Morphometric study of the neuromuscular synapses in the adult rat with special reference to the remodelling concept

There is increasing morphologic evidence that neuromuscular synapses are not rigid structures in the mature muscles of adult animals. On the contrary, they may be submitted to a continuous process of remodelling. In silver-impregnated sternocleidomastoid muscles of the young adult rat, we measured synaptic parameters such as nerve terminal length, the number of branching points of terminal arborization, and muscle fiber diameter, and used a morphometric approach to explore specific questions concerning neuromuscular remodelling. Quantitative data indicate that: (a) The complexity and maturation of the nerve endings in this muscle are very variable and the increase in branching points is not paralleled by an increase in terminal length; (b) Muscle fiber diameter is related only marginally to presynaptic parameters; (c) Accessory ending formation occurs when the original ending does not reach the mean size of endings in singly innervated areas; (d) The complexity of individual endings at dually innervated junctions is smaller than the mean development of singly innervated synapses, indicating the existence of some mutual inhibitory influence between closely spaced endings. Morphometric results suggest a continuous process of synaptic formation in this adult muscle.     

Pattern of arborization of the motor nerve terminals in the fast and slow mammalian muscles.

A silver impregnation method and a morphometric approach were used to define differences existing in the motor nerve terminal branching pattern between a fast-twitch muscle (extensor digitorum longus) and a slow-twitch one (soleus) of the normal adult rat. Because no single measure can describe precisely all geometrical properties (ie both topology and metrics) of the nerve terminals, we evaluated morphologic parameters defining length and angular characteristics in the different terminal segments classified according to their centrifugal order. The main results indicate that the distal free-end segments in the extensor digitorum longus muscle are shorter and less divergent than in the soleus nerve terminals. The endings in the two muscles have different fractal dimensions. Findings are discussed in the context of the hypothetical mechanisms governing motor nerve terminal size and complexity.     

Complex nerve terminal formation in the phasic muscles of frogs

Synapses with complex nerve terminals consisting of several terminal arbors of a single axon divided by myelin segments were investigated using histological and electrophysiological techniques during experiments on the cutaneous-pectotoralis muscles of different aged frogs. Numbers of synapses with complex nerve terminals were shown to increase during the postnatal developmental process. The relationship between the complexity of nerve terminals, summated length of terminals, and size of muscle fiber is described. Some terminal arborizations at complex nerve terminals originate from nodes of Ranvier; these are marked by low quantal secretion and a distinctive pattern of sodium current decay along the path of the terminals. The causes and mechanisms governing increased complexity of nerve endings in phasic muscles are discussed, together with transmitter release patterns at these endings. It is postulated that growth and myelination processes occur in parallel at the nerve terminal.A. A. Ukhtomskii Physiological Institute, Leningrad State University; S. A. Kurashov Medical Institute, RSFSR Ministry of Health, Kazan'. V. I. Ul'yanov State University, Kazan'. Translated from Neirofiziologiya, Vol. 22, No. 1, pp. 99–107, January–February, 1990.     

Topological differences along mammalian motor nerve terminals for spontaneous and alpha-bungarotoxin-induced sprouting

Spontaneous sproutings can be observed in end plates from normal adult vertebrate muscles and motor end plates develop increased growth signs and sprouts when target muscle cells become less active or paralysed. Nevertheless, very little is known about where in the motor nerve terminal arborization spontaneous and experimentally induced sprouts originate, their similarities and differences and also about their final maturation or elimination. In this study we investigate the topological properties of both spontaneous and alpha-bungarotoxin-induced sprouts (during different periods of intoxication and after recovery) along the motor nerve terminal branches of the Levator auris longus muscle of Swiss mice (between 48-169 day old). Muscles were processed for immunocytochemistry to simultaneously detect postsynaptic AChRs and axons. This procedure permits us to make an accurate identification of the fine sprouts and a morphometric study of the presynaptic branching pattern profile in control muscles, during the toxin action and after recovery from paralysis. The results show that in normal muscles, the initial and trunk segments (those between branch points) of the terminal arborization sprouted proportionally more branches when taking their relative lengths into account than the distal free-end segments. In contrast, every micrometer of alpha-bungarotoxin-treated muscles throughout the full terminal arborization have the same probability of generating a sprout. Moreover, the toxin-induced sprouts can consolidate as new branches once recovered from the paralysis without changing the total length of the nerve terminal arborization.     

