Ultrastructural relationships between choline acetyltransferase- and neuropeptide y-containing neurons in the rat striatum.

The relationships between cholinergic and neuropeptide Y-containing neuronal systems in the rat striatum were examined using a dual immunoperoxidase labelling method. These neurons were identified by their immunoreactivity to choline acetyltransferase and neuropeptide Y, respectively, and were visualized on the same sections using 3,3'-diaminobenzidine and benzidine dihydrochloride as distinct chromogens under two conditions: (i) neuropeptide Y detection by the 3,3'-diaminobenzidine diffuse brown reaction product and choline acetyltransferase detection by the benzidine dihydrochloride blue, granular reaction product; (ii) choline acetyltransferase detection by 3,3'-diaminobenzidine and neuropeptide Y detection by benzidine dihydrochloride. Although both neuropeptide Y- and choline acetyltransferase-immunoreactive cell bodies were simultaneously detected and were easily distinguishable whatever the conditions used, neuropeptide Y- and choline acetyltransferase-immunoreactive dendrites and axons could not be visualized on the same sections, since only the diaminobenzidine-labelled processes were detectable. Light microscopic observations on sections dual labelled with either method confirmed that choline acetyltransferase and neuropeptide Y immunoreactivities were localized in morphologically different populations of striatal neurons scattered throughout the striatum, choline acetyltransferase immunoreactivity being associated with large neurons and neuropeptide Y immunoreactivity with medium-sized neurons. In addition, the choline acetyltransferase-immunoreactive neurons were found to be more numerous than the neuropeptide Y-immunoreactive neurons and to be prevalent in the dorsolateral areas of the striatum, whereas neuropeptide Y-immunoreactive neurons were preferentially found in the ventromedial areas of this structure. Electron microscopic observations on sections processed under either condition revealed that choline acetyltransferase-positive terminals form synaptic contacts of the symmetrical type with neuropeptide Y-positive somata and proximal dendrites and that choline acetyltransferase-positive neurons are contacted by neuropeptide Y-positive terminals. These data show that the striatal neuropeptide Y- and choline acetyltransferase-containing neuronal systems have reciprocal synaptic interactions and provide morphological support for the hypothesis that striatal cholinergic and neuropeptide Y interneuron activities may be functionally linked.     

Plasmalemmal appositions between cholinergic and non-cholinergic neurons in rat caudate-putamen nuclei

We have observed that in rat caudate-putamen nuclei, neurons immunolabeled for choline acetyltransferase were sometimes in direct apposition to unlabeled perikarya and dendrites [Pickel V. M. and Chan J. (1990) J. Neurosci. Res. 25, 263-280]. Similar juxtapositions between plasmalemmas of nerve cells each receiving input from one common terminal have been associated with activation of certain central neurons [Theodosis D. T. and Poulain D. A. (1989) Brain Res. 484, 361-366]. Thus, we sought to determine the relative abundance and ultrastructure of the appositions and the frequencies of shared synapses between choline acetyltransferase-labeled and unlabeled neurons in the rat striatum. A monoclonal antibody raised against choline acetyltransferase was localized in semi-adjacent ultrathin sections through 24 neurons in the dorsolateral caudate-putamen nuclei. Five of these choline acetyltransferase-labeled perikarya showed direct somatic appositions with unlabeled neurons. The remaining 19 of the choline acetyltransferase-labeled perikarya did not show somatic appositions with unlabeled perikarya; however, when traced through multiple (20-100) semi-adjacent sections their dendrites always showed extensive plasmalemmal juxtapositions with one or more unlabeled perikarya. The apposed perikarya had round nuclei and other characteristics of medium, spiny neurons. The majority of the apposed cholinergic and non-cholinergic neurons were postsynaptic to at least one common unlabeled terminal. These terminals usually formed symmetric junctions. At sites of appositions, the plasmalemmas of choline acetyltransferase-immunoreactive soma or dendrites and unlabeled neurons were closely spaced without intervening astrocytic processes. The appositions lacked the ultrastructural features typical of gap-junctions, but did occasionally show parallel arrays of thin (1-2 nm) electron-dense bands. In both labeled and unlabeled perikarya, the nuclei were separated from the appositional zones by narrow (0.7-3.3 microns) rims of cytoplasm. This cytoplasmic rim contained subsurface cisternae and other less specialized smooth and rough endoplasmic reticulum, and vesicular structures. The findings suggest that in the caudate-putamen nuclei (1) the tonically active cholinergic neurons [Wilson C. J. et al. (1990) J. Neurosci. 10, 508-519] may modulate or be modulated by non-cholinergic spiny neurons through non-synaptic somatic or dendritic appositions, and (2) that both neurons may be simultaneously inhibited by shared afferent input. Activation of this system could facilitate coordinated movements through synchronization of cholinergic interneurons and spiny projection neurons containing GABA or other transmitters.     

