Urethral chromaffin cells

Summary The urethral mucosa of the rat, rabbit and guinea-pig was examined with both fluorescence and electron microscopy. Employing the former technique, numerous brightly fluorescing flask-shaped cells were observed amongst the basal cells of the urethral epithelium in all three species. In the electron microscope cells with a similar shape and distribution are distinguished by their content of membrane-limited dense granules, extensive Golgi membranes and bundles of filaments. In favourable planes of section short microvilli extend from the apical region of these cells which are joined to neighbouring urethral epithelial cells by zonulae occludentes. These fluorescent, granule-containing cells are classified as urethral chromaffin cells.Fluorescent nerves were not observed in relation to the urethral epithelium although the electron microscope revealed axons lying singly or in groups both beneath and between the urethral epithelial cells. Many of these axons appear varicose and contain small, agranular vesicles, a few large granulated vesicles and numerous mitochondria. Occasionally a vesicle-containing axon lay adjacent to a urethral chromaffin cell. While a direct autonomic innervation of these cells could not be discounted it is concluded that the majority of nerves probably perform a sensory function.     

The ultrastructure of the innervation of the intestinal wall in the teleosts Myoxocephalus scorpius and Pleuronectes platessa

Summary The innervation of the intestinal wall in the teleosts Myoxocephalus and Pleuronectes was examined electron microscopically. Two classes of axons can be identified. The first, which is in the majority, contains numerous 50–150 nm granular vesicles as well as some 40–50 nm agranular vesicles while the second contains predominantly the 40–50 nm agranular vesicles. Chromate/dichromate staining methods suggest that the first type is aminergic. Both types lie in close association with the perikarya of intrinsic myenteric neurons but only axons containing predominantly agranular vesicles have synaptic membrane specialisations. No axon bundles pass into the longitudinal muscle layer in Myoxocephalus gut and though some do in Pleuronectes, they do not closely approach the smooth muscle cells. Axons containing large granular vesicles lie in intimate contact with the myocytes of the circular muscle layer. Both axon types pass through the submucosa to form a plexus underneath the mucosal epithelium. Varicosities containing agranular or granular vesicles are separated from the epithelial cells by a gap of about 200 nm in which lies a basal lamina.     

Neurotensin immunoreactivity in the central nucleus of the rat amygdala: an ultrastructural approach

Neurotensin (NT) was demonstrated in the central nucleus of the rat amygdala (CNA) using a modification of the avidin-biotin complex immunohistochemical technique. Electron-dense reaction product (particles were 15-25 nm in diameter) was localized in perikarya, dendrites, axons, and axon terminals. It was found also associated with profiles of rough endoplasmic reticulum, mitochondria, microtubules, and small agranular as well as large granular vesicles. In distal dendrites, the reaction product was associated with microtubules, vesicles, and postsynaptic densities. Axon terminals of three types formed synaptic contracts with NT-immunoreactive neurons in the CNA: one was characterized by numerous round or oval agranular vesicles, the second by numerous pleomorphic vesicles, and the third by agranular vesicles that were loosely distributed and pleomorphic. All three types formed symmetric axosomatic and asymmetric axodendritic contacts. NT-immunoreactive axon terminals containing small round agranular vesicles stood out clearly from the intermingling profiles of immunonegative structures. We found numerous glomeruli, each consisting of a central NT-immunoreactive dendrite surrounded by all three types of axon terminals. We observed that some NT-immunoreactive terminals formed symmetric axoaxonal contacts with each other, providing evidence for the presence of local NT-to-NT circuits, whereas many others synapsed with axon terminals devoid of NT immunoreactivity.     

Varicose axons bearing “synaptic” vesicles on the basal lamina of the human seminiferous tubules

Summary Normal (infant and adult) and pathological testes were examined by electron microscopy in order to study testicular innervation. Nerves composed of non-myelinated fibres were abundant in the tunica vasculosa of the tunica albuginea. These nerves penetrated into the testicular septa reaching the interstitial tissue. This showed numerous non-myelinated nerve fibres running among the Leydig cells and blood vessels. Single axons or small groups of them, partially surrounded by Schwann cells, approached: 1) the Leydig cells, 2) the interstitial blood vessels, and 3) the seminiferous tubules. Single naked axons were also observed primarily in the proximity of the seminiferous tubules. These axons showed varicosities containing both small and large synaptic vesicles. The latter were less numerous and contained a central dense core. Small vesicles were agranular. Some varicose axons ran across the myofibroblast layer of the tunica propria reaching the basal lamina of the seminiferous tubules at the level of the Sertoli cells but not at the level of the spermatogonia. The intercellular space between Sertoli cell and axon membrane was about 150–200 nm.Profesor Agregado de Histología y EmbriologíaProfesor Adjunto de Citología e HistologíaProfesor Adjunto de Histología y Embriología     

