Round Vesicles Research Articles

Synaptic organization of the interstitial subdivision of the nucleus tractus solitarii and of its laryngeal afferents in the rat

The nucleus tractus solitarii, the first central relay for gustatory and a variety of visceral afferents, is also an integrative center for numerous functions. Its interstitial subdivision is involved in swallowing and respiratory reflexes. The ultrastructural characteristics of this subdivision and of its laryngeal afferents were investigated in adult rat by a serial-section study and by application of wheat germ agglutinin-horseradish peroxidase conjugate to the peripheral afferent fibers. The interstitial subnucleus contained scattered small neuronal cell bodies with such ultrastructural features as a large nucleus with deep indentations and an organelle-poor cytoplasm. On the basis of their size and vesicular content, the axon terminals were classified into three categories. Group I and group II terminals were small or large, respectively, and contained mainly small, round, and clear synaptic vesicles. Group III terminals were also small but contained small, pleomorphic, and clear vesicles. Axodendritic synapses were the most numerous. They were either asymmetrical, comprised of group I and II terminals, or symmetrical, comprised of group III terminals. More than 50% were part of complex synaptic arrangements in the form of rosettes or glomeruli. Axosomatic contacts involved both group I and group III terminals and were always symmetrical. A high frequency of axoaxonic synapses was found. They were symmetrical, comprised of group III terminals on group I or II terminals. Different types of symmetrical synaptic contacts made by dendrites were also found. This study indicates also that the ipsilateral interstitial subdivision constitutes the preferential site of termination for superior laryngeal afferents. The labeled axon terminals belonged exclusively to groups I and II and were involved in both axodendritic and axoaxonic synapses. Some of the axodendritic synapses were part of rosettes or glomeruli. All these synaptic arrangements may be considered a morphological substrate for important processing of afferent information in the nucleus tractus solitarii. They may account for some of the integrative functions of the interstitial subnucleus such as physiological processes triggered from the superior laryngeal nerve.

Glutamate‐immunoreactivity in identified vagal afferent terminals of the cat: a study combining horseradish peroxidase tracing and postembedding electron microscopic immunogold staining

Using electron microscopic immunohistochemistry we have shown that strong glutamate-immunoreactivity (glutamate-ir) is present in neuronal cell bodies of the nodose ganglion, axons in the tractus solitarius and afferent terminals in the nucleus tractus solitarii. Vagal afferent fibres were specifically labelled by transganglionic retrograde transport of horseradish peroxidase (HRP). Fifty-seven per cent of the HRP-labelled terminals in the dorsomedial medulla were found to contain a high level of glutamate-ir, suggesting that a population of vagal afferent fibres uses glutamate as a neurotransmitter substance. There were no apparent ultrastructural differences between glutamate-ir and non-glutamate-ir vagal afferent terminals, both classes mainly containing rounded vesicles and forming asymmetric synapses. However, some difference in their preference for postsynaptic target was noted. The great majority (83%) of non-glutamate-ir vagal afferent terminals made axodendritic synapses, but only just over half (57%) of the glutamate-ir vagal terminals made synaptic contact with dendrites. Approximately 13% of the HRP-labelled terminals were found to make synaptic contact with HRP-labelled dendrites or soma of motoneurones of the dorsal vagal motor nucleus, confirming the existence of monosynaptic connections between vagal afferent fibres and vagal motoneurones.

Distribution of GABA immunoreactivity in the retino-recipient layer of the viper optic tectum. A light and electron microscope quantitative study.

The distribution of GABA-immunoreactivity was investigated in the principal retino-recipient layer of the optic tectum in the snake Vipera aspis. This layer, the stratum griseum et fibrosum superficiale, contained an important proportion (approximately 50%) of small GABA-immunoreactive interneurons, characterized by a voluminous invaginated nucleus surrounded by a thin rim of cytoplasm poor in organelles and occasionally showing pleiomorphic synaptic vesicles, which could also be observed in some of the dendrites that contained synaptic vesicles. In the neuropile, the GABA-immunoreactive profiles containing synaptic vesicles could be subdivided into dendrites containing synaptic vesicles and axon terminals with pleiomorphic synaptic vesicles. The dendrites containing synaptic vesicles (23.4% of all profiles containing synaptic vesicles) were postsynaptic either to optic terminals (39.2%), GABA-immunoreactive axon terminals with pleiomorphic synaptic vesicles (48.2%) or to immunonegative (S1) boutons with round synaptic vesicles (12.6%). These dendrites were presynaptic to GABA-immunoreactive (18%) neurons or immunonegative (82%) neurons. The axon terminals with pleiomorphic synaptic vesicles, which represented 47.4% of all profiles, were predominantly (99%) GABA-immunoreactive and four types could be distinguished according to cytological criteria. These axon terminals made synaptic contacts for the most part (78%) with immunonegative profiles, and more rarely (22%) with immunoreactive neurons. These data are compared to those previously obtained in the homologous structure of other vertebrate species, birds and mammals in particular.