Descriptive morphology of developing fetal neostriatal allografts in the rhesus monkey: a correlated light and electron microscopic Golgi study.

Primate fetal striatal neurons were transplanted into the ibotenic acid lesioned rhesus monkey striatum. Ten weeks after transplantation the monkeys were transcardially perfused and graft tissue was histologically stained. Golgi impregnated, and processed for electron microscopy. The monkeys received magnetic resonance imaging (MRI) scans before lesioning, after lesioning, and ten weeks after transplantation to noninvasively study the striatal grafts. The study demonstrated that fetal striatal grafts, measuring up to 0.4 x 0.8 cm, can survive for extended periods of time in the non-human primate. Hematoxylin-eosin stained sections of the transplant demonstrated that neuronal, glial, vascular, and lymphocytic cells were present in the graft. The majority of the neurons had somatic diameters between 8 and 20 microns and were characterized by nuclei containing multiple nucleoli. A few neurons within the graft had somatic diameters up to 40 microns. These larger neurons exhibited more mature cytoplasm containing a moderate amount of Nissl substance. Some of the blood vessels within the graft were surrounded by a large number of plasma cells, but there was no evidence of hemorrhage or necrosis. Bielschowsky staining and Golgi impregnation of the transplanted tissue demonstrated that there were neurons at various degrees of differentiation. Some of the neurons had varicose dendrites, growth cones, and filopodia, which are all characteristics of immature neurons, while others had a much more mature appearance, including a moderate number of dendritic spines. Some of these neurons had an appearance typical of differentiating "medium spiny" neurons of the normal striatum. Electron microscopic analysis of the transplanted tissue and individual Golgi-impregnated neurons within the transplant confirmed that there were developing neurons within the graft. These neurons had an increased nuclear-to-cytoplasmic ratio and had nuclei containing multiple nucleoli. The neuropil surrounding these neurons was loosely organized and contained large areas of extracellular space. The neuropil exhibited developing dendrites, numerous growth cones, and mature synapses. In summary, the study demonstrated that fetal striatal allografts can survive for up to three months in the rhesus monkey and undergo normal differentiation as assessed by Golgi impregnation and electron microscopy.     

Input from the frontal cortex and the parafascicular nucleus to cholinergic interneurons in the dorsal striatum of the rat.

Evidence derived from many experimental approaches indicates that cholinergic neurons in the dorsal striatum (caudate-putamen) are responsive to excitatory amino acids. Furthermore, evidence from physiological experiments indicate that the excitatory input is derived from the cortex and/or the thalamus. The object of the present experiment was to anatomically test whether cholinergic neurons receive cortical and/or thalamic input in the dorsal striatum using a combined anteograde tracing and immunocytochemical approach at both the light- and electron-microscopic levels. Rats received injections of the anterograde tracers Phaseolus vulgaris-leucoagglutinin or biocytin at multiple sites in the frontal cortex or parafascicular nucleus of the thalamus. Sections of the striatum were stained to reveal the anterogradely transported markers and then immunostained to reveal choline acetyltransferase immunoreactivity. The striata of these animals contained dense networks of anterogradely labelled fibres that were dispersed throughout the neuropil and interspersed with the choline acetyltransferase-immunoreactive (i.e. cholinergic) perikarya and dendrites. The anterogradely labelled fibres were often closely apposed to the choline acetyltransferase-immunoreactive neurons. Examination of electron-microscopic sections failed to demonstrate cortical terminals in synaptic contact with the cholinergic neurons even when choline acetyltransferase-immunoreactive structures were examined that had first been identified in the light microscope as having cortical terminals closely apposed to them. In these cases it was often observed that the cortical terminal, although apposed to the membrane of the labelled neurone, made synaptic contact with an unlabelled spine that was in the vicinity. In contrast to the cortical input, analysis of material that was double-stained to reveal thalamostriatal terminals and choline acetyltransferase-immunoreactive structures, revealed that the thalamostriatal terminals were often in asymmetrical synaptic contact with the perikarya and dendrites of cholinergic neurons. It is concluded that the cholinergic neurons of the dorsal striatum, like those of the ventral striatum or nucleus accumbens [Meredith and Wouterlood (1990) J. comp. Neurol. 296, 204-221] receive very little or no input from the cortex but are under a prominent synaptic control by the thalamostriatal system. Those pharmacological effects of excitatory amino acids on the cholinergic systems of the striatum are therefore presumably related to the thalamostriatal and not the corticostriatal system.     