Localization of catecholamines and acetylcholinesterase in the terminal nerve fibres of the rabbit myometrium

Summary The autonomic nerves of the myometrium of the rabbit were studied in order to demonstrate simultaneously the adrenergic nature of an axon and the localization of acetylcholinesterase (AChE) in the same axons. The synaptic vesicles of the adrenergic axons and nerve terminals remained partially filled with the electron dense material typical for them after formaldehyde fixation and short incubation time for AChE. AChE stain was localized regularly on the axons which contained agranular synaptic vesicles and also on axons which contained dense cored synaptic vesicles beeing probably adrenergic. The role of AChE on the adrenergic axons is discussed.     

Effect of oculomotor and trigeminal nerve section on the ultrastructure of different myoneural junctions in the rat extraocular muscles

Summary The intramuscular nerves and myoneural junctions in the rat rectus superior, medialis and inferior muscles from 10 hours to about 10 days after section of the trigeminal and oculomotor nerves were studied with the electron microscope. Two different kinds of myoneural junctions are to be observed; one type derives from myelinated nerves and is similar to the ordinary myoneural junctions (motor end plates) of other striated skeletal muscles, while the other type derives from unmyelinated nerves, is smaller in size and has many myoneural synapses distributed along a single extrafusal muscle fibre.Section of the trigeminal nerve caused no changes in the myoneural synapses. After section of the oculomotor nerve degenerative changes occur in both the myelinated and unmyelinated nerves and in both types of myoneural junctions. In the axon terminals of both the myelinated and unmyelinated nerves the earliest changes are to be observed 10 to 15 hours after section of the nerve. First, swelling of the axoplasm, fragmentation of microtubules and microfilaments and swelling of mitochondria takes place, somewhat later agglutination of the axonal vesicles and mitochondria. The axon terminals are separated from the postsynaptic muscle membrane by hypertrophied teloglial cells about 24 hours after section of the nerve. The debris of the axon terminals is usually digested by the teloglial cells within 42 to 48 hours in both types of myoneural junction.Changes in the postsynaptic membrane are observed in the myoneural junctions of the unmyelinated nerves as disappearance of the already earlier irregular infoldings, whereas no changes take place in the infoldings of the motor end plates. The postsynaptic sarcoplasm and its ribosomal content increase somewhat.The earliest changes occur along unmyelinated axons 10 to 15 hours and along myelinated axons 15 to 24 hours after nerve section. The unmyelinated axons are usually totally digested within 48 hours, whereas the myelinated axons took between 48 hours and 4 days to disappear. The degeneration, fragmentation and digestion of the myelin sheath begin between 24 and 42 hours and still continues 10 days after the operation.The results demonstrate that in the three muscles studied structures underlying the physiologically well known double innervation of the extraoccular muscles are all part of the oculomotor system.We are grateful to Professor Antti Telkkä, M. D. Head of the Electron Microscope Laboratory, University of Helsinki, for permission to use the facilities of the laboratory.     

Perivascular nerve fibres and endothelial cells of the rat basilar artery: immuno-gold labelling of antigenic sites for type I and type III nitric oxide synthase

Ultrastructural localisation of type I (neuronal) and type III (endothelial) isoforms of nitric oxide synthase in perivascular nerve fibres (axons) and endothelial cells was studied in the Wistar rat cerebral basilar artery, using monoclonal antibodies either to type I or type III nitric oxide synthase and post-embedding colloidal-gold immunocytochemistry. Labelling signal (gold particles) for type I and type III nitric oxide synthase was localised both in axons and endothelial cells. In the axon profiles, labelling for either type I or type III nitric oxide synthase was localised in the axoplasm and the lumen and/or membrane of small agranular synaptic vesicles. In the endothelial cells, labelling for either type-I or type-III nitric oxide synthase was predominantly in the cytoplasm. The present qualitative data extends our previous study of cerebrovascular nerve fibres and endothelial cells employing monoclonal antibodies; the localisation of nitric oxide synthase in a subpopulation of synaptic vesicles in nitric oxide synthase-positive cerebrovascular nerves suggests that vesicular mechanisms may be involved in the production/release of nitric oxide. © 1998 Chapman and Hall     

