The myotendinous junction of the smooth feather muscles (mm. pennati)

Summary Smooth feather muscles (mm. pennati) consist of bundles of smooth muscle cells which are attached to the feather follicles by short elastic tendons. In addition, some muscle bundles are interrupted by elastic tendons. The elastic tendon is composed of longitudinally arranged elastic fibers which branch and wavy bundles of collagen fibrils. Smooth muscle cells of the muscle bundles are attached to each other by desmosome-like junctions and by fusion of the basal laminae. The cytoplasm of the muscle cells is characterized by conspicuous thick filaments and abundant thin and intermediate filaments. These are attached to band-like dense patches (dense bands) at the plasma membrane which are particularly broad at the tapering end of the muscle cell. The contact surface between smooth muscle cells and their elastic tendon is considerably increased (i) by deep finger-like invaginations and indentations located at the tapering muscle end, and (ii) by branching of the coarse elastic fibers into slender processes, which are attached to the richly folded surface of the muscle cell endings by peripheral microfibrils. This intimate interlocking closely resembles the myotendinous junctions in skeletal muscle. In addition to fibroblasts and fibrocytes, the myotendinous junction of the young growing chicks contains numerous so-called myofibroblasts, which are suggested to represent smooth muscle cells differentiating into fibroblasts of the developing tendon.Dedicated to Professor Dr. Helmut Leonhardt on the occasion of his 60th birthdaySupported by a grant from the Deutsche Forschungsgemeinschaft (Dr. 91/1)     

Reinnervation of regenerating smooth muscle cells in minced vas deferens of the guinea-pig

Summary In adult guinea-pigs, a portion of the wall of the vas deferens was removed, minced and replaced. This caused muscle cells to dedifferentiate, divide and redifferentiate. Reinnervation of redifferentiating cells was followed using electron microscopy and histochemistry. Adrenergic nerves were first observed to re-enter the regenerating area 5 days after operation, and close contacts (within 20 nm) with muscle cells were first seen at 10 days. The total number of adrenergic nerves per 100 muscle cells reached control values by 5 weeks, and by 15 weeks was higher than control levels. Cholinergic nerves first appeared in the regenerating area about 3–4 weeks after the operation. The total number of cholinergic nerves present had not reached control values even at 15 weeks, and no nerve muscle contacts within 20 nm were observed. The ratio of adrenergic to cholinergic nerves in the regenerating area was higher at 15 weeks than in control tissue.This work was supported by grants from the Wellcome Trust and the Medial Research Council     

Ultrastructure and innervation of the swimbladder of Tetractenos glaber (Tetraodontidae)

Summary The general structure, ultrastructure and innervation of the swimbladder of the smooth toadfish, Tetractenos glaber, were examined with light-microscopic, fluorescence-histochemical, and transmission electron-microscopic techniques. The structure of the swimbladder is similar to that of other euphysoclists. Fluorescence histochemistry showed adrenergic fibres in both the secretory and resorptive areas of the swimbladder. Transmission electron microscopy revealed two morphologically distinct axon profiles type-I profiles containing many small, flattened vesicles; type-II profiles containing both large, granular vesicles and rounded, small clear vesicles in varying proportions.The gas-gland cells and surrounding muscularis mucosae are innervated by both type-I and type-II fibres. Type-I fibres also innervate pre-rete arteries. The rete- and gas-gland capillaries do not appear to be innervated. Arteries running to the resorptive area are innervated by type-I fibres. Both type-I and type-II profiles make contact with the muscularis mucosae in the resorptive area. Only type-I fibres innervate the radial dilator muscle in the oval sphincter region, whereas only type II fibres innervate the circular muscle of the oval sphincter.Type-I fibres took up -methyl-noradrenaline, and could not be found after pre-treatment with 6-hydroxydopamine. They are, therefore, assumed to be adrenergic. Type-II fibres were tentatively identified, by exclusion, as cholinergic.     