Substance P immunoreactivity in the rat interpeduncular nucleus: synaptic interactions between substance P-positive profiles and choline acetyltransferase- or glutamate decarboxylase-immunoreactive structures.

The subnuclear and synaptic distribution of substance P immunoreactivity was examined in the rat interpeduncular nucleus at the light and electron microscope level. The nucleus possessed a prominent substance P-immunoreactive axonal plexus in the lateral and dorsomedial subnuclei, and in the dorsal cap of the rostral subnucleus. The density of substance P-immunoreactive axons in the remaining subnuclear divisions was sparse to moderate. Terminals of immunoreactive axons contained spherical vesicles and formed asymmetric contacts on dendritic processes exclusively. Immunoreactive neurons, restricted to the rostral subnucleus, possessed long, sparsely branched dendrites. Unlabelled terminals containing either spherical or pleomorphic vesicles contacted substance P-immunoreactive dendritic profiles. Axodendritic and axosomatic synapses containing substance P immunoreactivity pre- and postsynaptically were not observed. Ultrastructural evidence for synaptic relationships between substance P-containing profiles and those containing either choline acetyltransferase or glutamate decarboxylase was obtained by means of double antigen immunohistochemistry. Terminals of fasciculus retroflexus axons stained for choline acetyltransferase immunoreactivity formed asymmetric synaptic contacts with substance P-immunoreactive dendritic profiles. Few substance P-positive dendrites in the rostral subnucleus received terminals possessing glutamate decarboxylase activity. Unlabelled terminals containing either spherical or pleomorphic vesicles contacted substance P- and glutamate decarboxylase-immunoreactive dendritic profiles simultaneously. Terminals possessing either substance P or glutamate decarboxylase immunoreactivity formed synaptic contacts with dendritic processes of neurons in the lateral subnucleus. Many of the neurons within this subnuclear division contained glutamate decarboxylase. This study provides direct evidence of synaptic relationships between choline acetyltransferase-immunoreactive axons and substance P-immunoreactive dendritic profiles, and between substance P-positive axons and glutamate decarboxylase-immunoreactive dendrites. These findings reveal that two types of transmitter-specific axons of the fasciculus retroflexus innervate neuronal populations of the interpeduncular nucleus stained immunohistochemically for either substance P or glutamate decarboxylase.     

The cholinergic innervation of normal and transplanted superior colliculus in the rat: an immunohistochemical study

The distribution of choline acetyltransferase was determined in normal and transplanted rat superior colliculus with choline acetyltransferase immunohistochemistry. This distribution was compared to the pattern of histochemically detected acetylcholinesterase activity. To determine cholinergic input to the superficial superior colliculus, double labelling experiments combining retrograde tracing methods and choline acetyltransferase immunohistochemistry were carried out. No choline acetyltransferase-containing neurons were observed in the rat superior colliculus. A dense network of choline acetyltransferase-immunoreactive fibres and terminals was seen in the intermediate layers of the normal superior colliculus. The distribution was patchy and very similar to the pattern of acetylcholinesterase activity. Occasional fibres and terminals were seen in the deep tectal laminae. The superficial layers contained a low number of choline acetyltransferase-stained fibres and terminals but a comparatively high level of acetylcholinesterase activity. Following a unilateral injection of a tracer into the superficial superior colliculus, retrogradely labelled choline acetyltransferase-immunoreactive neurons were found in the dorsal and ventral subnuclei of the ipsilateral parabigeminal nucleus. As in the normal superior colliculus, choline acetyltransferase-positive neurons were not found in tectal transplants. However, choline acetyltransferase-immunoreactive fibres and terminals were seen in grafts but only in those which had extensive connections with the host midbrain. The pattern of staining most closely resembled that seen in the intermediate layers of the normal superior colliculus. The similar arrangement of choline acetyltransferase and acetylcholinesterase activity in the intermediate layers of normal rat superior colliculus provides further evidence for cholinergic innervation to these layers, probably originating in the dorsal and pedunculopontine tegmental nuclei. The data from the double labelling experiments indicate that the choline acetyltransferase-immunoreactive terminals observed in the superficial layers represent the terminal field of an ipsilateral cholinergic projection from the parabigeminal nucleus. Tectal grafts receive cholinergic innervation from the host. The evidence suggests that much of this input derives from the cholinergic nuclei in the brainstem tegmentum which normally project to the intermediate tectal layers.     