Light and electron microscopic observations of the autonomic innervation of the mouse gallbladder mucosa

Summary The autonomic innervation of the mouse gallbladder mucosa was studied using histo-and cytochemical methods. In a light microscopic investigation the distribution of acetylcholinesterase (AChE) activity and formaldehyde-induced fluorescence was studied histochemically. Nerve fibres and small varicosities showed concentrations of AChE activity very close to the epithelium in the subepithelial connective tissue. No adrenergic nerves were observed in the mucosa.When using the electron microscope and employing the potassium permanganate fixation/staining technique only one sort of axonal enlargement was encountered, viz. the cholinergic type. These varicosities contained numerous agranular vesicles (500–600 Å in diameter). No varicosities of the adrenergic (dense-cored vesicles) type were observed.Signs of increased secretory activity in the epithelium were observed in the first few minutes after cholinergic stimulation. After repeated in vivo stimulation, there was an almost total depletion of glycoprotein granules, best seen when using the cytochemical PA-CrA-silver technique. The findings suggest that the subepithelial connective tissue and the epithelium of the mouse gallbladder mucosa have a cholinergic innervation.     

Light and electron microscopic observations of the autonomic innervation of the mouse gallbladder mucosa. A histochemical, cytochemical, and secretory study.

The autonomic innervation of the mouse gallbladder mucosa was studied using histo- and cytochemical methods. In a light microscopic investigation the distribution of acetylcholinesterase (AChE) activity and formaldehyde-induced fluorescence was studied histochemically. Nerve fibres and small varicosities showed concentrations of AChE activity very close to the epithelium in the subepithelial connective tissue. No adrenergic nerves were observed in the mucosa. When using the electron microscope and employing the potassium permanganate fixation/staining technique only one sort of axonal enlargement was encountered, viz. the cholinergic type. These varicosities contained numerous agranular vesicles (500-600 A in diameter). No varicosities of the adrenergic (dense-cored vesicles) type were observed. Signs of increased secretory activity in the epithelium were observed in the first few minutes after cholinergic stimulation. After repeated in vivo stimulation, there was an almost total depletion of glycoprotein granules, best seen when using the cytochemical PA-CrA-silver technique. The findings suggest that the subepithelial connective tissue and the epithelium of the mouse gallbladder mucosa have a cholinergic innervation.     

Nervous system of the snailHelix aspersa

Summary The fine structure of neurosecretory nerves and endings associated with the sheath of the infraesophageal ganglion ofHelix aspersa is described. The sheath is a neurohemal organ whose vascularized stroma receives both monoaminergic and peptidergic endings. The latter occur along the surface of the nerves or scattered within the stroma. They include a complex population of vesicular profiles. The granular vesicles (1300–3000 Å in diameter) exhibit structural modifications that may be related to the intra-axonal release of their neurohormones. The agranular vesicles (500–2000 Å in diameter) occur in large numbers and lie mostly adjacent to the axon surface. Synaptoid specializations seem to represent active sites for the extracellular discharge of neurosecretory material. The monoaminergic endings so far studied lack synaptoid specializations and contain small granular (800–1300 Å in diameter) and agranular (700 Å in diameter) vesicles. Two kinds of non-neural cells are associated with the nerves: glial cells and melanocytes.Partly supported by Conicyt (Grant 105) and Comisión de Investigación Científica Universidad de Chile (Grant 48). The technical assistance of Mr. Arnold van Dun is gratefully acknowledged. We also thank the Department of Physics, Faculty of Physical Sciences and Mathematics, University of Chile, for the use of a Philips EM-300 electron microscope.     