Ultrastructural parameters of GABA-ergic and non-GABA-ergic synaptic contacts on neurons of the catsubstantia nigra

Nigrothalamic neurons were identified in the reticular part of thesubstantia nigra using labelling by the retrograde axonal transport of horseradish peroxidase. Nine parameters of the synaptic contacts (n=195) were analyzed, including the size and shape of terminals and size and type of synaptic vesicies. Sixty-six percent of axon terminals studied formed symmetric contacts and contained large polymorphic vesicles (group I). Two-thirds of these synapses were located on the distal dendrites, while one-third was distributed on the perikarya and proximal dendrites. Synapses of group II (29% of all synapses analyzed) exhibited asymmetric contacts and contained round agranular vesicles. Among these synapses, 79% were located on the distal dendrites, 19% were located on the proximal dendrites, and only 2% were located on the neuronal perikarya. Axon terminals of group III (5% of total population) exhibited symmetric contact and contained small polymorphic vesicles. High-resolution immunogold EM histochemistry indicated that 63% of the group-I axon terminals were GABA-positive. The majority of synapses on the labelled nigrothalamic neurons (21 contacts of 25) belonged to group I. Among these 21 synapses, 19 were axo-somatic and mostly GABA-positive. Within group II, 30% of synapses showed slightly expressed GABA-positivity.

Ultrastructural and immunocytochemical studies of neuromuscular junctions in oviduct of Locusta migratoria

The ultrastructure of nerve endings in the oviduct visceral muscles of Locusta migratoria was studied by electron microscopy and by immunogold labeling for two kinds of neuromodulators, the pentapeptide proctolin and FMRFamide-related peptides. Nerve endings contained electron-lucent round vesicles and two kinds of granules (round and avoid), and formed two types of synapses or release sites with the muscle. The morphologically distinct nerve endings were classified into three different categories based on the composition of synaptic vesicles and granules. Type-I nerve endings were dominated by electron-lucent round vesicles and contained only a few round electron-dense granules. Type-II nerve endings contained mostly electron-dense round granules and electron-lucent round vesicles. A few electron-dense ovoid granules were also present. Electron-dense ovoid granules dominated the type-III nerve endings, which usually contained less electron-lucent vesicles than either type-I or II nerve endings. Both proctolin and FMRFamide-like immunoreactivity was associated with electron-dense round granules. However, FMRFamide-like immunoreactivity was only found in the type-II nerve endings, while proctolin immunoreactivity was found within type-I nerve endings as well as in some type-II nerve endings. Immunological results therefore allow us to further divide type-II nerve endings into type-IIa (immunonegative for proctolin) and type-IIb (immunopositive for proctolin). Type-III nerve endings show no immunolabeling to either proctolin or FMRFamide.

Midbrain paralemniscal projections to the facial nucleus: an anatomical and immunohistochemical study.

Serial 30 microns-thick sections through the midbrain tegmentum were stained with cresyl violet. The PL was found to be situated along the medial edge of the lateral lemniscus. The PL consisted of small- (10-15 microns) and medium-sized neurons (25-35 microns), and was the most prominent at the caudal level of the superior colliculus. In order to confirm the existence of the inhibitory paralemniscal-facial pathway, a combined HRP and immunohistochemical technique was use in the rat. This experiment revealed that 10.9% of the total number of GABA immunoreactive PL neurons also labeled with HRP after HRP injection was made in the medial part of the facial nucleus (FN). Electron microscopic observations were carried out on the medial part of the facial nucleus (FN) after kainic acid injection was made into the contralateral PL in the cat. The majority of degenerating PL fibers were ranged from 0.5 to 3.1 microns in diameter and made synaptic contacts with somata, proximal dendrites and dendritic profiles. These fibers, containing either round or pleomorphic vesicles, formed asymmetrical or symmetrical synapses. It was of particular interest in the present study that 40.7% of the total number of degenerating fibers make synaptic contacts with large dendrites more than 3.0 microns in diameter.