Nerve cell clusters in dorsal striatum and nucleus accumbens of the male rat demonstrated by glucocorticoid receptor immunoreactivity

Glucocorticoid receptor-immunoreactive nerve cells have been analysed in the dorsal striatum and nucleus accumbens of the rat by means of a monoclonal antibody against rat liver glucocorticoid receptor. Glucocorticoid receptor immunoreactivity was present in the nuclei of the vast majority of the striatal nerve cells. The analysis of sections stained with glucocorticoid receptor antibody and cresyl violet showed that around 90% of the entire striatal neuronal population contained glucocorticoid receptor immunoreactivity. By means of the double immunoperoxidase technique evidence was provided that somatostatin- and choline acetyltransferase-immunoreactive nerve cells in the striatum do not contain glucocorticoid receptor immunoreactivity. The density of glucocorticoid receptor-immunoreactive nerve cells in the grey matter and the presence of clusters of glucocorticoid receptor-immunoreactive nerve cells have been investigated in three fields located in the medial and central dorsal striatum and nucleus accumbens at the coronal level A 8620 microns according to the K?nig and Klippel atlas using computer-assisted image analysis. Every aggregate containing three or more glucocorticoid receptor-immunoreactive nerve cells, which had an intercenter distance less than the mean diameter (10-11 microns) of the striatal cells, was considered an island. A higher density of both glucocorticoid receptor-immunoreactive nerve cell nuclei and islands was found in the nucleus accumbens with respect to dorsal striatal areas. The most frequent island formed consisted of three to ten nerve cells both in dorsal striatum and nucleus accumbens. Furthermore, some nucleus accumbens islands contained up to 100 nerve cells, whereas in the dorsal striatum the maximum number of glucocorticoid receptor-immunoreactive nerve cells per island ranged from 50 to 60. The present procedure proved to be a sensitive method to reveal clusters of chemically identified structures and provided evidence for a basic cytoarchitectonic organization of the dorsal striatum and nucleus accumbens of the rat. This paper also demonstrated that the vast majority, but not all, striatal nerve cells contained glucocorticoid receptor immunoreactivity, and thus may be under the control of circulating glucocorticoids. In fact, only small transmitter-identified neuronal populations, such as somatostatin- and choline acetyltransferase-immunoreactive nerve cells, were devoid of glucocorticoid receptor immunoreactivity.     

Cholinergic input to dopaminergic neurons in the substantia nigra: a double immunocytochemical study.

In order to determine whether the cholinergic fibres that innervate the substantia nigra make synaptic contact with dopaminergic neurons of the substantia nigra pars compacta, a double immunocytochemical study was carried out in the rat and ferret. Sections of perfusion-fixed mesencephalon were incubated first to reveal choline acetyltransferase immunoreactivity to label the cholinergic terminals and then tyrosine hydroxylase immunoreactivity to label the dopaminergic neurons. Each antigen was localized using peroxidase reactions but with different chromogens. At the light microscopic level, in confirmation of previous observations, choline acetyltransferase-immunoreactive axons and axonal boutons were found throughout the substantia nigra. The highest density of these axons was found in the pars compacta where they were often seen in close apposition to tyrosine hydroxylase-immunoreactive cell bodies and dendrites. In the ferret where the choline acetyltransferase immunostaining was particularly strong, bundles of immunoreactive fibres were seen to run through the reticulata perpendicular to the pars compacta. These bundles were associated with tyrosine hydroxylase-immunoreactive dendrites that descended into the reticulata. The choline acetyltransferase-immunoreactive fibres made "climbing fibre"-type multiple contacts with the tyrosine hydroxylase positive dendrites. At the electron microscopic level the choline acetyltransferase-immunoreactive axons were seen to give rise to vesicle-filled boutons that formed asymmetrical synaptic specializations with nigral dendrites and perikarya. The synapses were often associated with sub-junctional dense bodies. On many occasions the postsynaptic structures contained the tyrosine hydroxylase immunoreaction product, thus identifying them as dopaminergic. It is concluded that at least one of the synaptic targets of cholinergic terminals in the substantia nigra are the dendrites and perikarya of dopaminergic neurons and that in the ferret at least, the dendrites of dopaminergic neurons that descend into the pars reticulata receive multiple synaptic inputs from individual cholinergic axons.     