Activity-dependent plastic changes in the motor nerve terminals of the adult rat

Summary— Small and short-lasting physiologic variations in the locomotor activity of normal adult rats can induce remodelling in the motor nerve endings of the fast extensor digitorum longus muscle. The specificity and relative importance of the different plastic adaptations occurring in the presynaptic axonal tree have been studied, in silver impregnated nerve endings, by using an automatic image analysis treatment of the nerve terminals' geometric properties and a discriminant analysis of the morphometric parameters. Changes observed, like selective length variations in certain terminal segments and positional rearrangements, agree with a mechanism of neural connectivity regulation in the adult that arises as a consequence of normal neuromuscular activity.     

Modes of neuronal arbor enlargement in the ear of a postembryonic fish,Astronotus ocellatus

New hair cells are added during postembryonic life in several species of fishes and birds. The production of new hair cells appears to require enlargement of eighth nerve arbors during growth since, at least in fish, eighth nerve neurons are added more slowly than hair cells or not at all. This situation provides an intriguing opportunity to study the mechanisms of growth of the neuronal arbors. In this paper, we report the results of studies on the postembryonic growth of eighth nerve dendritic arbors in the saccular epithelium of the cichlid fish Astronotus ocellatus. Arbor sizes and shapes were compared in small and large fish using the axonal tracer cobaltouslysine. Our data suggest that postembryonic eighth nerve arbors enlarge in 2 ways. First, arbors add new terminal endings to their distal ends. Second, whole new branches appear to be added at locations up to hundreds of micrometers proximal to the terminal endings. These 2 modes of growth suggest that more than one mechanism may be operative in controlling arbor enlargement.     

Effects of lingual nerve section on morphometric characteristics of reticular nucleus neurons in the kitten brain stem

Computerized morphometric techniques were used to investigate each of 23 parameters in three types of brain stem reticular nucleus neurons in Golgi-stained frontal slices from the brain of 30-day-old kittens after uni- and bilateral lingual nerve section 5–7 days after birth. Particular statistically significant differences in some parameters were discovered in all types of cell. Certain group-specific differences in parameters could be most frequently distinguished in each category: distribution of loose dendritic endings through the dendritic area in reticular neurons, length of dendritic segments in branching cells, and distribution of foci of dendritic arborization in giant multipolar neurons. Unilateral lingual nerve section results in quantitatively more marked deviation from the normal state. It was only under these circumstances, moreover, that differences in overall length of dendrites could be seen, which could indicate a difference in the surface area of the cell.Institute of Higher Nervous Activity and Neurophysiology, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 23, No. 4, pp. 409–418, July–August, 1991.     

Effects of nerve growth factor on the development of the dendritic system of cholinergic neurons in dissociated culture of the rat septum]

Neurons dissociated from septal area of fetal (E18-19) rat brain were grown 14-days in culture. Cholinergic neurons were identified by cytochemical demonstration of acetyl cholinesterase. It was shown that the nerve growth factor added to the culture medium (50 u/ml) has increased the size of cell body of AchE-positive neurons, mean total length and arborization of dendrites and also the dendritic tree area.     

Data in favor of possible olfactory function of the antennal nerve and lateral lobe of protocerebrum of larva of the dragonfly Aeschna grandis

Using staining with methylene blue of larvae of dragonflies of the genus Aeschna sp. (2000 animals) the antennal nerve was shown to be connected with the lateral lobe of protocerebrum with a septum, through which sensory fibers enter the lobe. Near the lateral lobe of the antennal nerve, two enlargements are found, which contain motor neurons of antennal muscle as well as the incoming sensory fibers of antennal receptors and descending lateral bundles of fibers of lobes of mushroom bodies. In the lateral lobe of protocerebrum there is revealed arborization of neurons with terminal apparatuses similar to endings of the descending neuron of the glomerular antennal tract of the domestic fly.     