Cerebellar-responsive neurons in the thalamic ventroanterior-ventrolateral complex of rats: Light and electron microscopy

The morphology and synaptic organization of neurons in the ventroanterior-ventrolateral nucleus of rats was examined using in vivo intracellular staining techniques. Neurons were characterized electrophysiologically based on intrinsic membrane properties and synaptic responses to stimulation of motor cortex and cerebellar nuclei, as described in the companion paper. Cerebellar-responsive neurons were stained intracellularly with either horseradish peroxidase or biocytin. All stained ventroanteriorventrolateral nucleus neurons were identified as thalamocortical neurons on anatomical (and often electrophysiological) grounds, consistent with previous findings that rat ventroanterior-ventrolateral nucleus is interneuron-sparse. Ventroanterior-ventrolateral nucleus neurons had three to eight thick primary dendrites. Proximal dendrites often exhibited a tufted branching pattern, from which many thinner, higher order dendrites arose. Dendrites branched to form a funnel-like infiltration of the neuropil that resulted in a spherical, roughly homogeneous dendritic field. The axon originated from the cell body or a proximal dendrite and coursed laterally and dorsally to innervate motor cortex. One to five axon collaterals were emitted in the rostral dorsolateral sector of the thalamic reticular nucleus; collaterals were not observed in the ventroanterior-ventrolateral nucleus or other nuclei in dorsal thalamus. The synaptic organization of the ventroanterior-ventrolateral nucleus was examined with electron microscopy, including two intracellularly labeled ventroanterior-ventrolateral nucleus neurons that were shown electrophysiologically to receive monosynaptic inputs from the cerebellum. The neuropil of rat ventroanterior-ventrolateral nucleus lacked the complexity and diversity found in corresponding thalamic nuclei of felines and primates, due to the paucity of interneurons. Vesicle-containing dendrites, dendrodendritic synapses and glomeruli were not observed. Three broad classes of presynaptic terminals were identified. (1) Small round boutons: small boutons containing densely-packed, small round vesicles that formed asymmetric synapses predominantly with the distal dendrites of thalamocortical neurons. These were the most prevalent type of bouton in the ventroanterior-ventrolateral nucleus (78% of presynaptic elements) and likely arose from the cerebral cortex. (2) Large round boutons: large terminals with loosely packed small round vesicles that made multiple asymmetric synapses with proximal and intermediate dendrites. Large round boutons comprised 8% of the neuropil, and likely arose from the cerebellar nuclei. (3) Medium size boutons with pleomorphic vesicles: mediumsized profiles containing pleomorphic vesicles that formed symmetric synapses with proximal, intermediate and distal dendrites and, less frequently, with cell bodies. These boutons constituted 14% of the neuropil, and their morphology was similar to previous descriptions of GABAergic immunoreactive profiles in rat ventroanterior-ventrolateral nucleus, as well as boutons originating from the thalamic reticular nucleus (which is the primary source of GABAergic input to the ventroanterior-ventrolateral nucleus). The synaptic organization of thalamocortical neurons and the scarcity of interneurons in the ventroanterior—ventrolateral nucleus is discussed in relation to the critical role of the thalamic reticular nucleus in modulating the efficacy of cortical and cerebellar afferents in the ventroanterior-ventrolateral nucleus and in determining the functional state of ventroanterior-ventrolateral nucleus neurons.

Histological and ultrastructural localization of the kainate receptor subunits, KA2 and GluR6/7, in the rat nervous system using selective antipeptide antibodies.

Kainate receptors are found throughout many regions of the brain and presumably contribute to responses of neurons to glutamate and other excitatory amino acids. Two affinity-purified polyclonal antibodies that recognize the kainate binding subunits, KA2 and GluR6, were made using C-terminus peptides. A previous study demonstrated that each antibody is specific for its subunit, although antibody to GluR6 recognizes GluR7 to some extent (hence the designation GluR6/7). Vibratome sections immunostained with either antibody showed light to moderate staining in many structures in the brain as well as in cervical spinal cord, dorsal root and vestibular ganglia, and pineal and pituitary glands. Moderate levels were seen in the olfactory bulb, cerebral cortex, caudate/putamen, and hypothalamus, whereas much of the thalamus was stained lightly. In the hippocampus, CA3 pyramidal cells were stained more densely than CA1 pyramidal cells--the difference more evident with antibody to GluR6/7. In addition, neuropilar staining was densest in the stratum lucidum of the CA3 region. In the brainstem, staining was moderate to moderately dense in a number of sensory, motor, and reticular nuclei. The moderately dense staining in the reticulothalamic nucleus and pontine nuclei with antibody to GluR6/7 may represent its recognition of GluR7. In the cerebellum, staining was moderate in granular and molecular layers with antibody to KA2 and in the molecular layer with antibody to GluR6/7, whereas it was moderately dense to dense in the granular layer with the GluR6/7 antibody. Outside of the brain, densest staining was seen with antibody to KA2 in the intermediate lobe of the pituitary gland. Ultrastructural localization of immunostaining was examined in the hippocampus, cerebral cortex, and cerebellar cortex. Typically, major staining was in postsynaptic densities apposed by unstained presynaptic terminals with round or mainly round vesicles and in associated dendrites. The light microscope pattern of staining was fairly similar to that of previous [3H]kainate binding and in situ hybridization studies. In addition, comparison with previous studies on distribution of other types of glutamate receptors indicates that KA2 and GluR6/7 are found with various other subunits in many of the same cell populations throughout the nervous system.