The eye muscles and their innervation inChaetodon trifasciatus (Pisces,Teleostei, Chaetodontidae)

Synopsis InChaetodon trifasciatus, the large eye has the form of a thick disk rather than that of a globe. A deep cutaneous groove surrounds the eyeball, probably allowing rapid eye movements. The form and innervation of the three pairs of extraocular muscles are described. Each muscle is made of two types of fascicles of fibres, thick and thin. There is neither an anterior nor posterior myodome. The skull attachment of the obliques and of the inferior rectus is made on the thin sagittal ethmoidal membranous septum while that of the other recti occurs on osseous pieces of the skull. The attachment on the eyeball is made on the cartilaginous sclera. The ratio of the lengths of the antagonist muscles, superior vs. inferior oblique, superior vs. inferior rectus and medial vs. lateral rectus, is about 1.43:1. The three oculomotor nerves (III: common oculomotor, IV: trochlear and VI: abducens) as well as the ciliary system are described. For the following reasons, an analogy between the lateral rectus ofChaetodon trifasciatus and the lateral rectus + retractor bulbi of other vertebrates is indicated: (1) the nucleus of nerve III (which innervates four muscles) has four sectors, while that of IV (which innervates only the superior oblique) is made of one sector; (2) nerve VI consists of two roots corresponding to two groups of nerve cells of its motor nucleus and (3) in other vertebrates, nerve VI innervates both the lateral rectus and the retractor bulbi.     

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 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.     

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.     

Fine structure and fluorescence morphology of adrenergic nerves after 6-hydroxydopamine in vivo and in vitro

Summary In parallel fine structural, fluorescence histochemical and biochemical experiments the effect of 6-OH-DA administered in vivo and in vitro on the adrenergic nerves in the mouse iris was studied. As seen in the electron microscope, in vivo administration of 6-OH-DA causes a selective, rapid degeneration of the adrenergic axon terminals similar to that found after axotomy, whereas the cholinergic nerves are unaffected at all time intervals studied. Already 1 hr after the injection of 6-OH-DA the axonal enlargements swell and the size of the dense core of the granular vesicles is strongly reduced. Since the NA stores are almost completely depleted at this time interval, the small core present may be due to a reaction between 6-OH-DA and the fixative. From 2–4 hr after the injection increasing numbers of axonal enlargements with a high electron density are observed in the Schwann cell cytoplasm, which later are digested and completely absent about 48–72 hr after the 6-OH-DA injection. During the following weeks adrenergic axons reappear. This time course of degeneration obtained is considerably faster than that seen after axotomy in other studies. After incubation in 6-OH-DA containing media similar changes were observed in the axonal enlargements, starting already after 30 min of incubation. At this time-point there is a considerable reduction of endogenous NA and a severe damage of the membrane pump uptake mechanism. Incubation with 6-OH-DA and subsequent rinsing for 2 hr caused marked changes, including partly swelling of axons and partly shrinking of the axons into electron dense bodies.The fluorescence histochemical and biochemical results are in good agreement with the ultrastructural studies demonstrating a rapid loss of NA from the adrenergic nerve terminals and main axons and a long lasting depletion of the NA, with a gradual recovery to 75% 6 weeks after the injection.The investigation has been supported by research grants from the Swedish Medical Research Council (14X-2295, 14X-2887 and 04X-3881) Karolinska Institutet, Magnus Bergvalls and Carl-Berthel Nathorst Stiftelser. For generous supplies of drugs we are indebted to the following companies: AB Hässle (6-OH-DA, through Dr H. Corrodi), Pfizer (Niamid^®), Ciba (Serpasil^®). The skilful technical assistance of Miss Bodil Flock, Mrs Waltraut Hiort and Mrs Eva Lindqvist is gratefully acknowledged.     

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.     

Autonomic innervation of the ocular choroid membrane in the chicken

Summary The distribution pattern of adrenergic fibres innervating the ocular choroid membrane of the chicken was studied by means of fluorescence and electron microscopy. In addition, the origin of these fibres was investigated after superior cervical ganglionectomy. Adrenergic axons reach the choroid, partly forming the perivascular plexuses and partly running in the choroid nerves and the choroidal branches of the ciliary nerves. The axon terminals distribute to the smooth muscle cells of the arterial wall and to the extensive system of smooth muscle cells of the intervascular stroma. After unilateral ganglionectomy, fluorescent fibres almost completely disappeared, and degenerative changes could be observed in the terminal varicosities on both smooth muscle cell populations. These findings suggest that the adrenergic axons either originate from neurones within the ipsilateral superior cervical ganglion, or pass through this ganglion. The persistence of normal terminals in short- and long-term ganglionectomised animals shows that the vasal and intervascular muscle cells of the choroid membrane are provided with both an adrenergic and a cholinergic innervation.This work was supported by grant No 80.00442.04 from the Italian National Research Council (CNR)     