The structure of the coeliac ganglion of a teleost fish Myoxocephalus scorpius

Summary In order to compare the structure of a teleost sympathetic ganglion with those of other vertebrates, light, fluorescence histochemical and electron microscopy were carried out on the coeliac ganglion of the scorpion fish, Myoxocephalus scorpius. In common with studies on other vertebrates, fluorescence histochemistry distinguished two cell types: a) principal neurones which exhibited low levels of specific catecholamine fluorescence and comprise the majority of neurones in the ganglia, and b) smaller intensely fluorescent cells, some of which had processes tens of micrometers long.With the electron microscope, the principal cells were seen to make axodendritic and axosomatic synapses with axons containing mainly 30 nm agranular vesicles at the synaptic site while in other vertebrates usually only one or other synaptic association is present.Both the somata and the processes of intensely fluorescent cells contain 300–600 nm diameter vesicles many of which have electron dense cores. These cells are also innervated by axons containing 30 nm agranular vesicles.     

CORRELATION OF FINE STRUCTURE AND PHYSIOLOGY OF THE INNERVATION OF SMOOTH MUSCLE IN THE GUINEA PIG VAS DEFERENS

An electron microscope study of the innervation of smooth muscle of the guinea pig vas deferens was undertaken in order to find a structural basis for recent electrophysiological observations. The external longitudinal muscle coat was examined in transverse section. Large areas of the surfaces of adjacent muscle cells were 500 to 800 A apart. Closer contacts were rare. A special type of close contact suggested cytoplasmic transfer between neighbouring cells. Groups of non-myelinated axons from ganglia at the distal end of the hypogastric nerve ramified throughout the muscle. Some small axon bundles and single axons lay in narrow fissures within closely packed muscle masses. Many axons contained "synaptic vesicles." About 25 per cent of the muscle fibres in the plane of section were within 0.25 µ of a partly naked axon; of these 15 per cent were within 500 A of the axon, and about 1 per cent made close contact (200 A) with a naked axon. It is unlikely that every muscle fibre is in close contact with an axon, and it is not possible for every fibre to have many such contacts. Muscle fibres are probably activated by both diffusion of transmitter from naked portions of axons a fraction of a micron distant, and electrotonic spread of activity from neighbouring cells.     

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)     

Nerve ultrastructure of the arteries of the brain stem]

Ultrastructure of the nerve apparatus in the arteries of the brain base has been studied in cats. The structure of peri- and adventitial nerves has been investigated electron microscopically. Three types of efferent axons and four types of synaptic vesicles (small agranular and granular, large granular, large electron opaque vesicles) have been revealed. Vesicle-containing axons in the brain arteries approach the external smooth muscle cells of about 80 nm. Terminal axonal dilatations possessing direct and mediated connections with muscular cells of the middle tunica have been revealed.     

Substance P-containing axon terminals in the mucosa of the human urinary bladder: pre-embedding immunohistochemistry using cryostat sections for electron microscopy

The ultrastructure of substance P (SP)-containing axon terminals in the mucosa of the human urinary bladder was studied. Numerous SP-immunoreactive varicose nerve fibers were seen in the lamina propria, and most of them ran freely in the connective tissue. Many SP-immunoreactive nerve fibers were observed beneath the epithelium, and perivascular SP-immunoreactive nerves were also found in the submucosal layer. We observed a total of 305 SP-immunoreactive (IR) axon terminals, of which most (89.6%) were free nerve endings at the ultrastructural level; the rest of the SR-IR axon terminale were seen in the vicinity of the epithelium and blood vessels in the lamina propria. Varicose regions of SP-IR axon terminals contained large granular and small agranular synaptic vesicles, and most of them partially lacked a Schwann cell sheath. In some SP-IR varicosities, synaptic vesicles were concentrated in the region without any Schwann cell sheath. Long storage (for more than 1 month) of fixed-tissue pieces in sucrose before freezing has improved the ultrastructure of cryostat sections in pre-embedding immunohistochemistry. Trypsin digestion for the purpose of exposing antigenic sites was also employed before applying the first antiserum.     

Morphological changes in gland cells and axons resulting from stimulation of the salivary nerves of the Cockroach, Nauphoeta cinerea

The salivary glands of the cockroach, Nauphoeta cinerea (Olivier, 1789), are innervated and there is considerable evidence to suggest that dopamine is the neurotransmitter at the neuroglandular junction. As the gland is a bilaterally symmetrical structure it was possible to electrically stimulate the salivary nerve supplying the ipsilateral side of the gland whilst the contralateral side of the gland served as a convenient control. Saliva elicited from the glands by electrical stimulation of these nerves was collected and used to monitor the physiological state of the tissue. Glands were fixed for light and electron microscopy during secretion and it was observed that the ductules in peripheral acinar cells were distended in stimulated sides of the glands but not in contralateral unstimulated sides. This evidence implies that peripheral cells are responsible for the initiation of salivary fluid secretion. Changes were also observed in the catecholamine containing axons that innervate the glands. In stimulated axons a statistically significant reduction in numbers of small agranular vesicles was observed when compared with contralateral unstimulated controls and freshly fixed tissue. This was not the case with the larger granular vesicles of the same axons which showed no reduction in number as a result of stimulation. In addition it was also noted that the small agranular vesicles tended to aggregate and change their shapes in response to nerve stimulation. These results imply that the small agranular vesicles play a role in transmitter release.     

