chick chimeras. The dorsal aorta, carotid artery and the cardinal and jugular veins were isolated together with the vessel wall from quail embryos between embryonic day 2 to 15 and grafted into the coelom of chick hosts. Until embryonic day 7, all grafts yielded endothelial cells that colonized both host arteries and veins. After embryonic day 7, endothelial plasticity was progressively lost and from embryonic day 11 grafts of arteries yielded endothelial cells that colonized only chick arteries and rarely reached the host veins, while grafts of jugular veins colonized mainly host veins. When isolated from the vessel wall, quail aortic endothelial cells from embryonic day 11 embryos were able to colonize both host arteries and veins. Our results show that despite the expression of arterial or venous markers the endothelium remains plastic with regard to arterial-venous differentiation until late in embryonic development and point to a role for the vessel wall in endothelial plasticity and vessel identity.     

Prostacycin production by arterialized autogenous venous grafts in dogs

Arteries are capable of producing significantly larger quantities of protacyclin than are veins. To test the hypothesis, whether prostacyclin production by the vessel wall is related to blood pressure and flow, we measured the amounts of PGI₂ released and synthesized by venous segments transplanted for 6 weeks into the arterial circulation. These results were compared with the production of prostacyclin by normal veins and arteries. In 20 dogs a segment of jugular vein was interposed into the carotid system; a sham dissection was done on the opposite side. “Arterialized” vein grafts showed prominent intima lined by endothelium, medial smooth muscle cell proliferation and fibrotic proliferation in adventitia. Spontaneous and arachidonic acid- stimulated prostacyclin production (measured by radioimmunoassay for 6-keto-PGF_1α) was not significantly different between arterialized venous autografts and jugular veins. Significantly larger amounts of prostacyclin were synthesized by the carotid artery. Thus, histologic changes and rheologic effects occurring in vein grafts transposed to the arterial site do not affect prostacyclin production.     

Distribution and dilatory effect of vasoactive intestinal polypeptide (VIP) in human cerebral arteries

Vasoactive intestinal polypeptide (VIP)-containing nerve fibers were demonstrated in human pial arteries by immunocytochemistry. Fine varicose fibers were located in the adventitia close to the media layer. Measurements by radioimmunoassay revealed concentrations of VIP between 0.7 and 2.7 pmol/g in the major arteries at the base of the brain, obtained at autopsy. Isolated human pial arteries, obtained in conjunction with neurosurgery, relaxed in a concentration-dependent manner upon administration of VIP. The relaxation of the vessels amounted to 57 +/- 9% of the contraction elicited by prostaglandin F2 alpha (2.5 microM) with an EC50 value of (8.5 +/- 1.2) X 10(-9) M.     

VIP-immunoreactivity in the skin of various mammals: immunohistochemical, radioimmunological and experimental evidence for a dual localization in cutaneous nerves and merkel cells

In the present study VIP-immunoreactive (IR) nerve fibers were found in the skin of several mammalian species (cat, dog, pig and man). They supplied predominantly the arteries and arterial portions of arteriovenous anastomoses. Far fewer VIP-IR nerve fibers innervated veins and arterioles. Capillaries were supplied by VIP-IR fibers only in sweat and Meibomian glands. Some non-vascular VIP-IR nerve fibers were seen in contact with dermal smooth muscle strands. In eccrine sweat glands and in Meibomian glands VIP-IR fibers were targeting glandular cells. In addition, VIP-IR nerve fibers innervated the upper parts of facial hair follicles. In non-neuronal localization VIP-IR occurred in Merkel cells in all species and sites, while the intraepidermal axons consistently were not VIP-IR. Radioimmunoassay of different skin regions of cats also suggested both a neuronal and a Merkel cell origin of VIP-IR. Under physiological conditions VIP which is released from its neuronal and non-neuronal cutaneous pools may have an impact on thermoregulation by influencing blood flow and sweat production. It may also modulate axon-endings in Merkel cell-axon complexes and hair follicle receptors. Under pathological conditions an enhanced release of cutaneous VIP may lead to local inflammatory processes partly mediated via release of histamine from cutaneous mast cells.     