Vasomotor innervation in the hind limb muscles of the normal and 6-hydroxydopamine treated cat

Summary Vasomotor nervous distribution to arteries in the cat gastrocnemius and soleus muscles was investigated in the normal and 6-hydroxydopamine treated animal. Muscle samples were processed for the demonstration of acetylcholinesterase by a thiocholine method and for the presence of catecholamines by a formol fluorescence technique. In isolated perfused muscle preparations noradrenaline or adrenaline increased perfusion pressure while acetylcholine or isoprenaline decreased this pressure. A beaded fluorescent periarterial plexus (not visualised after reserpine treatment) was observed. Vasomotor axons, all of which were ACHE-negative, exhibited degenerative changes following 6-hydroxydopamine injection, while ACHE-positive axons outside the adventitial sheath were unaffected. No indication of cholinergic periarterial nerves was found and our evidence suggests that the vasomotor innervation is adrenergic sympathetic in nature.This work was supported by a grant from the Medical Research Council. During the tenure of a Wellcome Trust Fellowship, Dr. F. Joó is on leave of absence from Jozsef Attila University, Szeged, Hungary. The electron microscope is on permanent loan to one of us (J. D. L.) from the Wellcome Trust. We are most grateful to Mrs. G. Howells and Miss D. Morgan for technical assistance and to Mr. P. Hire for photographic help.     

Adrenergic innervation of the tunica media in the saphenous artery of the fetal and newborn guinea-pig

Summary Previous studies have demonstrated that adrenergic nerves are located in the medial-adventitial border of the muscular arteries. Observations made in this study have revealed that adrenergic nerves penetrate into the outer medial layer of the saphenous artery in fetal and newborn guinea-pigs, while in the adult these nerves are located in the medial-adventitial border. It is proposed that the adrenergic nerves located in the tunica media may have a trophic effect on the medial smooth muscle. It is further suggested that the final refinement of the dual control system of arterial walls, by nerves and circulating catecholamines, involves exclusion of adrenergic nerves from the tunica media.     

Homologies of the branchial arch muscles in Zacco platypus (Teleostei: Cypriniformes: Cyprinidae): Evidence from innervation pattern

Homologies of the branchial arch muscles in the cyprinid Zacco platypus are assessed based on their innervation. Muscles serving the first gill arch are innervated by branches of the glossopharyngeal (IX) nerve and those serving other arches by the vagal (X) nerve. Absence of the levator posterior is confirmed. Five pairs of muscles originating from the cranium and inserted onto the specialized 5th ceratobranchial, all unique to cyprinids, are innervated by the 4th branchial trunks of X, indicating that all pairs are derivatives of the sphincter oesophagi, involving reorganization from intrinsic to extrinsic elements. Homologies of some ventral branchial muscles are also discussed and the criteria for homology improved by clarifying the innervation pattern. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc.     

Small intensely fluorescent (SIF) cells and sympathetic nerves in the adult rabbit portal vein and during perinatal development

Summary The ontogenesis of small intensely fluorescent (SIF) cells and the adrenergic nerve plexus is described in stretch preparations of the rabbit portal vein. On the 25 to 26th days of gestation there was a predominance of SIF cells (8 to 30 m in diameter), but a few nerve fibres in bundles were also present. Each portal vein preparation contained 6 to 9 groups of cells. The distribution and number of SIF cells and nerve bundles remained constant until the 31st day of gestation at which stage the number of SIF cells had decreased, while the density of the nerve plexus had increased approximately 4-fold. The adult portal vein exhibited a dense adrenergic plexus, but SIF cells were absent from nine out of ten preparations.     

