Morphological correlates of electrotonic coupling in the magnocellular mesencephalic nucleus of the weakly electric fish Gymnotus carapo.

The magnocellular mesencephalic nucleus (MMN) of Gymnotus carapo was studied by electron microscopy. This particular nucleus, characteristic of weakly electric fish, contains two principal classes of neuron. (1) Large neurons (25-35 mum): these are rounded unipolar cells, with the perikaryon partially covered by a sheath of compact myelin. The axon leaves the neuron as a short thick unmyelinated process not resembling the initial segment of multipolar neurons. The axon branches profusely and becomes myelinated very close to its origin. The perikaryal surface not covered by the myelin sheath receives abundant club endings. The synaptic interface between club endings and large neurons is characterized by alternating gap junctions and attachment plaques. In addition, at the periphery of the club endings, "active" zones are generally present, and this synapse is therefore a "mixed" synapse. (2) Small neurons (5-12 mum): these are uni- or bipolar cells, scattered throughout the nucleus, and occasionally, grouped in small clusters. Gap junctions were not observed between neuronal perikarya in such clusters. The synaptic investment of small neurons is formed by long cup endings which almost completely encircle the perikarya. The synaptic interface between cup endings and the perikarya of small neurons is characterized by large areas of gap junctions. A single cup ending establishing gap junctions with two small neurons within the plane of the section was frequently observed and this arrangement provides a morphological basis for electrotonic coupling between small neurons by way of presynaptic fibres. In the neuropil of the MMN, there are abundant synaptic islands constituted by a large axon terminal in synaptic contact with small unidentified profiles; both synaptic elements are surrounded by numerous thin glial lamellae. At the synaptic interface, in the islands, both gap junctions and "active" zones are present. The synaptic islands must also be considered as "mixed" synapses. The morphological results presented here correlate with electrophysiological data (Szabo et al., 1975). The very short delay (0.8-1.3 ms) of the MMS response to the fish's own electric organ discharge can only be explained by the existence of electrotonic transmission along the neuronal chain of the electrosensory pathway. The presence of gap junctions between club endings and large neurons provides a morphological basis for electrotonic transmission at the mesencephalic level of the electrosensory pathway.     

Synaptic structure of the lateral line lobe nucleus in mormyrid fish

Electron microscopical observations reveal a complex synaptic structure (Fig. 1) in the nucleus of the lateral line lobe (nLLL). Different types of axosomatic and axoaxonic synapses are demonstrated to be in contact with the large cells. The results furnish morphological evidence for electrotonic transmission (by way of club endings with gap junctions) at this level of the electrosensory pathway of mormyrid fish. A new ultrastructural finding is the existence of presynaptic gap junctions on the unmyelinated surface area of the club endings.     

Electrophysiological study of the tangential vestibular nucleus of the chick embryo “in vitro”

Summary The two main neuron types of the tangential vestibular nucleus, the principal and the elongate cells, were investigated using intracellular recordings on brain slice preparations of 15–16 day chick embryos. Their identification was confirmed by intracellular injections of the fluorescent dye, Lucifer yellow. The passive membrane properties of these two neuron types were not sufficient criteria for their identification, whereas their responses to current pulse injections were distinctive. The elongate cells showed a nonlinear current-voltage relationship to hyperpolarizing currents and they fired repetitively at high frequencies without accommodation when the cells were depolarized. Neither of these two properties were observed in principal cells. Synaptic responses to vestibular nerve stimulation were different for the two neurons. The elongate cells responded gradually to increasing stimulus intensity, while the principal cells exhibited two clear post-synaptic responses: a low-threshold action potential initiated by an all-or-none EPSP, and a second, higher threshold, longer latency and graded EPSP. The latter EPSP is characterized by a slow rise time and a long duration. The findings obtained at this embryonic age are consistent with data concerning principal and elongate cell innervation by different caliber fibers comprising the vestibular nerve. The early EPSP recorded from the principal cells is attributed to the colossal fibers, which are vestibular afferents of large diameter forming calyciform contacts, the spoon endings, exclusively on the principal cells. The other synaptic responses recorded from the elongate and principal cells are due to the firing of different classes of small vestibular afferents. These experiments were performed in this new preparation at a critical age for the development of the spoon endings. In the 15–16 day embryo, gap junctions make their first appearance at the spoon endings joining the chemical synapses already present. According to our data, transmission at the onset of morphologically mixed synapses is monophasic suggesting that only one mode of transmission prevails.     