Immunocytochemical studies of GABAergic neurons in rat basal ganglia and their relations to other neuronal systems

GABAergic neurons were localized in the rat basal ganglia by glutamate decarboxylase (GAD) immunohistochemistry. In the striatum (caudato-putamen, accumbens nucleus) a medium density of GAD-positive terminals was observed; a small number of medium-to-large size neurons and the vast majority of medium-size neurons were GAD immunoreactive. In addition, opioid peptide-like immunoreactivity was colocalized in a subclass of GAD-positive medium-size striatal neurons. The pallido-nigral system (GP, VP, EP, SNR) displayed a high density of GAD-positive axon terminals which synapsed upon dendrites and nerve cell bodies. The majority of pallido-nigral neurons also were GAD-immunoreactive. In contrast, the substantia nigra pars compacta and the subthalamic nucleus contained only few GAD-immunoreactive neurons.     

Direct synaptic projections to the myenteric ganglion of the rat subdiaphragmatic esophagus from the dorsal motor nucleus of the vagus

We have examined the ultrastructure of the myenteric ganglion of the subdiaphragmatic esophagus and determined whether the ganglion neurons receive direct projections from the dorsal motor nucleus of the vagus (DMV) using wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) as an anterograde tracer. The neurons (22.2 microm x 13.3 microm) of myenteric ganglion in the esophagus contained dark cytoplasm having many free ribosomes, mitochondria, and an oval nucleus, and received only a few axon terminals contacting somata. All axon terminals formed asymmetric synaptic contacts with dendrites or somata. Approximately 85% of the axon terminals contacting dendrites and about 50% of the axon terminals contacting somata contained pleomorphic vesicles, while the rest contained round synaptic vesicles. When WGA-HRP was injected into the DMV, anterogradely labeled fibers and terminals were found in the myenteric ganglia. The WGA-HRP labeled terminals were large (1.97 microm) and contained round clear vesicles and small granular vesicles. These labeled terminals contacted exclusively the small dendrites, but not the somata. These results suggest that the DMV neurons project directly to the myenteric ganglion neurons and regulate the esophageal muscles via the ganglion neurons.     

Nitrergic neurons make synapses on dual-input dendritic spines of neurons in the cerebral cortex and the striatum of the rat: implication for a postsynaptic action of nitric oxide

Pre-embedding electron microscopic immunocytochemistry was used to examine the ultrastructure of neurons containing nitric oxide synthase and to evaluate their synaptic relationships with target neurons in the striatum and sensorimotor cerebral cortex. Intense nitric oxide synthase immunoreactivity was found by light and electron microscopy in a type of aspiny neuron scattered in these two regions. The intensity of the labeling was uniform in the soma, dendrites and axon terminals of these neurons. In both forebrain regions, nitric oxide synthase-immunoreactive neurons received synaptic contacts from unlabeled terminals, which were mostly apposed to small-caliber dendrites. The unlabeled symmetric contacts were generally about four times as abundant as the unlabeled asymmetric contacts on the nitric oxide synthase-immunoreactive neurons. Terminals labeled for nitric oxide synthase were filled with synaptic vesicles and were observed to contact unlabeled neurons. Only 54% (in the cerebral cortex) and 44.3% (in the striatum) of the nitric oxide synthase-immunoreactive terminals making apposition with the target structures were observed to form synaptic membrane specializations within the plane of the randomly sampled sections. The most common targets of nitric oxide synthase-immunoreactive terminals were thin dendritic shafts (54% of the immunoreactive terminals in the cortex and 75.7% of the immunoreactive terminals in the striatum), while dendritic spines were a common secondary target (42% of the immunoreactive terminals in the cortex and 20.6% of the immunoreactive terminals in the striatum). The spines contacted by nitric oxide synthase-immunoreactive terminals typically also received an asymmetric synaptic contact from an unlabeled axon terminal.These findings suggest that: (i) nitric oxide synthase-immunoreactive neurons in the cortex and striatum preponderantly receive inhibitory input; (ii) nitric oxide synthase-containing terminals commonly make synaptic contact with target structures in the cortex and striatum; (iii) spines targeted by nitric oxide synthase-containing terminals in the cortex and striatum commonly receive an asymmetric contact as well, which may provide a basis for a synaptic interaction of nitric oxide with excitatory input to individual spines.     

