A histochemical study on the innervation of the cerebral blood vessels in bats

Summary The adrenergic and cholinergic nerves innervating the cerebral blood vessels of four species of Japanese chiropterids (Rhinolophus ferrumequinum, Murina leucogaster, Vespertilio superans and Miniopterus Schreibersi) have been investigated using specific histochemical techniques. In all these species of bats arteries of the internal carotid system are poorly developed, whereas those of the vertebro-basilar system are well developed. The adrenergic and cholinergic nerves innervating these cerebral arteries, however, all originate from the stem nerve bundles entering the cranial cavity along the internal carotid artery. Both nerve plexuses are among the densest of any vertebrate species so far investigated. Adrenergic nerve plexuses are usually composed of complicated meshworks of fine fibres, while cholinergic ones are composed of rather longitudinally arranging meshworks of both thick and thin fibres, exhibiting a very high acetylcholinesterase activity. Small parenchymal arteries and arterioles are also dually innervated by adrenergic and cholinergic nerve fibres of peripheral origin. Intracerebral capillaries, on the other hand, are in several places directly connected with both adrenergic and cholinergic fibres of parenchymal origin. Capillaries in the cerebral and cerebellar cortices, diencephalon and cochlear nucleus in V. superans exhibit a heavy non-nervous acetylcholinesterase activity in their walls, but in R.ferrumequinum and M. schreibersi, the response is weak or negative, except for that in the cochlear nucleus.     

A histochemical study of the innervation of cerebral blood vessels in the snake

Summary Dual innervation of snake cerebral blood vessels by adrenergic and cholinergic fibres was demonstrated with the use of histochemical methods. Although the nerve plexuses are somewhat less dense, the essential features of innervation of the blood vessels are similar to those of mammals with the exception that the adrenergic plexuses are more prominent than the cholinergic plexuses. The major arteries of the cerebral carotid system have a rich nerve supply. However, the innervation is less rich in the basilar and poor in the spinal (vertebral) arteries. Although the arteries supplying the right side of head are poorly developed, three pairs of arteries, cerebral carotids, ophthalmics and spinals, supply the snake brain. The carotids and ophthalmics are densely innervated and are accompanied by thick nerve bundles, suggesting that the nerves preferentially enter the skull along those arteries. Some parenchymal arterioles are also dually innervated. Connection between the brain parenchyma and intracerebral capillaries via both cholinergic and adrenergic fibres was observed. In addition cholinergic nerve fibres, connecting capillaries and the intramedullary nerve fibre bundles, were noticed. Capillary blood flow may be influenced by both adrenergic and cholinergic central neurons. The walls of capillaries also exhibit heavy acetylcholinesterase activity. This may indicate an important role for the capillary in the regulation of intracerebral blood flow.     

Adrenergic innervation of the umbilical vessels

Summary The intra- and extrafetal portions of the umbilical vessels in the guinea-pig and the umbilical cord of man, mouse and rabbit have been investigated by means of the Falck-Hillarp method for the fluorescence microscopical demonstration of catecholamines.The umbilical cord was found to be devoid of nerves in all species investigated. Adrenergic nerves are present only in the immediate vicinity of the umbilicus.The intrafetal portions of the umbilical artery and umbilical vein receive adrenergic nerves, the distribution pattern of which is different for each vessel. In the guinea-pig the ductus venosus is an intrahepatic branch of the vena umbilicalis. No adrenergically innervated sphincter has been detected in the initial segment of the ductus venosus. Regional variations in the pattern of innervation of the intrafetal portion of the umbilical vein are paralleled by regional differences in the construction pattern of the vessel's wall. Regional differences in the noradrenaline concentration (measured by fluorometry) which correspond to the fluorescence microscopical findings have been detected in umbilical vessels: low noradrenaline content of the umbilical cord, high concentrations in the intrafetal sections of the umbilical vessels. The noradrenaline concentration of the guinea-pig umbilical artery is three times that of the umbilical vein.Supported by the Joachim Jungius-Gesellschaft der Wissenschaften, Hamburg.For continuous advice and constructive criticism I am indebted to Prof. Dr. Dr. E. Horstmann.     

The adrenergic innervation of the pulmonary vasculature,the lung and the thoracic aorta,and on the presence of aortic bodies in the domestic fowl (Gallus gallus domesticus L.)