The relationship between some measures of synaptic ultrastructure as a function of distance from the soma on lamprey reticulospinal neurons

Summary Synaptic junctions located on the dendrites of lamprey (Petromyzon marinus) reticulospinal neurons labelled with intracellularly-injected horseradish peroxidase were studied. The normal ultrastructure of the synaptic junctions was defined and several quantitative measures made from each junction in order to test the hypothesis that distally-located synapses are ultrastructurally different from those located at proximal dendritic sites. A total of 820 contacts from one neuron and 279 from a second neuron ranging from 20 to 340 m from the soma were quantified. The vast majority of the presynaptic endings contained round, clear-cored vesicles and formed an asymmetrical membrane differentiation with the postsynaptic dendrite. A small fraction of the population contained flattened or pleomorphic vesicles and these synapses were equally distributed with respect to distance from the soma. Many of the terminals contained a few large dark- and clear-cored vesicles. Four quantitative measures of each synaptic contact were made. These included vesicle number, length of differentiated membrane, vesicle area and terminal area. Four ratios relating the different quantitative measures were also calculated. Each ratio or measurement from the synaptic junctions was plotted as a function of distance from the soma to determine if differences existed at any distance. It was found that synaptic junctions are uniformly similar and that distal junctions did not differ significantly (P > 0.05) from those at proximal dendritic sites. It is concluded that if distal synapses do compensate for their remote location they do this in some other way, possibly by increasing the number of synaptic contacts made by each presynaptic axon.     

Electron microscopy of synapses in reptile spinal cord

The spinal cord of the reptile Anolis carolinensis was examined by electron microscopy. Motor neurons appear as multipolar cells 30-60 micrometer in diameter. Two types of synaptic endings are endings are present on motor neurons. The first type is characterized by distinct synaptic clefts measuring 15-20 nm between pre- and postsynaptic membranes, and by clear presynaptic vesicles. The second type of synapse, which is less common, is characterized by gap junctions between pre- and postsynaptic membranes. At these synapses, there are also clusters of clear vesicles close to the presynaptic membrane adjacent to the gap junction. These findings indicate that both chemical and electrical synaptic transmission are present in the spinal cord of Anolis.     

Dual transmission at morphologically mixed synapses: Evidence from postsynaptic cobalt injections

Two experimental approaches have been utilized to test the possibility that morphologically mixed synaptic terminals of the eighth nerve fibers mediate both electrotonic and chemical excitation of the goldfish Mauthner cell. First, the spatial distributions of electrotonic and chemical postsynaptic potentials, evoked by stimulation of the eighth nerve, have been determined with intracellular recordings from the Mauthner cell soma and several locations along the lateral dendrite. In some instances, both synaptic components were maximal at distal dendritic recording sites. In that region, it appears that the only presynaptic terminals with morphological characteristics consistent with excitatory chemical transmission are the large myelinated club endings, which actually establish mixed synapses with the lateral dendrite. Second, we have analyzed the effects of postsynaptic Co²⁺ injections on these synaptic responses. With high iontophoretic currents, there was a rapid uncoupling of the electrotonic component. However, with smaller current intensities, uncoupling is accompanied, or preceded, by a transient reduction in the later chemically mediated postsynaptic potentials. This latter effect on chemical transmission is only observed if the postsynaptic potentials are associated with electrotonic synaptic inputs. We speculate that Co²⁺ diffuses across the gap junctions and into the presynaptic terminals, acting there to reduce evoked transmitter release.The results of these two experimental approaches support the hypothesis that mixed synapses on the lateral dendrite of the Mauthner cell do actually mediate transmission by both chemical and electrical modes.     