Ultrastructural identification of cholinergic neurons in the external cuneate nucleus of the gerbil: acetylcholinesterase histochemistry and choline acetyltransferase immunocytochemistry

Summary Using acetylcholinesterase histochemical and choline acetyltransferase immunocytochemical localization methods, this study has provided conclusive evidence for the existence of cholinergic neurons in the external cuneate nucleus of gerbils. By light microscopy, both acetylcholinesterase and choline acetyltransferase labelling was confined to the rostral portion of the external cuneate nucleus. Ultrastructurally, acetylcholinesterase reaction products were found in the nuclear envelope, cisternae of rough endoplasmic reticulum and Golgi saccules of some somata and large dendrites as well as in the membranes of small dendrites, myelinated axons and axon terminals. These neuronal elements were also stained for choline acetyltransferase; immunoreactivity was associated with nuclear pores, nuclear envelope, perikaryal membrane and all the membranous structures within the cytoplasm. Of the total choline acetyltransferase-labelled neuronal profiles analysed, 79% were myelinated axons, 15% dendrites, 4% somata and 2% axon terminals. The immunostained axon terminals consisted of two types containing either round (Rd type; 62.5%) or pleomorphic (Pd type; 37.5%) vesicles. Both were associated directly with choline acetyltransferase-positive dendrites. In contrast to the paucity of choline acetyltransferase-labelled axon terminals, numerous choline acetyltransferase-positive myelinated axons were present. It may thus be hypothesized that most, if not all, of the external cuneate nucleus cholinergic neurons are projection cells; such cells may give rise to axonal collaterals which synapse onto their own dendrites for possible feedback control. Choline acetyltransferase-positive dendrites were contacted by numerous unlabelled presynaptic boutons, 60% of which contained round or spherical synaptic vesicles (Rd boutons) and 40% flattened vesicles (Fd boutons), suggesting that these neurons are under strong inhibitory control. The preferential concentration of cholinergic components in the rostral external cuneate nucleus may be significant in the light of the highly organized somatotopy in the external cuneate nucleus and its extensive efferent projections to medullary autonomic-related nuclei. Our results suggest that the cholinergic neurons may be involved in somatoautonomic integration.     

Distribution of DARPP-32 in the basal ganglia: an electron microscopic study

Summary DARPP-32, a dopamine and cyclic AMP-regulated phosphoprotein, has been studied by light and electron microscopical immunocytochemistry in the rat caudatoputamen, globus pallidus and substantia nigra. In the caudatoputamen, DARPP-32 was present in neurons of the medium-sized spiny type. Immunoreactivity for DARPP-32 was present in dendritic spines, dendrites, perikaryal cytoplasm, most but not all nuclei, axons and a small number of axon terminals. Immunoreactive axon terminals in the caudatoputamen formed symmetrical synapses with immunolabelled dendritic shafts or somata. Neurons having indented nuclei were never immunoreactive. In the globus pallidus and substantia nigra pars reticulata, DARPP-32 was present in myelinated and unmyelinated axons and in axon terminals. The labelled axon terminals in these regions formed symmetrical synaptic contacts on unlabelled dendritic shafts or on unlabelled somata. These data suggest that DARPP-32 is present in striatal neurons of the medium-sized spiny type and that these DARPP-32-immunoreactive neurons form symmetrical synapses on target neurons in the globus pallidus and substantia nigra. The presence of DARPP-32 in these striatal neurons and in their axon terminals suggests that DARPP-32 mediates part of the response of medium-size spiny neurons in the striaturn to dopamine D-l receptor activation.     