Morphometric analysis of hypoxia-induced synaptic activity in intrapulmonary neuroepithelial bodies

Summary A morphometric analysis has demonstrated ultrastructural changes induced by hypoxia in the epithelial cells and the intracorpuscular nerve endings of the presumed chemoreceptive intrapulmonary neuroepithelial bodies (NEB) of neonatal rabbits.Acute hypoxia stimulates an exocytosis of epithelial dense-core vesicles (DCV) at the level of the morphologically afferent or sensory (type 1 a) intracorpuscular nerve endings of the NEB. Assuming the epithelial cells to be chemoreceptive, this phenomenon could represent a transduction of sensory stimuli.In the morphologically efferent or motor (type 2 and type 1 b) intracorpuscular nerve endings of the NEB, acute hypoxia causes a depletion of synaptic vesicles and an increase in the amount of membrane-bounded cisternae and multivesicular bodies, suggestive of an enhanced synaptic activity of these nerve endings. It is proposed that the chemoreceptor cells could thus in turn be modulated centrifugally by their efferent-like intracorpuscular nerve endings.It has been proposed in our earlier studies that the NEB probably are intrapulmonary chemoreceptors with local secretory activities, reacting to the composition of the inhaled air. By the release of serotonin and peptide substances they may produce a local vasoconstriction in hypoxically aerated lung areas, enabling an intrapulmonary regulation of the V/Q ratio. The present study provides evidence that, in addition to this local effect, NEB could generate centripetal nerve impulses via exocytosis of epithelial DCV at the afferent-like intracorpuscular nerve endings. At the same time they could be modulated by the CNS via their efferent-like intracorpuscular nerve endings.With respect to their innervation, numerous similarities appear to exist morphologically and functionally between the carotid body and the intrapulmonary NEB.     

Further observations concerning the motor innervation of the tensor tympani muscle of the cat

The cat tensor tympani muscle presented an uncommon ultrastructural organization of neuromuscular junctions compared with those in the other striated muscles. In cross sections, individual neuromuscular junctions had very extended contact area of the nerve terminal and muscle fiber, the terminal bouton was covering as a "calyx" the postjunctional muscle fiber. Long basal lamina was interposed between them. The sarcolemma at the level of the nerve terminal had multiple infoldings along its length, or smooth postjunctional muscle membrane was found beneath endings on both fiber types.     

Ultrastructure of the joint receptors in the frog (Rana temporaria).

The ultrastructure of sensory nerve endings was examined in joint capsules of large limb joints in three adult frogs (Rana temporaria). The joint receptors are represented by the only one kind of sensory nerve endings--by free nerve endings. The unmyelinized preterminal desintegrates into single terminals. This branching is bound on the most peripheral cell of the Schwann cell by means of mesaxons, they pass from the pericaryum of the Schwann cell peripherally. The branches of the nerve terminal are surrounded by a cover of 1...3 cytoplasmatic processes of the Schwann cell. The surface lamella is covered by a distinct basal membrane. Bundles of collagenous fibrils pass along the branches of the nerve terminal. Quite naked nerve endings were not observed. The axoplasma of the nerve terminal contains strikingly few cell organels. Besides axially passing neurofilaments and neurotubules only sporadic mitochondria and clear vesicles were observed. The accumulation of mitochondria, characteristic for the axoplasma of nerve terminals, was observed in no case. Free nerve endings which were found in the joint capsules of the frog belong among so called "free penicillate nerve endings".     

Changes in the ultrastructure of Merkel's nerve endings of the sinus hair of rats after denervation

Merkel nerve endings of sinus hairs in the upper lip of the rat were observed after having cut the nervus infraorbitalis. The nerve terminal of the Merkel nerve ending has already been degenerated 24 hours after denervation and was phagocytised by neighbouring keratinocytes. The Merkel cells did not change in structure after having lost their nerve terminal even within 169 days after nerve crush. Their position in the stratum basale of the sinus hairs remained constant; the number, size and position of the osmiophilic granules in the cytoplasm of the Merkel cells did not change. These results may show, that Merkel cells are not modified keratinocytes. The possibilities of their origin are discussed.     