The mormyrid brainstem--III. Ultrastructure and synaptic organization of the medullary "pacemaker" nucleus

The ultrastructure and synaptic organization of the presumed medullary pacemaker nucleus, nucleus c of the weakly electric mormyrid fish, Gnathonemus petersii has been investigated. Nucleus c consists of about 12-15 small (20-25 micron) neurones (P-cells), which form a group situated ventrally to the medullary relay nucleus and embedded in a neuropil of myelinated fibres and dendritic processes. The P-cells often exhibit an enhanced electron density of their cytoplasm and dendroplasm. They possess several dendrites of different diameter, a short, thin axon initial segment and a thickly myelinated axon running in dorsal direction. The pacemaker neurons are interconnected by complex electronic coupling, established by somatosomatic, dendrosomatic and dendrodendritic gap junctions. Perikarya and dendrites are frequently interconnected serially by gap junctions; dendrites showed sometimes triadic gap-junction arrangement. It is suggested that this high degree of electrotonic coupling amongst the pacemaker cells represents the first level of the highly ordered synchronization processes which characterize the electric discharge command system of Gnathonemus. Pacemaker cells receive synaptic input from club endings with mixed synapses and from bouton-like terminals with chemical synapses, both of them originating from medium-sized myelinated fibres and contacting mainly neuronal perikarya and dendritic processes. The axon initial segment receives only few synaptic inputs. Bouton-like terminals were found to be of two types according to their vesicle content, namely, boutons with ovoid, clear synaptic vesicles forming Gray type-1 synapses and boutons with pleomorphic clear synaptic vesicles forming Gray type-2 synapses. Different functional roles for the two types of boutons in modulating pacemaker cell activity are suggested.     

Neuro-epitheliomuscular cell and neuro-neuronal gap junctions inHydra

Summary Gap junctions have been described ultrastructurally between neurons and epitheliomuscular cells and between neurons and their processes in the hypostome peduncle and basal disc ofHydra. All gap junctions examined inHydra exhibit two apposed plasma membranes having a 2–4 nm gap continuous with the extracellular space. The gap junctions are variable in length from 0.1–1.6 m and appear linear or V-shaped in section. Neuronal gap junctions inHydra occur infrequently as compared to chemical synapses. Electron microscopy of serial sections has demonstrated the presence of adjacent electrical and chemical synapses (neuromuscular junctions) formed by the same neuron. In addition multiple gap junctions were present between two neurons. This is the first ultrastructural demonstration of electrical synapses in the nervous system ofHydra. Such synapses occur in neurons previously characterized as sensory-motor-interneurons on the basis of their chemical synapses; these neurons appear to represent a type of stem cell characterized by having both electrical and chemical synapses.     

Neuronal and synaptic organization of the outer plexiform layer of the pigeon retina

The organization of the outer plexi-form layer (OPL) of the pigeon retina is described by electron microscopy and Golgi impregnation. Six types of photoreceptor, four types of horizontal cell, eight types of bipolar cell, and an interplexiform cell type were found by Golgi impregnation. The OPL was tri-stratified due to the endings of the photoreceptors at three different levels. This stratification was reflected in the laminar arrangement of the dendrites of the horizontal and bipolar cells. Electron microscopy showed that the synaptic endings of the photoreceptors made ribbon synapses, both triads and dyads, and basal junctions with the process of second-order neurons. Horizontal cells formed conventional chemical synapses, while horizontal cell axon terminals were extensively linked by gap junctions.     

Synaptology of the command (pacemaker) nucleus in the brain of the weakly electric fish, Sternarchus (Apteronotus) albifrons

