Convergent vasopressinergic and hippocampal input onto somatospiny neurons of the rat lateral septal area

Electron microscopic immunocytochemistry, was combined with acute anterograde axon degeneration, following transection of the fimbria-fornix, to describe the innervation of somatospiny neurons by vasopressin-immunoreactive and degenerated hippocamposeptal axon terminals in the rat lateral septal area. Vasopressin-immunopositive boutons characterized by symmetric synaptic membrane specializations, and the degenerated hippocamposeptal axon terminals which form asymmetric synaptic contacts, frequently terminate on the same dendritic and somatic profiles, and particularly on the somata of somatospiny neurons. Although hippocamposeptal fibers predominantly form axospinous synapses in the lateral septal area, they terminate mainly on the dendritic shafts and soma of the vasopressin-receptive neurons. Of 720 vasopressin-immunoreactive terminals in the mediolateral part of the lateral septal area, 80% form synaptic contacts with dendritic shafts; 50% on small (distal) dendritic profiles and 30% on large (proximal) dendrites. Synaptic contacts between vasopressin-immunoreactive terminals and dendritic spines were not observed. The remaining 20% of immunoreactive boutons formed axosomatic synaptic contacts with a total of 58 neurons; 31% of these neurons exhibited somatic spines in the plane of the section analysed. Previous studies have demonstrated that in the lateral septal area vasopressin modulates the action of the excitatory amino acid-containing hypocamposeptal fibers, and also plays a role in the maintenance of long term potentiation evoked by fimbria-fornix stimulation. The convergent vasopressinergic and hippocampal input onto the same somatospiny neurons of the lateral septal area suggests that these neurons are targets of these physiological actions.     

Tyrosine hydroxylase-immunoreactive boutons in synaptic contact with identified striatonigral neurons, with particular reference to dendritic spines

Tyrosine hydroxylase-immunoreactive fibres in the rat neostriatum were studied in the electron microscope in order to determine the nature of the contacts they make with other neural elements. The larger varicose parts of such fibres contained relatively few vesicles and rarely displayed synaptic membrane specializations; however, thinner parts of axons (0.1-0.4 micron) contained many vesicles and had symmetrical membrane specializations, indicative of en passant type synapses. By far the most common postsynaptic targets of tyrosine hydroxylase-immunoreactive boutons were dendritic spines and shafts, although neuronal cell bodies and axon initial segments also received such input. Six striatonigral neurons in the ventral striatum were identified by retrograde labelling with horseradish peroxidase and their dendritic processes were revealed by Golgi impregnation using the section-Golgi procedure. The same sections were also developed to reveal tyrosine hydroxylase immunoreactivity and so we were able to study immunoreactive boutons in contact with the Golgi-impregnated striatonigral neurons. Each of the 280 immunoreactive boutons examined in the electron microscope displayed symmetrical synaptic membrane specializations: 59% of the boutons were in synaptic contact with the dendritic spines, 35% with the dendritic shafts and 6% with the cell bodies of striatonigral neurons. The dendritic spines of striatonigral neurons that received input from immunoreactive boutons invariably also received input, usually more distally, from unstained boutons that formed asymmetrical synaptic specializations. A study of 87 spines along the dendrites of an identified striatonigral neuron showed that the most common type of synaptic input was from an individual unstained bouton making asymmetrical synaptic contact (53%), while 39% of the spines received one asymmetrical synapse and one symmetrical immunoreactive synapse. It is proposed that the spatial distribution of presumed dopaminergic terminals in synaptic contact with different parts of striatonigral neurons has important functional implications. Those synapses on the cell body and proximal dendritic shafts might mediate a relatively non-selective inhibition. In contrast, the major dopaminergic input that occurs on the necks of dendritic spines is likely to be highly selective since it could prevent the excitatory input to the same spines from reaching the dendritic shaft. One of the main functions of dopamine released from nigrostriatal fibres might thus be to alter the pattern of firing of striatal output neurons by regulating their input.     