Parvalbumin immunoreactivity in the thalamus of guinea pig: Light and electron microscopic correlation with gamma‐aminobutyric acid immunoreactivity

The relationship of the calcium binding protein parvalbumin (PV) with gamma-aminobutyric acidergic (GABAergic) neurons differs within different thalamic nuclei and animal species. In this study, the distribution of PV and GABA throughout the thalamus of the guinea pig was investigated at the light microscopic level by using immunoperoxidase methods. Intense PV labelling was found in all the GABAergic neurons of the reticular nucleus and in scattered GABAergic neurons in the anteroventral nucleus, whereas GABAergic interneurons in the ventrobasal and lateral geniculate nuclei were not PV labelled. At the electron microscopic level, preembedding immunoperoxidase for PV was combined with postembedding immunogold for GABA. In the ventrobasal nucleus, four types of profiles were recognized: 1) terminals with flattened vesicles and forming symmetric synapses, which were labelled with both PV and GABA and could therefore be identified as afferents from the reticular nucleus; 2) boutons morphologically similar to presynaptic dendrites of interneurons, labelled only with GABA; 3) large terminals with round vesicles and asymmetric synapses, labelled only with PV, which contacted GABAergic presynaptic dendrites in glomerular arrangements and resembled ascending excitatory afferents; and 4) terminals unlabelled by either antiserum. In the ventrobasal nucleus of the guinea pig a double immunocytochemical labelling permits therefore the differentiation of two populations of GABAergic vesicle-containing profiles, i.e., the terminals originating from reticular nucleus (that are double labelled) and the presynaptic dendrites originating from interneurons (that are GABA-labelled only). The possibility to differentiate GABAergic inputs from the reticular nucleus and from interneurons can shed light to the functional interpretation of synaptic circuits in thalamic sensory nuclei.

Enrichment of glutamate immunoreactivity in lemniscal terminals in the ventropostero lateral thalamic nucleus of the rat: An immunogold and WGA‐HRP study

The ventropostero lateral nucleus (VPL) is a thalamic somatosensory center receiving inputs from limbs and trunk; some of this input is via terminals of the dorsal column medial lemniscal pathway. These fibers convey non-noxious somesthesic information. In this study the neurochemical content of lemniscal afferents in VPL of rats was investigated at the electron microscopic level by combining anterograde transport of horseradish peroxidase conjugated to wheat germ agglutinin, injected in the dorsal dorsal column nuclei, with postembedding immunogold labeling for glutamate (Glu). Anterograde labeling in VPL was detected only in myelinated axons and in large terminals containing round synaptic vesicles, interpreted as lemniscal afferents. Quantitative evaluation of gold particle density showed enrichment of Glu immunolabeling in the identified lemniscal terminals with respect to other neuronal profiles. Observation of serial sections immunoreacted for Glu demonstrated consistency of labeling, whereas in alternate sections immunoreacted for Glu and for the inhibitory amino acid GABA these two antigens were always present in distinct types of terminals. These findings are in agreement with several lines of evidence, obtained with different experimental approaches, supporting the hypothesis that Glu plays a major role in conveying sensory stimuli to the thalamus from second order neurons in the dorsal column nuclei.

Muscle and Nerve Terminal Fine Structure of a Primitive Crustacean, the Cephalocarid Hutchinsoniella macracantha.

Abdominal muscles of the cephalocarid Hutchinsoniella macracantha resemble the striated muscle fibers of other crustaceans, having regularly aligned sarcomeres that average 5 μm in length; thick, wavy Z-lines; and orbits of eight thin filaments surrounding a thick filament. However, unlike most crustacean muscle fibers, the cephalocarid muscle fibers are not subdivided into myofibrils by elaboration of the longitudinally oriented sarcoplasmic reticulum. Consequently, elements of the transverse tubule and sarcoplasmic reticulum in the form of triads occur scattered over the entire fiber. Motor innervation is by means of scattered nerve terminals, populated with round synaptic vesicles, indicative of excitatory axons. By lacking myofibrils, the cephalocarid and ostracod muscle represents a much simpler condition than the myofibril-rich muscles of the other crustacean classes and signifies a primitive condition in its resemblance to the onycophoran muscle.

Fine structure of the supramammillary nucleus of the rat: analysis of the ultrastructural character of dopaminergic neurons.