Fine structural survey of the intermediate subnucleus of the nucleus tractus solitarii and its glossopharyngeal afferent terminals

The intermediate subnucleus of the nucleus tractus solitarii (imNTS) receives somatosensory inputs from the soft palate and pharynx, and projects onto the nucleus ambiguus, thus serving as a relay nucleus for swallowing. The ultrastructure and synaptology of the rat imNTS, and its glossopharyngeal afferent terminals, have been examined with cholera toxin-conjugated horseradish peroxidase (CT-HRP) as an anterograde tracer. The imNTS contained oval or ellipsoid-shaped, small to medium-sized neurons (18.2×11.4 μm) with little cytoplasm, few cell organelles and an irregularly shaped nucleus. The cytoplasm often contained one or two nucleolus-like stigmoid bodies. The average number of axosomatic terminals was 1.8 per profile. About 83% of them contained round vesicles and formed asymmetric synaptic contacts (Gray’s type I), while about 17% contained pleomorphic vesicles and formed symmetric synaptic contacts (Gray’s type II). The neuropil contained small or large axodendritic terminals, and about 92% of them were Gray’s type I. When CT-HRP was injected into the nodose ganglion, many labeled terminals were found in the imNTS. All anterogradely labeled terminals contacted dendrites but not somata. The labeled terminals were usually large (2.69±0.09 μm) and exclusively of Gray’s type I. They often contacted more than two dendrites, were covered with glial processes, and formed synaptic glomeruli. A small unlabeled terminal occasionally made an asymmetric synaptic contact with a large labeled terminal. The large glossopharyngeal afferent terminals and the neurons containing stigmoid bodies characterized the imNTS neurons that received pharyngeal afferents.     

Ultrastructural evidence that substance P neurons form synapses with noradrenergic neurons in the A7 catecholamine cell group that modulate nociception.

Potent antinociception can be produced by electrical stimulation of spinally projecting noradrenergic neurons in the A7 catecholamine cell group and this effect is blocked by intrathecal injection of alpha2-adrenoceptor antagonists. Microinjection of substance P near A7 neurons also produces antinociception that is blocked by intrathecal injection of alpha2-adrenoceptor antagonists. These observations suggest that substance P produces antinociception by activating noradrenergic A7 neurons. However, it is not known whether this effect of substance P is produced by a direct or an indirect action on A7 neurons. Although light microscopic studies have demonstrated the existence of both substance P-containing axon terminals and neurokinin-1 receptors in the region of the A7 cell group, it is not known whether substance P terminals form synapses with noradrenergic A7 neurons. These experiments used double-labeling immunocytochemical methods and electron microscopic analysis to determine whether substance P-containing axons form synapses with noradrenergic neurons in the A7 cell group. Pre-embedding immunocytochemistry, combined with light and electron microscopic analysis, was used to provide ultrastructural evidence for synaptic connections between substance P-immunoreactive terminals labeled with immunoperoxidase and tyrosine hydroxylase-immunoreactive A7 neurons labeled with silver-enhanced immunogold. Tyrosine hydroxylase labeling was found in perikarya and dendrites in the A7 region, and substance P labeling was found in axons and synaptic terminals. Substance P-labeled terminals formed asymmetric synapses with tyrosine hydroxylase-labeled dendrites, but only a few of these were present on tyrosine hydroxylase-labeled somata. Substance P-labeled terminals also formed asymmetric synapses with unlabeled dendrites, and many unlabeled terminals formed both symmetric and asymmetric synapses with tyrosine hydroxylase-labeled dendrites. These results demonstrate that substance P neurons form a significant number of synapses with the dendrites of noradrenergic A7 neurons and support the conclusion that microinjection of substance P in the A7 cell group produces antinociception by direct activation of spinally projecting noradrenergic neurons.     

Ultrastructural changes in the dorsal motor nucleus of monkey following bilateral cervical vagotomy

Summary The neurons of the dorsal motor nucleas (DMN) of the monkey (Macaca fascicularis) were of two main types: small (13 × 8 m) and medium-sized (20 × 13 m). The latter, which were the predominant form, contained a pale oval nucleus surrounded by organelle-rich cytoplasm. Between one and three long principal dendrites per section profile arose from eac¹ of the somata. Both axosomatic and axodendritic synapses were seen on these cells although the latter were more common.No structural changes were noted in the DMN 1–3 days after bilateral cervical vagotomy. Some of the dendrites of the medium-sized axotomized vagal neurons appeared darkened 5–10 days after the operation. With longer surviving intervals, i.e. 21 and 28 days after operation, darkened dendrites were more commonly seen and the cytoplasmic density of these dendrites was dramatically enhanced. Their mitochondria were pale and some of them also showed vesiculation. Both normal and degenerating axon terminals were seen to form synaptic contacts with the darkened dendrites. The degenerating axon terminals were characterized by the clumping of their round agranular vesicles. Both darkened dendrites and degenerating axon terminals were phagocytosed by hypertrophied astrocytes and activated microglial cells. Blood elements infiltrating into the DMN were a possible source for some of the neural macrophages.It was concluded from the present study that the dendrites of the vagal neurons were the first structures to degenerate in axotomy and these were subsequently removed by glial elements. Degenerating axon terminals on the darkened dendrites could represent endings of the central processes of peripheral vagal ganglion cells that had undergonetransganglionic degeneration after damage to their peripheral processes.     