Innervation of the arteriovenous anastomoses in the dog tongue

Summary Profiles of nerve plexuses in the arteriovenous anastomoses of the dog tongue were investigated by both transmission and scanning electron microscopy. Three-dimensional morphology of the vascular nerves was examined after removal of the connective tissue components by the HCl-hydrolysis method. Tight bending and a rich nerve supply were the most characteristic features of the anastomosing channels. The tunica media consisted of an outer circular layer of typical smooth-muscle cells and an inner region containing longitudinal plicae of ramified smoothmuscle cells. The tunica adventitia was exclusively occupied by nerve bundles; fibroblasts were poorly developed. Numerous nerve bundles of variable size were coiled around the anastomosing channels, and occasional bundles ran crosswise over the U-shaped bent vessels.     

The innervation and fine structure of paraneuronic cells in an amphibian pulmonary artery

Summary The pulmonary artery of Bufo marinus contains large numbers of bipolar cells situated in the tunica adventitia and in the outer layers of the media. These cells show a bright green-yellow fluorescence (emission spectra 485 nm) after formaldehyde pre-treatment suggesting that they contain a primary monoamine. The most characteristic fine-structural feature of these cells is the presence of numerous dense-cored vesicles (80—300 nm diameter) in their cytoplasm. The cells are in close contact (20 nm gap) with both agranular and granular nerve fibres. Both EM-cytochemical and formaldehyde-induced fluorescence tests indicate that the granule-containing nerve fibres are adrenergic. The agranular nerve fibres form discrete synaptic contacts with pre-and post-synaptic membrane thickenings on the cells. This was never observed with respect to the adrenergic fibres. Each process of the cells is about 45 m long. The processes do not bear any special relationship to either vessels of the arterial vasa vasorum or medial smooth muscle cells. Their location in the wall of the artery suggests that they are functionally significant with respect to activity of the arterial media.     

Structural alterations of blood vessels in hypertensive rats

Vascular changes in the mesenteric arteries were examined in three animals models for human essential hypertension. These models are: spontaneously hypertensive rats, which develop hypertension with age; Dahl model of genetic, salt-dependent hypertensive rats; and deoxycorticosterone-salt hypertensive rats. Morphometric measurements of the arterial wall components (e.g., endothelium, media) were carried out in the elastic arteries, muscular arteries, and arteriolar vessels from the mesenteric bed. The observed changes were correlated with the stages of hypertension development and the effect of antihypertension therapy, including sympathectomy. Specific emphasis was made to determine whether the changes observed were primary in nature, and related to the causes of hypertension, or they were secondary adaptive changes. A comparison of the three models showed that common changes in the intima, media, and adventitia were present in the three models. Alterations in the endothelium (e.g., enlargement of subendothelial space, necrotic changes), adventitia (collagen increase), and hypertrophy of the smooth muscle cells are secondary adaptive changes, because these changes occur subsequent to the development of hypertension, and antihypertensive therapy also prevent these changes from taking place. In contrast, hyperplasia of the smooth muscle cells is a primary change, because it occurs prior to the onset of hypertension. Functionally, alteration in the media is probably the most important change, because it can cause hyperreactivity of the arteries in response to stimulation. Damage to the endothelial cells may play a role in the maintenance of hypertension during the later phase. Alteration in adventitia is a passive change, which does not appear to have a major role in hypertension.(ABSTRACT TRUNCATED AT 250 WORDS)     

The vasculature of the rat penis: a scanning electron microscopic and histologic study.

As a basis for understanding the mechanism of erection in an animal model frequently used in research in reproductive biology, the angioarchitecture of the penis of the rat has been described using scanning electron microscopy. Study of the penile vasculature of the rat indicates that the corpora cavernosa penis and the corpus spongiosum are independent erectile tissues, each with its own arterial and venous vessels. The large vascular spaces and abundant smooth muscle of the penile crura are compatible with its role in regulating blood flow to more distal penile tissues. Helicine arteries of the crura, but not the parent deep penile artery or arteries elsewhere, have muscular cushions in their walls. The venous drainage of the penile crura is via subtunical veins which are thought to be compressed during erection to elevate pressure within the penis. Large, paired cavernous veins drain the shaft of the penis. A unique method for inhibiting blood flow from the penis is indicated by the division of the cavernous veins into smaller channels prior to joining the subtunical venous plexus. Erectile tissue in the bifid origins of the corpus spongiosum has abundant cavernous muscle, while in the remainder of the corpus spongiosum little smooth muscle lines the cavernous spaces. The cavernous spaces on either side of the urethra coalesce to form vessels, each of which communicates with cavernous spaces in the glans. In addition, a bypass of the glans is effected by communication of these vessels directly with the deep dorsal vein. The apparent absence of muscular pads in vessels of the spongiosum, the relative paucity of cavernous smooth muscle, and the ample venous drainage provided by the deep dorsal vein may account for the lack of a venous occlusive mechanism similar to that of the corpora cavernosa penis.     