Summary The Falck-Hillarp technique for the localisation of biogenic amines has been used to examine the adrenergic innervation of the thoracic vasculature and lung, and to demonstrate the occurrence of aortic bodies in the domestic fowl. The proximal pulmonary vein is very densely innervated but distally the innervation becomes sparse. The pulmonary artery is sparsely innervated over its whole length. The bronchial muscle of the lung has little adrenergic innervation and fluorescent cell bodies are absent from the lung. The thoracic aorta receives a moderate adrenergic innervation. In the region of the aortic arch and pulmonary arteries groups of fluorescent cells are common. Extramedullary chromaffin cells and small, intensely fluorescent cells occur within these groups. In the media of the aorta and pulmonary artery other types of fluorescent cells are found. These results are discussed in the light of previous observations.Part of this work was performed while the author was a postdoctoral research fellow of the National Heart Foundation of Australia. His thanks are due to Prof. G. Burnstock for use of laboratory facilities.     

Comparative study of the autonomic innervation of the mammalian ovary,with particular regard to the follicular system

Summary The autonomic innervation of the ovary was studied in 12 mammalian species utilizing the cholinesterase method in combination with pseudocholinesterase inhibition for the cholinergic component, and glyoxylic acid histochemistry together with fluorometric determination of noradrenaline for the adrenergic component. Ovaries from cow, sheep, cat, and guinea pig were very richly supplied with adrenergic nerves in the cortical stroma, particularly enclosing follicles in various stages of development. In the follicular wall the nerve terminals were located in the theca externa, where they ran parallel to the follicular surface. Numerous adrenergic terminals also surrounded ovarian blood vessels. The adrenergic innervation was of intermediary density in the human ovary and in the pig, dog, cat, and opossum. Ovaries from rabbit, mouse and hamster had a sparse adrenergic nerve supply. The amount of intraovarian adrenergic nerves agreed well with the tissue concentration of noradrenaline in the various species. The cholinergic innervation was generally less well developed, but had the same distribution as the adrenergic system around blood vessels and in the ovarian stroma, including follicular walls.     

The adrenergic innervation of the renal artery and vein of the rat

Summary The adrenergic innervation of the extrarenal blood vessels of the rat left kidney was investigated by fluorescence histochemistry and by electron microscopy. The trunk of the renal artery proximal to the aorta is elastic and appears to be very sparsely innervated. In contrast, near the kidney the renal artery—which divides into 3 to 4 large branches of the muscular type possesses a dense adrenergic innervation. The adrenergic terminal axons are situated in the adventitia close to the external elastic lamella, but only rarely in close contact with smooth muscle cells. In most instances several terminal axons are grouped and enclosed by a Schwann cell, single axons being rare. All terminal axons are able to take up and to store 5-hydroxydopamine which strongly suggests that they are adrenergic. The innervation of the renal vein is more sparse than that of the muscular arteries but somewhat denser than that of the elastic artery. In addition, close to the origin of the renal artery the presence of small intensively fluorescent (SIF) cells as well as of some adrenergic ganglion cells is noted. The latter are situated in the adrenergic nonterminal axon bundles, which run parallel to the blood vessels.It is concluded that the uneven adrenergic innervation along the artery as well as individual variations in the branching of the artery are the main causes of the unusually high individual variations of the NA content of this organ such as used in pharmacological experiments.     

Effect of the ovulatory cycle on oviductal sperm storage in the domestic fowl

Anovulatory domestic hens (pregnant mare serum-treated) and normally cyclic domestic hens were artificially inseminated with 0.034 ml of pooled semen. A subsequent microscopic assessment of the uterovaginal sperm storage glands on days 1, 3, 6, 9 and 12 post-insemination indicated that the sperm glands emptied over time in the normally cyclic hens, but not in the anovulatory hens. The data suggest that events associated with ovulation and/or oviposition are important to the sperm gland emptying process.     