The supramammillary nucleus projecting to widespread regions contains dopaminergic and non-dopaminergic neurons. The present study provided a comprehensive electron microscopic analysis of these dopaminergic and non-dopaminergic neurons in the supramammillary nucleus of the rat. The normal supramammillary nucleus was composed of round or spindle-shaped, small and medium-sized neurons (12.7 x 8.0 microns, 78.0 microns 2) containing a light oval nucleus with invaginated envelope, mitochondria, Golgi apparatus, lysosomes, less-developed rough endoplasmic reticulum, and no Nissl bodies. The majority of terminals (more than 70%) in the normal neuropil were small (diameter less than 1.0 microns) and contained round vesicles forming asymmetric synaptic contacts. The terminals often contained dense-cored vesicles. To determine the morphological features of dopaminergic neurons, we examined the ultrastructural localization of tyrosine hydroxylase (TH) immunoreactivity, which is the synthetic enzyme of dopamine, and compared TH-immunoreactive neurons to non-TH-immunoreactive neurons. Their shape and size were similar. The average number of axosomatic terminals in a sectional plane was 5.0 in TH-neurons and 2.4 in non-TH-neurons; the bouton covering ratio was 16.5% in the former and 8.6% in the latter. Both numbers were significantly larger in TH-neurons than in non-TH-neurons. Serial ultrathin sections of these neurons revealed that the average total number of axosomatic terminals was 55.7 in the TH-neuron and 28.4 in the non-TH-neuron. Characteristic lamellar bodies and sub-surface cisternae were often present in TH neurons. There were no TH-labeled terminals. These results indicate that dopaminergic neurons receive more inputs than neurons containing other neurotransmitters.

Synaptic terminal coverage of primate triceps surae motoneurons

This study examined the synaptic terminal coverage of primate triceps surae (TS) motoneurons at the electron microscopic level. In three male pigtail macaques, motoneurons were labeled by retrograde transport of cholera toxin-horseradish peroxidase that was injected into TS muscles bilaterally and visualized with tetramethylbenzidine stabilized with diaminobenzidine. Somatic, proximal dendritic, and distal dendritic synaptic terminals were classified by standard criteria and measured. Overall and type-specific synaptic terminal coverages and frequencies were determined. Labeled cells were located in caudal L5 to rostral S1 ventral horn and ranged from 40 to 74 microns in diameter (average, 54 microns). The range and unimodal distribution of diameters, the label used, and the presence of C terminals on almost all cells indicated that the 15 cell bodies and associated proximal dendrites analyzed here probably belonged to alpha-motoneurons. Synaptic terminals covered 39% of the cell body membrane, 60% of the proximal dendritic membrane, and 40% of the distal dendritic membrane. At each of these three sites, F terminals (flattened or pleomorphic vesicles, usually symmetric active zones, average contact length 1.6 microns) were most common, averaging 52%, 56%, and 58% of total coverage and 56%, 57%, and 58% of total number of cell bodies, proximal dendrites, and distal dendrites respectively. S terminals (round vesicles, usually asymmetric active zones, average contact length 1.3 microns) averaged 24%, 29%, and 33% of coverage and 33%, 35%, and 36% of number at these three sites, respectively. Thus, S terminals were slightly more prominent relative to F terminals on distal dendrites than on cell bodies. C terminals (spherical vesicles, subsynaptic cisterns associated with rough endoplasmic reticulum, average contact length 3.5 microns) constituted 24% and 11% of total terminal coverage on cell bodies and proximal dendrites, respectively, and averaged 11% and 6% of terminal number at these two locations. M terminals (spherical vesicles, postsynaptic Taxi bodies, some with presynaptic terminals, average contact length 2.7 microns) were absent on cell bodies and averaged 3% and 7% of total coverage and 2% and 5% of terminals on proximal and distal dendrites, respectively. Except for M terminals, which tended to be smaller distally, terminal contact length was not correlated with location. Total and type-specific coverages and frequencies were not correlated with cell body diameter. Primate TS motoneurons are similar to cat TS motoneurons in synaptic terminal morphology, frequency, and distribution. However, primate terminals appear to be smaller, so that the fraction of membrane covered by them is lower.