An ultrastructural study of the polyp and strobila of Atorella japonica (Cnidaria,Coronatae) with respect to muscles and nerves

Atorella japonica were observed by TEM to examine the nerve plexus in the capitulum of the polyp and the cross-striated muscle cells of the strobila. The nerve plexus included a number of neuromuscular junctions and many interneural synapses. Neuromuscular junctions contained two types of synaptic vesicle: clear and small (ca 75 nm diam.), and dense cored and large (ca 120 nm diam.). The first type of vesicle always appeared near the presynaptic membrane and the second type was distributed behind the former. In interneural synapses, two types of vesicle which were similar to neuromuscular synaptic vesicles were recognized. They were distributed in a pattern similar to that of the neuromuscular synaptic vesicles, but these vesicles were found on both sides of the two synaptic membranes.     

Postnatal ontogenic development of the adrenergic innervation pattern in the rat portal vein

Summary The postnatal development of the adrenergic innervation pattern in the rat portal vein has been studied with the histochemical fluorescence method of Hillarp and Falck.Stretch preparations and transverse freeze-dried sections of intact portal veins were studied from rats during the first 5 weeks of life and from adult rats. Orientation of undifferentiated smooth muscle cells into two layers was observed at 4 days of age. Dominance of the thick outer longitudinal muscle layer was apparent at two weeks of age. A terminal adrenergic nerve plexus with some varicosities was restricted outside the media at the end of the first week. Ingrowth of penetrating non-terminal adrenergic nerve fibers through the longitudinal muscle layer occurred during the second week of age when the main terminal nerve plexus was developing between the two muscle layers. After 3 weeks of age the adult pattern of a two-dimensional adrenergic plexus between the muscle was established. In the adult rat pharmacological treatment with nialamide and noradrenaline revealed the thin, penetrating non-terminal adrenergic nerve fibers in the longitudinal muscle layer which were poorly visible otherwise.The present observations strongly indicate that the main adrenergic plexus between the two muscle layers emanates directly from the outer axonal plexus. These findings are discussed regarding possible trophic interactions between ingrowing sympathetic adrenergic vasomotor nerves and maturing vascular smooth muscle.Supported by grants from the Swedish Medical Research Council (grants No. 14X-2207, O4P-4173, 3884), Magn. Bergwall's Foundation, G. & M. Lindgren's Foundation, the Medical Faculty of University of Göteborg. The technical assistance of Miss Serney Bööj, Mr. Pär-Anders Larsson and Miss Ann Kjellstedt is gratefully acknowledged     

Phylogeny and co-speciation in feather mites of the subfamily Avenzoariinae (Analgoidea: Avenzoariidae)

A cladistic analysis of phylogenetic relationships is carried out for the feather mite subfamily Avenzoariinae. The analysis is made at two different taxonomic levels, for 19 genera of the all family Avenzoariidae and for taxa of species rank for the Avenzoaria and Bychovskiata generic groups. A subsequent comparative analysis of phylogenetic hypotheseis for the subfamily Avenzoariinae and recently accepted phylogenetic hypotheses of the shorebirds Charadriiformes indicates co-speciation of feather mites with their hosts. As a result of the comparative analysis it is suggested, that the subfamily Avenzoariinae originated from an ancestor of the order Charadriiformes and co-speciated with this host order. The expected pattern of parallel evolution is disturbed by different evolutionary events, such as host shifts, extinction of mites and differential evolutionary rates of mite lineages in different phyletic branches of feather parasites.     

Smooth muscles in relation to the gill skeleton of Perca fluviatilis: organization and innervation

Summary Two laminae composed of smooth muscles, elastic tissue and collagen have been described in relation with the gill skeleton in Perca fluviatilis. A transverse smoothmuscle lamina joins the base of the cartilage rods of the two opposite hemibranchs. A longitudinal smooth-muscle lamina runs parallel to the afferent branchial artery and joins the cartilage rods from one filament to the other. In both laminae, the formaldehyde-induced fluorescence technique (Falck-Hillarp) reveals a network of nerve fibers displaying a green fluorescence characteristic of catecholamines. At the ultrastructural level, the presence of nerve endings containing clear and granular vesicles, and the degeneration of these endings after 6-hydroxydopamine treatment confirm the aminergic nature and the sympathetic origin of this innervation. Surgical denervation brings evidence that the innervation of both laminae is supplied by the metatrematic branches of the branchial nerves. The role of these smooth-muscle laminae remains speculative.