The high frequency electric emission of the weakly electric fish Sternarchus (Apteronotus) albifrons depends on the pacemaker activity of a specific brainstem nucleus located in the ventral part of the rhombencephalic reticular formation. The general morphology and fine structure of this nucleus has been investigated, with particular reference to its synaptic connections.Three neuronal components could be distinguished in the nucleus; namely large cells of 80–100 μm diameter, small cells of 30–50 μm diameter and bundles of thin, myelinated fibres. These elements are embedded in a network of thick myelinated fibres. The large cells have a few small and short dendrites whereas the small neurons have long branching dendrites. Large and small neurons possess thick myelinated axons, but only those of the latter show branching patterns and send collaterals which have intranuclear courses only. Two types of synaptic terminals have been found on both neurons: large club endings exclusively with gap junctions and small bouton-like terminals with polarized chemical synapses. Serial semi-thin and ultra-thin sections revealed that the large club endings belong to the pacemaker cells, whereas the small terminals are found in the thin myelinated axons of extranuclear origin.The findings indicate that the small neurons are connected 1) to each other and 2) to the large neurons, by way of their large myelinated axons. Both, small (pacemaker) as well as large (relay), neurons receive chemical synapses from myelinated fine fibers probably originating from higher encephalic centers. Thus, electric organ discharge rhythm can be modulated at the level of pacemaker as well as of the relay cells. No somatosomatic, dendrodendritic or dendrosomatic connections were found between large, small or large and small cells.     

The mormyrid brainstem--II. The medullary electromotor relay nucleus: an ultrastructural horseradish peroxidase study

The medullary relay nucleus of the mormyrid weakly electric fish Gnathonemus petersii is a stage in the command pathway for the electric organ discharge. It receives input from the presumed command or pacemaker nucleus and projects to the electromotoneurons in the spinal cord. Its fine structure and synaptology were investigated by electron microscopy. The origin of the terminals contacting the cell membrane of the neurons of this nucleus was determined by horseradish peroxidase (HRP) injections into different brain structures, namely into the bulbar command- and mesencephalic command-associated nuclei. Twenty-five to thirty large cells of about 45 micron in diameter constitute the medullary electromotor relay. Each cell has a kidney-shaped, lobulated nucleus, a large myelinated axon with a short initial segment and several long, richly arborizing primary dendrites. Many, if not all, cells are interconnected with large somatosomatic or dendrosomatic, dendrodendritic and dendroaxonic gap junctions. These junctions often occur in serial or triadic arrangements. The relay cells receive large club endings as well as small boutons. The club endings are found mainly on the soma and primary dendrites and are morphologically mixed synapses. The boutons are characterized by synapses which are only chemical and are distributed all over the cell membrane, but with a definitely higher frequency on secondary dendrites and more distal parts of dendritic processes. Horseradish peroxidase injections into the mesencephalic command-associated nucleus reveal a large number of labelled boutons on the secondary dendrites of the relay cells. Injections into the bulbar command-associated nucleus label the same type of boutons as mesencephalic injections, but also label club endings on relay cell soma and primary dendrites. The results support the conclusion made on the basis of previous light microscopical observations that boutons originate from the bulbar command-associated nucleus, whereas the club endings issue from the presumed pacemaker nucleus (nucleus c). The club endings of the bifurcating axons of this nucleus are labelled by retro- and anterograde transport of horseradish peroxidase; the bifurcating axons project simultaneously to the bulbar command-associated nucleus and the medullary relay nucleus.     

Anatomical study and HRP identification of electromotoneurons and motoneurons in the spinal cord of Gymnarchus niloticus

Two types of neurons were identified by light and electron microscopy in the spinal cord of Gymnarchus niloticus following HRP injections at different peripheral sites: one, situated in the mediodorsal region, is labelled after injection into the electric organ; the other, situated in the lateroventral region, is labelled in addition after injections extending beyond to the lateral muscle. The cells of the first type, thus shown to be electromotoneurons (EMNs), are spherical and do not have dendritic processes; some of them are connected by somato-somatic gap junctions. The EMNs are surrounded by dense glial processes and embedded in a network of myelinated axons of large diameter. Synaptic contacts with the EMNs are of axosomatic type, exhibiting mostly gap junctions and, less frequently, mixed or chemical junctions. Some endings establish gap junctions simultaneously, with two electromotoneurons, the somatic membrane of which may be joined. The initial segment does not bear any endings. The second type of cells, identified as motoneurons, are pyriform and have large dendritic processes. The motoneurons are surrounded by myelinated axons of small diameter. They receive axosomatic endings with chemical synapses.     