Synaptology of neurosecretory cells in the nucleus paraventricularis of the domestic fowl

Summary Processes of magnocellular neurosecretory cells (MNCs) are easily identifiable on the basis of their content in neurosecretory granules in the neuropil of the rostral division of the paraventricular nucleus (PVN) of the domestic fowl. In specimens sacrificed during the winter the synaptic organization of the neuropil and the pattern of synapses ending on neurosecretory processes were studied at the ultrastructural level. Synapses in the rostral part of the PVN neuropil may be divided into three main categories on the basis of their morphology and their content of clear and dense-core synaptic vesicles. These different types of terminals can be attributed to aminergic, peptidergic or other types of synapses. The percent distribution of synapses within these categories differs when all synapses observed in the neuropil or only those ending on MNC processes are compared. Present ultrastructural data obtained in birds support two physiological hypotheses already suggested for mammals, i.e., the probable existence of a recurrent pathway to MNCs via an interneuron, and the importance of aminergic and peptidergic input in regulating the electrical activity of MNCs.This work was partly supported by a CNR grant (n. 81.00377.04)     

Development of the mitochondrial apparatus and blood supply of skeletal muscle fibers during ontogenesis of domestic fowl

The diameter, length, and numerical density of capillaries, diameter of muscle fibers, size and numerical density of mitochondrial profiles, and relative volume of mitochondria in them were determined in the chicken red oxidative gastrocnemius and white glycolytic pectoral muscle during development from day 10 of embryogenesis to six month of postnatal life. The bulk blood flow was measured in these muscles by hydrogen clearance during postembryonic development. During embryogenesis, the fibers of gastrocnemius muscle develop and grow at a higher rate, while during postembryonic development, those of the pectoral muscle develop faster. The density of mitochondrial profiles increases during embryogenesis and decreases after hatching, while their mean size increases, especially in the oxidative fibers, but it somewhat decreases in 6-month old chicks. Redistribution of mitochondria across the fiber section during development takes place in both muscles: they are localized predominantly in the center in 18-day embryos and in the periphery, especially in the gastrocnemius fibers, in 6-month old chicks. At hatching, the length of capillaries is similar in both muscles, but as chicks grow, the proportion of longer (more than 600 µm) capillaries in the pectoral muscle sharply increases, while their density and bulk blood flow decrease. Ratios were determined between structural parameters of the capillary bed and mitochondria, on the one hand, and oxygen consumption (ml/min per 1 mm fiber and 100 g muscle mass), on the other.__________Translated from Ontogenez, Vol. 36, No. 2, 2005, pp. 135–144.Original Russian Text Copyright © 2005 by Belichenko, Korostyshevskaya, Maksimov, Shoshenko.     

A variant in the restriction endonuclease cleavage pattern of mitochondrial DNA in the domestic fowl, Gallus gallus domesticus

Summary. The cleavage patterns of mitochondrial DNAs (mtDNAs) were investigated from 15 lines of domestic fowls, Gallus gallus domesticus . using 11 restriction endonucleases. The cleavage patterns with 10 restriction endonucleases were identical in all the lines. A variant was found in a line of White Leghorn in the pattern with Mspl digestions. Cleavage patterns of the red jungle fowl, Gallus gallus gallus , were identical to the common patterns shown by the 14 lines of domestic fowls.     

Organization of the sympathetic innervation in liver tissue from monkey and man

Summary The sympathetic innervation of the liver of monkey and man has been investigated in a combined fluorescence histochemical, chemical and electron microscopical study. By means of the Falck-Hillarp fluorescence method a dense network of monoamine-containing nerve fibers was visualized in liver tissue of monkey and man. The nerve fibers ran in close contact to both hepatocytes and blood vessels. Chemical quantitations showed high concentrations of noradrenaline in both human and monkey liver. Microspectrofluorometry of the intraneuronal monoamine resulted in spectra characteristic of a catecholamine. For the electron microscopical study the dopamine analogue, 5-hydroxydopamine, was used to label the catecholamine terminals in both human and monkey liver. The nerve profiles, identified as catecholamine-containing, were demonstrated in a perivascular location and in close contact to hepatocytes. No synaptic membrane specializations were present between nerve fibers and hepatocytes. The general ultramorphology and intralobular distribution pattern of nerves in the liver of monkey and man were similar. The present results prove the existence of a sympathetic innervation of hepatocytes and blood vessels in the liver of man and monkey.     

Innervation of arteriovenous anastomoses in the brood patch of the domestic fowl

Summary The innervation of blood vessels in the brood patch (thoracic skin) of the domestic fowl was studied by use of the catecholamine fluorescence technique, acetylcholinesterase staining, and the immunoperoxidase technique for demonstration of vasoactive intestinal polypeptide (VIP). Large arteries and veins were sparsely innervated, whereas arteriovenous anastomoses (AVAs) were densely innervated by adrenergic, acetylcholinesterase-positive, and VIP-immunoreactive nerve fibres. The rich supply of different vasomotor nerves to AVAs emphasizes the importance of these vascular shunts in regulating blood flow and, in turn, the transport of heat to the brood patch. Furthermore, the presence of VIP-immunoreactive nerve fibres in the vasculature of the brood patch suggests that VIP might be the mediator of the previously reported cold-induced vasodilatation.     