Quantitative studies of rat carotid body type I cell nerve endings

The results of a stereological and morphometric analysis of rat carotid body type I cell nerve endings are described. 66.9% of endings possessed symmetrical junctions. Of the remaining endings, 3.6% were presynaptic and 26% were postsynaptic to type I cells; 3.6% of endings had a reciprocal configuration. Apart from membrane specialisations, no other ultrastructural criteria were found to distinguish the different types of endings. Ventilation with 100% and 10% oxygen showed that the hypoxic mixture reduced synaptic vesicle concentration in the nerve endings; this effect was independent of the innervation to the carotid body.     

Structure of the sensory innervation of the anal canal in the pig. A light- and electron-microscopical study

The sensory innervation of the anal canal of the pig was investigated by light and electron microscopy. The distribution of the different types of sensory nerve endings correlates with the histology of different zones: (1) After the rectal mucosa there was a zone lined with nonkeratinized stratified squamous epithelium. (2) A middle zone was lined with keratinized stratified squamous epithelium. Here the dermis already showed a papillary and reticular layer. (3) The last zone showed hairy skin with a high hair density. The following nerve endings were found: Free nerve endings reached the stratum superficiale in nonkeratinized squamous epithelium and the stratum granulosum in the keratinized squamous epithelium. Dermal free nerve endings were found in all zones near the epithelium and two different types were identified as those derived from C-fibers and those from A-delta-fibers. Merkel nerve endings showed different features depending on their location. Few Merkel-like cells were found in the epithelium of the anal crypts. Typical Merkel Tastscheiben were located at the base of epithelial ridges or pegs in zones 2 and 3. The number of Merkel cells varied up to 200. The myelinated afferent fiber supplied 10-15 Merkel cells. Merkel cells were also found regularly in the outermost layer of the external rooth sheath of hair follicles at about the same level as perifollicular nerve endings. Lamellated corpuscles were found in the dermis of all zones except the cranial part of zone 1, where the anal crypts are located. Generally they consisted of a central nerve terminal which may be branched. Each terminal was surrounded by an inner core of concentrically arranged lamellae of the terminal Schwann cell and one or several inner cores were included in a capsule of perineural cells. The size of the corpuscle, the regularity of the inner core and the number of capsular layers depended on the location of the corpuscle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Morphology and chemical characteristics of subepithelial laminar nerve endings in the rat epiglottic mucosa

The laminar nerve endings are distributed in the laryngeal mucosa, and described as sensory receptors evoked by laryngeal pressure changes. The present study aimed to determine detailed morphological characteristics of the laryngeal laminar endings of the rat. Immunohistochemistry for Na(+)-K(+)-ATPase, α(3) subunit, showed that laminar endings were distributed in the entire laryngeal surface of the epiglottis. The parent axons of the endings were thick in diameter, and they were branched and continued to the endings. In some cases, several endings from different parent axons fused into a large complex structure of 500 μm in width. The laminar endings were also immunoreactive for vesicular glutamate transporter 1 (vGLUT1) and vGLUT2, but not for P2X(3) purinoceptor. Around the laminar endings, terminal Schwann cells with immunoreactivity for S-100 protein were closely associated with axon terminals. Use of scanning electron microscopy with alkaline maceration method showed that the terminal Schwann cells consisted of a rounded perinuclear region and lamellar cytoplasmic processes. Ultrastructurally, axon terminals with numerous mitochondria were partly covered with Schwann cell sheath, and some terminals intruded into the epithelial layer. Clear vesicles of 50 nm in diameter were also observed especially in small cytoplasmic processes of 400 nm to 1 μm in size. The results in the present study suggested that the laminar endings in epiglottic mucosa have morphological characteristics of slowly adapting mechanoreceptors and contribute to sensation of laryngeal pressure via mucosal tension.     