Electron microscopic observations of the mesencephalic nucleus of the fifth nerve in the Selachian brain

Summary The mesencephalic nucleus of the trigeminal nerve (mes V) in the brain of the skate (Raja oscellata) was studied by electron microscopy. Mes V neurons are large (40–80 m diameter) and are located in the periventricular grey matter. Their perikaryal cytoplasm is rich in Golgi apparatus, small mitochondria, rough endoplasmic reticulum, polysomes and bundles of neurofilaments. A striking feature is the presence of masses of glycogen granules, at times surrounded by membrane wrappings and lysosomal bodies.Two types of conventional synaptic contacts were made onto mes V perikarya and dendrites. One had round, agranular vesicles and usually also contained dense-cored vesicles, the other had flattened, pleomorphic, agranular vesicles and usually lacked dense-cored vesicles. Additional membrane complexes consisting of a region of gap junction flanked by sites of desmosomal attachment were observed to link neighbouring mes V neurons. Somato-somatic, dendro-somatic, axo-somatic, and dendro-dendritic junctions were noted. Except for the somato-somatic union, one or more chemical synapses were located close to the sites of gap junctions.     

Synaptology of the medullary command (pacemaker) nucleus of the weakly electric fish (Apteronotus leptorhynchus) with particular reference to comparative aspects

Summary The general organization and synaptology of the medullary command (pacemaker) nucleus (MCN) was investigated in the high frequency weakly electric fish, Apteronotus leptorhynchus. This study was undertaken in order to establish differences and similarities between the MCN of A. leptorhynchus and that of the closely related species, Apteronotus albifrons which has been studied previously. The basic morphology and synaptology of the MCN in A. leptorhynchus is similar to that of A. albifrons. The MCN of A. leptorhynchus consists of large (relay) and small (pacemaker) cells; both cell types receive synaptic input or large club endings with electrotonic gap junctions and bouton-like terminals with polarized chemical synapses. Club endings originate from thick meyelinated fibres belonging to the small (pacemaker) cells, whereas the bouton-like terminals issue from thin myelinated fibers of extranuclear origin. Via their club endings, the small (pacemaker) cells are connected both to each other and to the large (relay) cells. Besides the similarities, there are distinct and characteristic differences between the MCN of the two species, which mainly concern the synaptology of the nucleus. In A. leptorhynchus, the large (relay) cells possess long dendritic processes, covered exclusively with bouton-like terminals; the axon initial segment of large (relay) cells receives boutons, in addition to club endings. Small (pacemaker) cells have short dendritic protrusions receiving input from club endings and boutons; furthermore, the small pacemaker cells axon initial segment receives both club endings and bouton-like terminals. These differences are discussed in terms of the functional organization of the MCN in certain gymnotoids and draw attention to the fact that the morphological and ultrastructural aspects of the central command of the electric organ discharge reveal several differences not only between different gymnotoid fish (Apteronotus and Eigenmannia) but also between closely related species such as A. albifrons and A. leptorhynchus.     

The cholinergic innervation of the rat substantia nigra: a light and electron microscopic immunohistochemical study

Summary Putative cholinergic axons and synaptic endings were demonstrated in the substantia nigra (SN) of the rat by light and electron microscopy on the basis of the localization of choline acetyltransferase (ChAT) immunoreactivity. The distribution of ChAT immunoreactivity in the SN as demonstrated by light microscopy revealed a modest network of ChAT-immunoreactive beaded axons in the SNc, in comparison to a relatively sparse distribution in the SNr. These axonal profiles were most dense in the middle of the rostral-caudal extent of the SNc and appeared to be concentrated in the middle third of the medial-lateral extent. By electron microscopy, unmyelinated, small diameter (0.25 m) ChAT-immuno-reactive axons were observed interspersed among numerous other non-immunoreactive axons in the SNc. ChAT-immunoreactive synaptic endings were observed in juxtaposition to small caliber (0.5 m) non-immunoreactive dendrites, and contained numerous spheroidal synaptic vesicles and occasional mitochondria. Synaptic contact zones were characterized by an accumulation of synaptic vesicles along the presynaptic membrane, and a prominent postsynaptic densification producing an asymmetrical pre-/postsynaptic membrane profile typical of excitatory synapses. These findings provide direct evidence for a cholinergic innervation of the SN, and suggest that this input may have an excitatory effect on neuronal elements in the SNc.     