Fibroblast tissue factor: calcium and ionophore induce shape changes, release of membrane vesicles, and redistribution of tissue factor antigen in addition to increased procoagulant activity

The coagulant activity of tissue factor (TF) in the membranes of cultured cells is increased after physical disruption of the cells, and a similar increase can be elicited by treatment of cell cultures with calcium ionophore and calcium. We observed that the supernatants of cultures treated with calcium and ionophore 4-Br-A23187 contained more TF activity than those of control cultures. Phase contrast microscopy showed that cultures treated with ionophore and calcium contained rounded cells, membrane vesicles, and cell fragments. Laser-activated fluorescence microscopy of cells stained for tissue factor antigen showed that 4-Br-A23187, in the presence of 5 mmol/L calcium, caused progressive changes in cell morphology. Treatment of cultures with thrombin receptor agonist peptide caused a transient increase in cytoplasmic calcium, but had no short-term effect on TF activity or cell morphology. These combined results show that 4-Br-A23187, with extracellular calcium, increases TF activity concomitant with dramatic changes in cell morphology and the plasma membrane. The effect of increased cytoplasmic calcium on TF expression may therefore be similar in mechanism to other models of cell injury and may be caused by the effects of a sustained increase of cytosolic calcium on cellular elements that influence membrane stability and the distribution of TF per se.

Light and electron microscopic immunocytochemical localization of AMPA‐selective glutamate receptors in the rat spinal cord

alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-type glutamate receptors are probably the most widespread excitatory neurotransmitter receptors of the central nervous system, and they play a role in most normal and pathological neural activities. However, previous detailed studies of AMPA subunit distribution have been limited mainly to the brain. Thus, a comprehensive study of AMPA receptor subunit distribution was carried out on sections of rat spinal cord and dorsal root ganglia, which were immunolabeled with antibodies made against peptides corresponding to C-terminal portions of the AMPA receptor subunits: GluR1, GluR2/3, and GluR4. In the spinal cord, labeling was most prominent in the superficial dorsal horn, motoneurons, and nuclei containing preganglionic autonomic neurons. Immunostaining also was observed in neurons in other regions including those known to contain Renshaw cells and Ia inhibitory cells. Although overall immunostaining was lighter with antibody to GluR1 than with GluR2/3 and 4, there were neurons that preferentially stained with antibody to GluR1. These "GluR1 intense" neurons were usually fusiform and most concentrated in lamina X. In dorsal root ganglia, immunostaining of ganglion cell bodies was moderate to dense with antibody to GluR2/3 and light to moderate with antibody to GluR4. Possible neuroglia in the spinal cord (mainly GluR2/3 and 4) and satellite cells in dorsal root ganglia (GluR4) were immunostained. Electron microscopic studies of the superficial dorsal horn and lateral motor column showed staining that was restricted mainly to postsynaptic densities and associated dendritic and cell body cytoplasm. In dorsal horn, colocalization of dense-cored vesicles with clear, round synaptic vesicles was observed in unstained presynaptic terminals apposed to stained postsynaptic densities. Subsynaptic dense bodies (Taxi-bodies) were associated with some stained postsynaptic densities in both the superficial dorsal horn and lateral motor column. Based on several morphological features including vesicle structure and presence of Taxi-bodies, it is likely that at least some of the postsynaptic staining seen in this study is apposed to glutamatergic input from primary sensory afferent terminals.

Ischemia‐Induced delayed‐onset paraplegia is accompanied by an unusual form of synaptic degeneration in the lumbosacral segments: An experimental light and electron microscopic study in dogs

We studied the effect of high thoracic aorta cross-clamping, complete transverse section of the spinal cord at Th6 level, and combined hemisection at Th6 level followed later by high thoracic aorta cross-clamping upon the morphology and number of identified presynaptic knobs in lumbosacral segments in dogs. In animals surviving 48-72 hours after high thoracic aorta cross-clamping the occurrence of an unusual form of boutons accompanied by periboutonal halo in L3-S1 segments was found. According to the bouton size and light as well as electron microscopic appearance, four types, i.e., light giant (T1), dark enlarged (T2), light giant with periboutonal halo (T3), and giant disintegrating (T4) boutons were detected after 48 and 72 hour reperfusion. The appearance of four boutonal types in the lumbosacral segments is caused by spinal cord ischemia secondary to high thoracic aorta cross-clamping followed by 48 or 72 hour reperfusion. At the end of the sixth reperfusion day no signs of enlarged and giant boutons were detected in L3-S1 segments. A statistically significant increase of enlarged and giant boutons was noted at the end of the third reperfusion day in comparison with 48 hour survival. After spinal cord transection at midthoracic (Th6) level, followed by 72 hour survival, no such unusual synaptic knobs could be found in L3-S1 segments. The laminar distribution pattern of T1-T4 types based on light microscopic analysis and confirmed electron microscopically is characteristic and strictly bound to those spinal cord gray matter layers which serve as main termination sites of the descending cortical, brain stem, as well as long propriospinal projections in the lumbosacral segments (laminae V-VII). A statistically significant increase of enlarged and giant boutons was found in the intermediate zone (lamina VII). Hemisection at midthoracic level (Th6) followed later by 30 minute high thoracic aorta cross-clamping and 48 hour reperfusion caused a marked decrease of enlarged and giant boutons in L3-S1 segments on the hemisectioned side in comparison with the contralateral one. Large amounts of irregularly arranged round vesicles and tubular profiles were disclosed in the boutonal matrix of T1, T3, and T4 types in L3-S1 segments of animals subjected to 30 minute high thoracic aorta cross-clamping followed by 72 hour reperfusion. Accumulation of tubular and membranous materials was invariably seen in the bulbous enlargement of the terminal axonal branch.(ABSTRACT TRUNCATED AT 400 WORDS)