Observations on the fine structure of the baroreceptors and adrenergic innervation of the guinea-pig carotid sinus

We examined the fine structure of the baroreceptors and the adrenergic innervation of the guinea-pig carotid sinus. The tunica adventitia contained many nerve bundles whose perineuria enclosed unmyelinated nerve fibers, alone or together with myelinated nerve fibers. Baroreceptors, which lay close to elastic and collagen fibers in the adventitia and media, were surrounded by “terminal” cells with ultrastructural features characteristic of Schwann cells and contained inclusions of various types. Morphologic features of the baroreceptors included densely packed mitochondria, osmiophilic lamellated and homogeneous bodies, clear and granular vesicles, lamellar systems, glycogen granules, neurofilaments, neurotubuli, and vacuolated mitochondria. In animals that had been treated with 6-hydroxydopamine, occasional electrondense endings (or fibers) were observed in the adventitial layer. The baroreceptors in the guinea-pig carotid sinus appear to have most of the morphologic features reported for other species.     

Distribution of adrenomedullin-containing perivascular nerves in the rat mesenteric artery

Distribution of adrenomedullin (AM)-containing perivascular nerve fibers was studied in rat mesenteric arteries. Many fibers containing AM-like immunoreactivity (LI) were observed in the adventitia. AM-LI fibers were abolished by cold storage denervation or capsaicin but not 6-hydroxydopamine. Double immunostainings showed colocalization of AM-LI with calcitonin gene-related peptide (CGRP)-LI. The dorsal root ganglia had many AM-positive cells and AM mRNA detected by RT-PCR. Electron microscopy study revealed high proportions of immunogold labeling for AM and colocalization of both AM-LI and CGRP-LI in unmyelinated nerve axons. These results suggest that AM-containing perivascular nerves are distributed in the rat mesenteric artery.     

Subset of cells immunopositive for neurokinin-1 receptor identified as arterial interstitial cells of Cajal in human large arteries

In the adventitia of large arteries, dendritic cells are located between nerve fibers, some of which contain substance P. The aim of the present study was to examine whether neurokinin 1 receptor (NK-1R) was expressed by dendritic cells in the arterial wall. Parallel sections of aortic and carotid artery segments were immunostained with anti-NK-1R and cell-type-specific antibodies. Dendritic cells in the arterial wall expressed NK-1R, albeit at a low level. Other cells, which intensely expressed NK-1R, were located along the border between the media and adventitia. They did not co-express any dendritic cell markers, including fascin, CD1a, S100, or Lag-antigen, and were negative for CD68, CD3, and mast cell tryptase. These NK-1R⁺ cells were laser-capture microdissected and studied by means of electron-microscopic analysis. The microdissected cells were in direct contact with nerve endings, and their ultrastructure was typical of the interstitial cells of Cajal present in the gastrointestinal tract. Further systematic electron-microscopic analysis revealed that the cells displaying the features typical of interstitial cells of Cajal were a basic element of the human arterial wall architectonics. Arterial interstitial cells of Cajal were negative for c-kit but they expressed vasoactive intestinal peptide receptor 1 (VIPR1). Destructive alterations of contacts between arterial interstitial cells of Cajal and nerve endings were observed in arterial segments with atherosclerotic lesions. The functional significance of the arterial interstitial cells of Cajal and their possible involvement in atherosclerosis and other vascular diseases need clarification.This work was supported by the St Vincents Clinic Foundation, Sydney, Australia.     

Artery and vein size is balanced by Notch and ephrin B2/EphB4 during angiogenesis

A mutual coordination of size between developing arteries and veins is essential for establishing proper connections between these vessels and, ultimately, a functional vasculature; however, the cellular and molecular regulation of this parity is not understood. Here, we demonstrate that the size of the developing dorsal aorta and cardinal vein is reciprocally balanced. Mouse embryos carrying gain-of-function Notch alleles show enlarged aortae and underdeveloped cardinal veins, whereas those with loss-of-function mutations show small aortae and large cardinal veins. Notch does not affect the overall number of endothelial cells but balances the proportion of arterial to venous endothelial cells, thereby modulating the relative sizes of both vessel types. Loss of ephrin B2 or its receptor EphB4 also leads to enlarged aortae and underdeveloped cardinal veins; however, endothelial cells with venous identity are mislocalized in the aorta, suggesting that ephrin B2/EphB4 signaling functions distinctly from Notch by sorting arterial and venous endothelial cells into their respective vessels. Our findings provide mechanistic insight into the processes underlying artery and vein size equilibration during angiogenesis.     