The fine structure of nerve endings on rat thyroid follicular cells

Summary Axon profiles in thyroid glands obtained from adult male Wistar rats were studied electron-microscopically, using common and serial thin sections.Bouton profiles of nerve fibers, resembling the terminal or en passant type, often appeared closely associated with vascular smooth muscle cells via basement membranes. These structures are probably adrenergic, since they contained mainly small-core vesicles (mean diameter: 41.2 nm), in addition to a few large-core (mean diameter: 88.4 nm) and flattened vesicles.Nerve fibers containing microtubules and sometimes mitochondria and vesicles were seen lying between basement membranes and follicular cells. The incidence of nerve fiber contacts on profiles of follicular cells was 0.0177±0.0092 (S.D.). Using serial sections, follicles were seen to have up to two nerve endings, separated from the plasma membranes of the follicular cells by a gap of 22 nm. They contained mainly flattened vesicles and several large-core vesicles (mean diameter: 95.1 nm). Small-core vesicles were rarely seen in these nerve endings. Furthermore, subsurface cistern-like rough endoplasmic reticulum was found immediately under the plasma membranes of follicular cells facing membranes of nerve endings. These results suggest that the nerve fibers in contact with follicular cells are different from the adrenergic type.     

Ultrastructural observation of possible nerve endings in rat sebaceous gland

Summary Neural elements within the parenchyma of the sebaceous gland have not been reported previously. Nerve endings have been observed only in the connective tissue surrounding the gland or in close association with the undifferentiated basal cells.In this study, electron microscopy revealed the possible presence of nerve endings (or terminal portions of neural elements) in the suprabasal level of functional sebaceous glands of pinnae of white rats. Morphologically, there are two distinct types of nerve endings. Type 1 is bordered by a membrane of relatively irregular contour and contains a single mitochondrion, various-sized vesicles, numerous microtubules, fine neurofilament-like fibrils, and occasional ribosome-like granules. Type II is also bordered by a membrane, but its contour was relatively smooth and rounded. Moreover, Type II contains many mitochondria, varying in size, density, and the arrangement of cristae. While ribosome-like granules are scattered throughout the structure in relative abundance, there are scarcely any fine neurofilament-like fibrils or microtubules. Whether these two structures are sensory or autonomic fibers could not be determined by electron microscopic examination.     

Calretinin-immunoreactive laminar nerve endings in the laryngeal mucosa of the rat

The distribution of laminar nerve endings that contained immunoreactive calretinin was examined in the laryngeal mucosa of the adult rat. In whole-mount preparations, the immunoreactive laminar endings were distributed in the supraglottic region but not in the subglottic region. The laminar endings that arose from thick nerve fibers with or without swellings were identified as corpuscles with many variform terminal arborizations. They appeared to be located at the interface between the epithelium and the subepithelial connective tissue. The terminals were scattered under the basal lamina of the epithelium, and some of them were located within the epithelial layer. Immunoelectron microscopy revealed that both sub- and intraepithelial immunoreactive terminals that were filled with mitochondria were partly or totally ensheathed by Schwann cell processes. The denervation experiments, in which the superior laryngeal nerve was cut unilaterally or bilaterally, suggested that the laminar endings originate from the superior laryngeal nerve with strict ipsilateral innervation. The laminar endings might be associated with detection of changes in pressure in the laryngeal cavity or chemical stimuli.     

Morphology and somatometric growth of long-tailed macaques (Macaca fascicularis fascicularis) in Singapore

Crab-eating, or long-tailed, macaques [ Macaca fascicularis (Raffles, 1821)] have been studied extensively throughout their distribution in South and South-east Asia. Despite this extensive body of research, the island population of long-tailed macaques from Singapore remains virtually undescribed. In the present study, we compare the morphometric variability and patterns of growth observed in a population sample from Singapore with a composite sample from Thailand, north of the Isthmus of Kra. The results of our analyses indicate that there are statistically significant differences between the two populations in adult size and shape. For both males and females, the Singapore population is smaller than the Thai population. Relative to body length, the Singapore macaques exhibit significantly longer tails, and, relative to cranial length, they exhibit significantly more narrow faces than the Thai macaques. Although levels of sexual dimorphism for most morphometric traits are very similar, indicating similar levels of male–male competition for females, the Singapore males exhibit a significantly larger testicular volume relative to body weight, suggestive of an alternative male reproductive strategy. In addition to adult somatometric size and shape, comparisons of growth patterns relative to age and body size reveal significant differences between the two population samples. Combined, these results suggest either that statistically significant differences in adult morphology and patterns of growth can occur in presumably reproductively cohesive subspecies, or the Singapore macaques may be taxonomically distinct.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 92 , 675–694.