Synaptic innervation of neurons in the internal pallidal segment by the subthalamic nucleus and the external pallidum in monkeys

In order to better understand the way by which the subthalamic nucleus interacts with the globus pallidus to control the output of the basal ganglia, we carried out a series of experiments to investigate the pattern of synaptic innervation of the pallidal neurones by the subthalamic terminals in the squirrel monkey. To address this problem we used the anterograde transport of biocytin. Following injections of biocytin in the subthalamic nucleus, rich plexuses of labelled fibres and varicosities formed bands that lay along the medullary lamina in both segments of the ipsilateral pallidum. At the electron microscopic level, two populations of biocytin-containing terminals were identified in the internal pallidum (GPi). A first group of small to medium-sized terminals (type 1; mean cross-sectional area +/- S.D. = 0.41 +/- 0.04 microns 2) contained round vesicles and formed asymmetric synapses with dendritic shafts (95%) of mixed sizes (maximum diameter ranging from 0.3 to 4.0 microns) and spine-like structures (5%). The second group of terminals (type 2) contained pleiomorphic vesicles, had a larger cross-sectional area (mean +/- S.D. = 0.9 +/- 0.4 micron 2) and formed symmetric synapses predominantly with perikarya (41%) and large dendrites (57%). In some cases, the two types of terminals converged at the level of single GPi neurones. Postembedding immunogold method revealed that the type 2 terminals displayed gamma-aminobutyric acid (GABA) immunoreactivity, whereas the type 1 terminals did not. In the external pallidum (GPe), injections in the subthalamic nucleus labelled both type 1 or type 2 terminals. However, the labelled type 2 boutons were much less abundant in GPe than in GPi. The presence of biocytin-labelled perikarya in GPe and the fact that the type 2 terminals displayed GABA immunoreactivity led us to suspect that these terminals were derived from axons of GPe neurones. In agreement with this hypothesis, injections of Phaseolus vulgaris-leucoagglutinin (PHA-L) in GPe labelled terminals in GPi that displayed the morphological features and a pattern of synaptic organization similar to the type 2 terminals. In conclusion, the results of our study demonstrate that the subthalamopallidal terminals form asymmetric synapses that are distributed along the dendritic tree of GPe and GPi neurones. In contrast, the GPe projection to GPi gives rise to large GABA-containing terminals that form symmetric synapses predominantly with the proximal region of pallidal neurones.(ABSTRACT TRUNCATED AT 400 WORDS)

Ultrastuctural studies of the primate lateral geniculate nucleus: Morphology and spatial relationships of axon terminals arising from the retina, visual cortex (area 17), superior colliculus, parabigminal nucleus, and pretectum of Galago crassicaudatus

The electron microscopic autoradiographic tracing method has been used to examine the morphology and postsynaptic relationships of five projections (retina, cortical area 17, superior colliculus (tectal), parabigeminal nucleus, and pretectum) to the dorsal lateral geniculate nucleus of the greater bush baby Galago crassicaudatus. Retinal terminals have been examined in the contralaterally innervated layer of each of the three matched pairs [parvi- (X-cell), magno- (Y-cell), and koniocellular (small, W-cell)] of geniculate layers. These terminals are large and contain pale mitochondria and round vesicles (RLPs). RLPs are presynaptic to juxtasomatic regions of parvi- and magnocellular neurons. In contrast, RLPs innervate more distal regions of koniocellular neurons. Labeled cortical, tectal, and parabigeminal terminals are relatively small and contain round vesicles and dark mitochondria. Cortical terminals in each of the three representative layers are presynaptic to small diameter dendrites. No convergence of cortical and retinal terminals has been seen in any layer. Labeled tectal and parabigeminal terminals are found primarily in the koniocellular layers, but the latter are also seen in all other layers. Tectal and parabigeminal terminals have been observed converging with retinal terminals on dendrites of some koniocellular neurons. Labeled pretectogeniculate terminals contain densely packed pleomorphic vesicles, dark mitochondria, and a dark cytoplasmic matrix. These terminals, which are present in each of the representative layers, are presynaptic to conventional dendrites and profiles containing loosely dispersed pleomorphic vesicles and a pale cytoplasmic matrix.