SEM observations of the elastic networks in canine femoral artery

Scanning electron microscopy was used to study the normal architectural arrangement of elastic tissue in a medium-sized muscular artery. Selective NaOH sonication digestion or formic acid digestion was used to expose and isolate the elastic networks in the femoral arteries of four healthy dogs. The digested segments were neutralized and freeze-dried before mounting for scanning electron microscopy (SEM) observation. The fenestrated internal elastic lamina (IEL) had a smooth surface with scattered regions of the fine elastic fibers that made up lacy networks protruding from the luminal surface. Prominent ellipsoid fenestrae, randomly scattered across the surface, were grouped into small and large sizes based on their mean diameter. The openings of most fenestrae were bridged by elastic fibers to give the fenestrae a sieve-like appearance. Large, transversely oriented, fusiform gaps were randomly scattered along the length of the IEL. These gaps, filled in by an elastic fiber network, sometimes spanned as much as a quarter of the vessel circumference. It is suggested that these gaps represent splits in the IEL that have been repaired. The tunica media contained a complex network of anastomosing elastic fibers and lamellae that were primarily circumferential in orientation. A well-defined external elastic lamina formed a solid sheet at the junction of the tunica media and the tunica adventitia. The tunica adventitia contained 8-10 incomplete lamellae of large, interconnecting, longitudinally oriented fibers. The architecture of the elastic network in canine femoral artery was compared with that previously described in medium-sized canine veins and in the rat femoral artery.     

The cardiac muscle in the pulmonary vein of the rat: A morphological and electrophysiological study

The pulmonary veins of albino Wistar rats were studied by means of light and electron microscopy. The media of larger veins consists of cardiac muscle fibers which extend until the vessels attain about 100 μ in diameter. This coat consists of external longitudinal fibers and internal circular fibers. The vasa vasorum are well developed and the capillaries show pseudofenestrations. The numerous adrenergic and cholinergic nerve endings do not form typical motor end-plates as seen in skeletal muscles. The ultrastructure of these media muscle fibers is similar to that of rat hearts. The smooth muscle layer of larger pulmonary veins is not continuous as it is in smaller veins where it forms cushions. Comparisons of albino rats and other rodents reveal striking differences. Action potential shape and propagation velocity (0.5–1.2 m/s) along the myocardial coat of the pulmonary vein were similar to those observed in the left atrium and so was their sensitivity to locally applied acetylcholine. The physiological direction of propagation in rat pulmonary veins is toward the lung. This finding lends support to the hypothesis of a rhythmic, valve-like action of the striated musculature of the pulmonary venous wall during the systole and a possible role in the capacitance of the pulmonary circulation.     

Denervation, but Not Decentralization, Reduces Nerve Growth Factor Content of the Mesenteric Artery

Abstract: In the present study we applied an improved nerve growth factor (NGF) extraction method to examine the effects of denervation and sympathetic decentralization on NGF levels in vascular tissue. Adult male Wistar Kyoto rats underwent mesenteric arterial denervation or splanchnic nerve transection. Four days after operation, animals were killed, and the mesenteric artery and coeliac-superior mesenteric ganglia were removed. The arterial adventitia was stripped from the media to measure NGF levels in nerve and smooth muscle separately. A high concentration of NGF was detected in the normal artery, 90% of which was in the adventitial layer. Surgical denervation significantly reduced the NGF levels in the artery and ganglia by 78 and 71%, respectively. However, within the artery the level of NGF was reduced in the adventitia but not in the media. Thus, the large reduction of NGF content resulted from the loss of nerve plexus from the artery. In contrast, decentralization did not alter the NGF content in the artery, in either the adventitia or media. Our results are in marked contrast to previous studies reporting elevated levels of NGF following denervation. This discrepancy is explained by the ability of our new procedure to extract much greater amounts of NGF from the tissue.