Localization of VIP-immunoreactive nerves in airways and pulmonary vessels of dogs,cats, and human subjects

Summary VIP-containing neurons were localized in lungs from dogs, cats, and human subjects by means of the indirect immunofluorescence technique. Nerve fibers and terminals were observed in the smooth muscle layer and glands of airways, and within the walls of pulmonary and bronchial vessels, especially at the medial-adventitial junction. VIP-positive nerve cell bodies were identified in ganglia located in the walls of bronchi. These findings provide an anatomic basis for the possible modulation of airway and pulmonary vascular function by this neuropeptide.Supported by National Research Service Award HL 5914 and by Lung Center Award HL 14187 from NHLBI, USPHS     

A histochemical study of keratinization in the domestic fowl (Gallus gallus)

The microanatomy of the epidermis of the domestic fowl is described and related to the distribution of various histochemical constituents involved in keratinization.
The avian horny layer over the back is composed of a loose network of structurally solid horny cells. This is in contrast to most mammalian epidermal horny cells in which structural keratin is found only in the peripheral cytoplasm, and the interior of the keratinocyte contains soluble products of cytolysis with possibly some free keratin filaments dispersed in the fluid material.
The avian tarsal epidermal horny scales show similarities to both the scales of lizards and snakes and to mammalian tail scales which appear to be homologous structures.
It is suggested that a thin layer of cells containing no detectable disulphide bonds, found in the tarsal scale region of the young chick, is probably mechanically weak and may function as a fission plane for sloughing of the horny layer. A specialized epidermis and thickened horny layer is developed in the fowl on the plantar underside of the toes, but this is quite different in structure from the mammalian plantar epidermis.
The overlapping of zones rich in ribonucleic acid (RNA) and bound cysteine (SH) in the growing feather suggests that protein synthesis and the preparatory stages to keratin disulphide bonding normally occur concurrently in feather formation. This is in contrast to the growing hair which has a region rich in RNA followed immediately before it becomes keratinized by a discrete keratogenous zone weak in RNA but rich in bound cysteine.     

The neuronal and extra-neuronal localisations of biogenic amines in the cervical region of the domestic fowl (Gallus Gallus domesticus L.)

Summary The Falck-Hillarp technique has been used to demonstrate the neuronal and extra-neuronal localisations of biogenic amines in the cervical region of the domestic fowl. Adrenergic cell bodies were found in the superior cervical ganglion and in the ganglia of the cervical paravertebral chain. The axons of the latter ran into the corresponding spinal nerves and thus to the periphery. Very few adrenergic fibres were found in the interganglionic portions of the cervical paravertebral chain. The precarotid branch of the glossopharyngeal nerve, and the vagus nerve, below its junction with the former, contained numbers of adrenergic fibres. The retrocarotid nerve-trunk from the superior cervical ganglion was composed of adrenergic fibres. With the exception of the parathyroid gland, the adrenergic nerves seen in the branchial derivatives (thymus, thyroid and ultimobranchials) appeared to be associated with blood vessels. Under normal conditions the cells of the ultimobranchial body were nonfluorescent, but after injection of 6-hydroxydopamine the cells were brightly fluorescent. The carotid body was devoid of adrenergic nerves other than those with blood vessels, but the cells of the carotid body were brightly fluorescent. Various fluorescent cell types were found throughout the cervical region, particularly in association with the vasculature. I should like to thank Prof. G. Burnstock (Department of Zoology, Melbourne University) in whose department this work was carried out and Dr. R. D. Hodges (Wye College, London University) for his indispensable advice on the disposition of the avian ultimobranchial body. The author held a Postdoctoral Research Fellowship of the National Heart Foundation of Australia during part of this study.     