Gross and microscopic anatomy of the canine intrinsic cardiac nervous system

A three-dimensional description of the distribution and organization of the canine intrinsic cardiac nervous system was developed in order to characterize its full extent physiologically. The anatomy of the canine intrinsic cardiac nervous system was investigated in 67 mongrel dogs by means of visualization following methylene blue staining as well as by light and electron microscopic analyses. Collections of ganglia associated with nerves, i.e., ganglionated plexuses, were identified in specific locations in epicardial fat and cardiac tissue. Distinct epicardial ganglionated plexuses were consistently observed in four atrial and three ventricular regions, with occasional neurons being located throughout atrial and ventricular tissues. One ganglionated plexus extended from the ventral to dorsal surfaces of the right atrium. Another ganglionated plexus, with three components, was identified in fat on the left atrial ventral surface. A ganglionated plexus was located on the mid-dorsal surface of the two atria, extending ventrally in the interatrial septum. A fourth atrial ganglionated plexus was located at the origin of the inferior vena cava extending to the dorsal caudal surface of the two atria. On the cranial surface of the ventricles a ganglionated plexus that surrounded the aortic root was identified. This plexus extended to the right and left main coronary arteries and origins of the ventral descending and circumflex coronary arteries. Two other ventricular ganglionated plexuses were identified adjacent to the origins of the right and left marginal coronary arteries. Intrinsic cardiac ganglia ranged in size from ones comprising one or a few neurons along the course of a nerve to ones as large as 1 x 3 mm estimated to contain a few hundred neurons. Intrinsic cardiac neuronal somata varied in size and shape, up to 36% containing multiple nucleoli. Electron microscopic examination demonstrated typical autonomic neurons and satellite cells in intrinsic cardiac ganglia. Many of their axon profiles contained large numbers of clear, round, and dense-core vesicles. Asymmetrical axodendritic synapses were common. The canine intrinsic cardiac nervous system contains a variety of neurons interconnected via plexuses of nerves, the distribution of which is wider than previously assumed.

Extraordinary synapses of the unipolar brush cell: An electron microscopic study in the rat cerebellum

A neglected type of neuron, termed the unipolar brush cell, was recently characterized in the granular layer of the mammalian cerebellar cortex with several procedures, including light and electron microscopic immunocytochemistry utilizing antibodies to calretinin and neurofilament proteins. Although certain features of the unipolar brush cells were highlighted in these studies, the internal fine structure was partially obfuscated by immunoreaction product. In this study, rat cerebella were prepared for electron microscopy after perfusion fixation and Araldite embedding, and folia of the vestibulo-cerebellum, where unipolar brush cells are known to be enriched, were studied by light microscopy in semithin (0.5-1 micron) sections and by electron microscopy in ultrathin sections. Unipolar brush cells were easily identified in semithin sections immunostained with antibodies to GABA and/or glycine, and counterstained with toluidine blue. The unipolar brush cells have a pale cytoplasm and are GABA and glycine negative, while Golgi cells are darker and appear positive for GABA and, for the most part, also for glycine. Sets of identification criteria to differentiate unipolar brush cells from granule and Golgi cells in standard electron micrographs are presented. The unipolar brush cells possess many distinctive features that make them easily distinguishable from other cerebellar neurons and form unusually conspicuous and elaborate synapses with mossy rosettes. The unipolar brush cell has a deeply indented nucleus containing little condensed chromatin. The Golgi apparatus is large and the cytoplasm is rich in neurofilaments, microtubules, mitochondria, and large dense core vesicles, but contains few cisterns of granular endoplasmic reticulum. In addition, unipolar brush cells contain an unusual inclusion, which invariably lacks a limiting membrane and is made up of peculiar ringlet subunits. The cell body usually emits a thin axon and is provided with a single, large dendritic trunk that terminates with a paintbrush-like bunch of branchlets. Numerous nonsynaptic appendages emanate from the cell body, the dendritic stem, and the branchlets. The appendages contain rare organelles and lack neurofilaments. The branchlets contain numerous mitochondria, neurofilaments, large dense core vesicles, and clusters of clear, small, and round synaptic vesicles. They form extensive asymmetric synaptic junctions with one or two mossy fibers, which indicates minimal convergence of excitatory inputs. Under the postsynaptic densities, the branchlet cytoplasm displays a microfilamentous web. Besides their contact with mossy rosettes, the branchlets form symmetric and asymmetric synaptic junctions with presumed Golgi cell boutons that contain pleomorphic synaptic vesicles, indicating that the unipolar brush cells receive an inhibitory modulation. Some of these junctions are unusually extensive.(ABSTRACT TRUNCATED AT 400 WORDS)