Junctional systems in the pineal gland of the Wistar rat (Ratus ratus). A freeze-fracture and thin section study

Intercellular junctions between neighbouring pinealocytes, glial cells, glial cells and pinealocytes as well as between nerve endings and parenchymal cells of the pineal gland of Wistar rats were investigated on freeze-fracture replicas and thin sections by transmission electron microscopy. Gap junctions, tight junctions and the annular gap junctions have been revealed. In addition, chemical synapses between nerve endings and pinealocytes have been observed.     

Serial and triadic synapses in the cerebellar nuclei of the cat

Summary A quantitative electron microscopic investigation of the nucleus interpositus in cat cerebellum reveals that about 1.5% of all observed synapses are established between synaptic vesicle-bearing profiles. It is shown by serial sections that 70% of these synaptic complexes are triadic arrangements and 30% are serial synapses. Further analysis discloses that the first presynaptic element in the triadic and serial synapses may be one of four different axonal types: (A) Purkinje-cell axons; (B) and (C) afferent fibers containing large round vesicles and originating from the brain stem (probably mossy and climbing fiber collaterals); and (D) axon terminals containing small round vesicles. Indirect evidence suggests that type D profiles are the recurrent axon collaterals of the projective neurons. The second, postsynaptic and presynaptic, vesicle-bearing process in these complexes is either a class D terminal, or a somewhat more dendrite-like profile (Class E) containing flattened vesicles, and identified as belonging to processes of local Golgi type II interneurons.     

Identification under the electron microscope of climbing fibers and their synaptic contacts

Summary An attempt is made to identify, under the electron microscope, the climbing fibers of the cerebellum (in the cat) and their synaptic contacts with Purkinje cells and other cortical neurons. — Two kinds of axonal profiles, having synaptic contacts with primary and secondary dendrites of Purkinje neurons, can be recognized: One being terminal fibers densely packed with neurofilaments, having mainly contacts de passage with the dendrite surface, with small accumulations of synaptic vesicles at the presynaptic side of the contact. The others are rather knob-shaped contacts filled with synaptic vesicles and poor in neurofilaments. In chronically isolated folia, in which only local neurons and their processes have survived, all filamentous profiles have disappeared while vesicular ones are not appreciably reduced in number. It is inferred from this, that the neurofilamentous profiles correspond to climbing fibers, whereas the vesicular ones could be the endings of outer stellate axons, recurrent Purkinje axon collaterals, or ascending basket axon collaterals. — Similar two kinds of axon-terminal profiles are found in synaptic contact with Golgi and basket cell bodies. As in chronically isolated folia only the vesicular profiles survive, it is inferred that the climbing fiber has axo-somatic terminals on Golgi cells and basket cells as well. Previous information of this kind, gained with the light microscope and with degeneration studies, is thus substantiated with the aid of the electron microscope. The vesicular presynaptic profiles on Golgi and basket neurons are in the first case certainly and in the second with high probability endings of recurrent Purkinje axon collaterals. — The few axosomatic synapses found on outer stellate neurons may also be terminals of climbing fibers, but degeneration evidence for this is not conclusive. — The observations are summarized and evaluated from the functional point of view in a diagram, with consideration to recent physiological information on the function of climbing fibers.     

The organization of the outer plexiform layer in the retina of the cat: electron microscopic observations

Summary The outer plexiform layer of the cat retina has been examined by electron microscopy of random and serial ultrathin sections in order that neural profiles might be positively identified and their synaptic relationships studied. Photoreceptors are interconnected by means of gap junctions as are the A horizontal cells. B horizontal cells and axon terminals do not appear to be engaged in any synapses apart from those with photoreceptors, while A horizontal cells make rare junctions with cone bipolars only. Interplexiform cell processes probably account for all the conventional chemical synapses in the outer plexiform layer of cat retina.