Morphometry of the galliform cecum: A comparison between Gambel's quail and the domestic fowl

Summary Tissues from the proximal, middle, and distal regions of the ceca of Gambel's quail and domestic fowl were examined by scanning and transmission electron microscopy. Cellular and subcellular structures, including epithelial cell height, mitochondrial volume fraction, microvillous surface area, proportion of goblet cells, and junctional complex characteristics, were quantified by a variety of stereologic procedures and other measurement techniques. The mucosal surface of quail cecum shows a much more highly developed pattern of villous ridges and flat areas than that of fowl cecum. The fowl has significantly greater cell heights than the quail in all cecal regions. The mitochondrial volume fraction does not differ significantly with species or region, but mitochondria are concentrated on the apical side of the nucleus. In both species, the proximal cecal region has the greatest microvillous surface area. All 3 components of junctional complexes, including zonula occludens, zonula adhaerens, and macula adhaerens, are quantified. When all factors are considered, the quail cecum appears to have morphological characteristics consistent with a greater potential capacity for absorption than the fowl cecum.     

The ultrastructure of the intestinal nerve of Remak in the domestic fowl

Summary An ultrastructural study was made of the neurons, satellite cells and vesiculated axons of the intestinal nerve of the domestic fowl. Broad membrane-to-membrane contacts between adjacent nerve cell bodies were sometimes observed. The cell bodies and processes were not always separated from the extracellular space by a capsule of satellite cells. Following fixation using potassium permanganate, catecholamine (CA)-containing neurons in the intestinal nerve, unlike those in the lumbar parasympathetic ganglia, did not possess any small granular vesicles (SGV). Following exposure to noradrenaline, SGV could be demonstrated in the cell bodies of the juxta-ileal ganglia but not the juxta-rectal ganglia of the intestinal nerve. Non-CA axons were examined in tissue from birds that had been pretreated with 6-hydroxydopamine. Approximately one half of the non-CA axons formed axo-somatic contacts. Most of the non-CA axons contained varying proportions of small clear vesicles, large clear vesicles and large granular vescles. Statistical analysis showed that the non-CA axons could not be subdivided according to their vesicle content. CA-axons contained many SGV and were found in close apposition to neuronal somata and processes, and in the neuropil.     

Distribution and vasomotor effects of peptide HI (PHI) in feline cerebral blood vessels in vitro and in situ

Peptide HI (PHI)-immunoreactive nerve fibres were numerous around cerebral blood vessels of the cat. The number and distribution resemble that previously found for vasoactive intestinal polypeptide (VIP), a peptide with which PHI co-exists in pial arteries, at least in some segments. PHI and VIP elicit dilatation in a concentration-dependent manner in isolated middle cerebral arteries; the maximum effects were similar but VIP was considerably more potent. Neither effect was blocked by atropine, cimetidine or propranolol, confirming an action at a non-adrenergic, non-cholinergic site. In chloralose-anaesthetized cats PHI and VIP elicited concentration-dependent dilatations; the magnitude of responses was similar, however, considerably more PHI was necessary to elicit the same response as that of VIP. The results suggest that though both peptides are co-localized and may act at the same receptor, VIP is a more likely candidate for eliciting dilatation during physiological conditions.     

Neuronal degeneration in the pineal ganglion during the post-hatching development of the domestic fowl

Summary The frequency of pineal ganglia associated with the pineal tract, and the numbers of acetylcholinesterasepositive neurons in these ganglia were studied in the domestic fowl during the post-hatching period by means of the acetylcholinesterase method. Furthermore, the degeneration of nerve cells in pineal ganglia of 40-day-old domestic fowl was investigated in detail at the electron-microscopic level. The rate of pineal organs containing one or more ganglia was 50% in 2- to 13-day-old, 38% in 40-day-old, and only 10% in 1-year-old domestic fowl. In parallel, the number of acetylcholinesterase-reactive nerve cells that constitute individual pineal ganglia decreased after hatching. Various degrees of neuronal degeneration were found in the pineal ganglia: swelling of the endoplasmic reticulum, electron-dense degeneration of the cytoplasm, and pyknosis of the nerve cell nucleus. Clusters of macrophages containing numerous lysosomes filled with debris-like material were scattered in the ganglion. In addition, plasma cells were observed in association with degenerating nerve cells. These results confirm the suggestion that the loss of acetylcholinesterase-positive nerve cells in the pineal ganglia of the domestic fowl is due to naturally occurring, programmed neuronal cell death. This process is discussed with reference to phenomena of cell death observed in other components of central nervous system.Fellow of the Alexander von Humboldt Foundation, Bonn, Federal Republic of GermanyThe authors are indebted to Professor A. Oksche and Dr. H.-W. Korf (Giessen) for stimulating discussions