Ultrastructural evidence for exocytosis in the median eminence of the rat

Summary Exocytosis has been demonstrated by electron microscopy in the external zone of the median eminence of the rat. Exocytotic profiles have been observed in nerve fibres characterized by the presence of granular vesicles with median diameters of 90–103 nm and agranular vesicles of about 50 nm. In addition to the small agranular vesicles, coated vesicles of the same size have been found in many nerve fibres, suggesting that at least part of the agranular vesicles in the median eminence originate by micro-pinocytosis. The nature of the fibres showing exocytosis is discussed. Attention is drawn to the possibility of identifying types of fibres in the median eminence by the occurrence of exocytosis.The technical assistance of Mrs. R. M. Y. Hartsteen is acknowledged with gratitude. We would like to thank Prof. J. Moll for his helpful criticism. We also thank Miss P. Delfos and Mr. W. van den Oudenalder for photographic assistance.     

The effect of 17-B-estradiol on the fine structure of the adrenergic axons of the rabbit myometrium

Summary The effects of 17-B-estradiol on the fine structure of the autonomic nerves of the rabbit oviduct and myometrium were studied by means of KMnO₄-fixation. The main changes due to prolonged estrogen treatment were the following: (1) the dimensions of the axons increased, (2) the amount of smooth endoplasmic reticulum in the axons increased, (3) the amounts of both large granular vesicles (LGV) and large agranular vesicles (LAV) in the axons increased and (4) electron dense grains were found within the endoplasmic tubuli of the axons. Furthermore, it seems probable that the amount of small granular vesicles was also considerably higher after two weeks of estrogen treatment.The mechanism of estrogen action on the storage of transmitter within the axon is discussed. In conclusion, the present results suggest that both the axoplasmic transport and the peripheral formation of storage sites for noradrenaline are stimulated by the increased estrogen level.This work was supported by grants from Orion, Helsinki and from Finska Läkaresällskapet.     

Ultrastructural changes in the gracile nucleus of the spontaneously diabetic BB rat.

The present study describes the structural changes in the gracile nucleus of the spontaneously diabetic BB rat. At 3-7 days post-diabetes, axons, axon terminals and dendrites showed electron-dense degeneration. Degenerating axons were characterized by swollen mitochondria, vacuolation, accumulation of glycogen granules, tubulovesicular elements, neurofilaments and dense lamellar bodies. Degenerating axon terminals consisted of an electron-dense cytoplasm containing swollen mitochondria, vacuoles and clustering of synaptic vesicles. These axon terminals made synaptic contacts with cell somata, dendrites and other axon terminals. Degenerating dendrites were postsynaptic to normal as well as degenerating axon terminals. At 1-3 months post-diabetes, degenerating electron-dense axons, axon terminals and dendrites were widely scattered in the neuropil. Macrophages containing degenerating electron-dense debris were also present. At 6 months post-diabetes, the freshly degenerating neuronal elements encountered were similar to those observed at 3-7 days. However, there were more degenerating profiles at 6 months post-diabetes compared to the earlier time intervals. Terminally degenerating axons were vacuolated and their axoplasm appeared amorphous. It is concluded that degenerative changes occur in the gracile nucleus of the spontaneously diabetic BB rat.     

Ultrastructural investigation on the innervation of the Herbst corpuscle

Summary Nerve fibres, running longitudinally as well as circularly between the core lamellae in the Herbst corpuscle are described.These fibres are morphologically different from the central afferent axon. They are most frequently observed in the outer part of the core, and contain inter alia numerous agranular vesicles measuring approx. 450 Å in diameter, dense core vesicles with a diameter approx. 800 Å and microtubuli (250 Å). Occasional specialized junctions are seen between the nerves and the neighbouring lamellae.This study was supported by the Norwegian Council of Agricultural Research.