大鼠纹状体parvalbumin阳性中间神经元的超微结构

目的:研究纹状体parvalbumin (Parv)阳性中间神经元在神经通路上的突触连接.方法:利用免疫组织化学和神经示踪方法标记SD大鼠纹状体Parv及其相关神经元,光镜和电镜观察阳性神经元的结构和位置关系.结果:Parv阳性中间神经元中等大小,散在分布于纹状体,以背外侧居多.光镜免疫双标记显示Parv阳性中间神经元与皮质、丘脑和中脑黑质的传入轴突终末形成明显的形态位置上的邻近关系,其轴突终末则与纹状体不同类型投射神经元在光镜下也形成邻近关系.Parv阳性中间神经元的免疫电镜观察显示阳性产物主要游离于胞体、树突和轴突的胞质内.Parv阳性中间神经元的胞体和树突均接受大量的非对称型突触传入.Parv阳性轴突终末平均大小为(0.62±0.28)μm,可见其与纹状体神经元的树突、胞体和树突棘形成对称型突触,其中与树突形成的突触占69.64%,与胞体和树突棘形成的突触分别为26.78%和3.58%.结论:纹状体Parv阳性中间神经元形态学上与皮质、丘脑、黑质以及纹状体投射神经元形成突触连接,提示其可能在调节纹状体信息传入和输出过程中具有重要作用.     

Fetal neostriatal transplants in the rat: a light and electron microscopic Golgi study.

Fetal striatal neurons were transplanted into the ibotenic acid lesioned rat striatum. Three months after transplantation the grafted tissue was Golgi-impregnated and examined at the light microscopic level to determine the morphological characteristics of the transplanted neurons. Golgi-impregnated neurons were then gold-toned and examined at the electron microscopic level. The transplanted neurons were classified by both somatic size and somatic and dendritic morphology, which demonstrated that at least seven distinct cell types are present in striatal grafts. Type I large neurons had aspinous somata, sparsely spined dendrites, and indented nuclei, whereas type II large neurons displayed somatic spines, sparsely spined dendrites, and indented nuclei. Type I medium neurons exhibited aspinous somata and proximal dendrites, heavily spined distal dendrites, and unindented nuclei. Type II medium neurons had somatic spines, sparsely spined dendrites, and indented nuclei. Type III medium neurons had aspinous somata, poorly branched and sparsely spined dendrites, and indented nuclei, while type IV medium neurons had aspinous somata, highly branched and sparsely spined dendrites, and indented nuclei. Type V medium neurons displayed aspinous somata, varicose dendrites, and indented nuclei. These results demonstrate that transplanted fetal striatal neurons differentiate into morphologically and ultrastructurally distinct striatal cell types.     

大鼠中脑导水管周围灰质腹外侧区5-羟色胺样、P物质样和亮氨酸-脑啡肽样免疫反应阳性亚微结构的电镜观察

本文用免疫电镜技术研究了大鼠中脑导水管周围灰质腹外侧区内5-HT样、SP样和L-ENK样的免疫反应阳性亚微结构。5-HT样免疫反应阳性的胞体较多,常见5-HT样阳性树突与阴性轴突终末形成多为非对称性的轴-树突触;偶见阳性轴突终末与阴性树突以及阴性轴突终末与阳性胞体分别构成轴-树和轴-体突触.SP样阳性胞体数目较少,可见少量含多形性小泡的阴性轴突终末与之形成轴-体突触;由阴性轴突终末与阳性树突所形成的轴-树突触最常见;阳性轴突终末与阴性胞体和阳性树突分别构成轴-体突触和轴-树突触。L-ENK样阳性胞体数目也较少,L-ENK样阳性树突与阴性轴突终末所形成的轴-树突触最多见,可见L-ENK样阳性胞体与阴性轴突终末构成轴-体突触;偶见阳性轴突终末与阴性树突形成轴-树突触。上述各种突触均主要含圆形小泡,有时有少量扁平小泡、椭圆形小泡和颗粒囊泡。