Catecholaminergic neurons in medullary nuclei are among the post-synaptic targets of descending projections from infralimbic area 25 of the rat medial prefrontal cortex

The infralimbic (IL) 'visceromotor' area of the rat medial prefrontal cortex projects to strategic subcortical nuclei involved in autonomic functions. Central among these targets are the nucleus tractus solitarius (NTS) and the rostral ventrolateral medulla (rVLM). By combining tract-tracing using the anterograde tracer biotinylated dextran amine (BDA) with immunolabeling for tyrosine hydroxylase (TH; an enzyme marker of catecholaminergic neurons), a limited proportion of BDA-labeled IL axonal boutons in the NTS and rVLM was found to be closely associated with TH immunopositive (+) target structures. Such structural appositions were mainly located proximally over the labeled dendritic arbors of identified TH+ neurons. Quantitative ultrastructural examination revealed that in NTS, TH+ dendritic shafts comprised 7.0% of the overall post-synaptic target population innervated by BDA-labeled IL boutons, whereas TH+ dendritic spines represented 1.25% of targets. In rVLM, TH+ shafts represented 9.0% and TH+ spines 2.5% of IL targets. Labeled IL boutons established exclusively asymmetric Gray Type 1 (presumed excitatory) synaptic junctions. The results indicate that subpopulations of catecholaminergic neurons in the NTS and rVLM are among the spectrum of post-synaptic neurons monosynaptically innervated by descending 'excitatory' input from IL cortex. Such connectivity, albeit restricted, identifies the potential direct influence of IL cortex on the processing and distribution of cardiovascular, respiratory and related autonomic information by catecholaminergic neurons in the NTS and VLM of the rat.     

Total number and distribution of inhibitory and excitatory synapses on hippocampal CA1 pyramidal cells

The integrative properties of neurons depend strongly on the number, proportions and distribution of excitatory and inhibitory synaptic inputs they receive. In this study the three-dimensional geometry of dendritic trees and the density of symmetrical and asymmetrical synapses on different cellular compartments of rat hippocampal CA1 area pyramidal cells was measured to calculate the total number and distribution of excitatory and inhibitory inputs on a single cell.A single pyramidal cell has approximately 12,000 microm dendrites and receives around 30,000 excitatory and 1700 inhibitory inputs, of which 40 % are concentrated in the perisomatic region and 20 % on dendrites in the stratum lacunosum-moleculare. The pre- and post-synaptic features suggest that CA1 pyramidal cell dendrites are heterogeneous. Strata radiatum and oriens dendrites are similar and differ from stratum lacunosum-moleculare dendrites. Proximal apical and basal strata radiatum and oriens dendrites are spine-free or sparsely spiny. Distal strata radiatum and oriens dendrites (forming 68.5 % of the pyramidal cells' dendritic tree) are densely spiny; their excitatory inputs terminate exclusively on dendritic spines, while inhibitory inputs target only dendritic shafts. The proportion of inhibitory inputs on distal spiny strata radiatum and oriens dendrites is low ( approximately 3 %). In contrast, proximal dendritic segments receive mostly (70-100 %) inhibitory inputs. Only inhibitory inputs innervate the somata (77-103 per cell) and axon initial segments. Dendrites in the stratum lacunosum-moleculare possess moderate to small amounts of spines. Excitatory synapses on stratum lacunosum-moleculare dendrites are larger than the synapses in other layers, are frequently perforated ( approximately 40 %) and can be located on dendritic shafts. Inhibitory inputs, whose percentage is relatively high ( approximately 14-17 %), also terminate on dendritic spines.Our results indicate that: (i) the highly convergent excitation arriving onto the distal dendrites of pyramidal cells is primarily controlled by proximally located inhibition; (ii) the organization of excitatory and inhibitory inputs in layers receiving Schaffer collateral input (radiatum/oriens) versus perforant path input (lacunosum-moleculare) is significantly different.     

Dendritic and somatic appendages of identified rubrospinal neurons of the cat

Giant neurons of the red nucleus of the cat were stained intracellularly with horseradish peroxidase and examined using light microscopy, electron microscopy of thin sections, and high voltage electron microscopy of thick sections (2-5 microns). Special attention was paid to the arrangement of dendritic spines and other appendages relative to the distribution of synaptic contacts from known sources. In the region of the neuron known to receive synaptic contacts from the nucleus interpositus of the cerebellum (soma and proximal 200-300 microns of dendrites), the dendrites were relatively unbranched, and free of long spines or complex appendages. The surface of the neurons in this region was covered with a dense layer of short thin appendages that invaginated or penetrated between the synaptic terminals that cover this part of the cells. The small spines received synapses of the types associated both with the cerebellar afferent fibers and with the local inhibitory interneurons. These same terminals made synaptic contacts directly onto the surface of the neurons and onto the lateral surfaces of the spines, suggesting that the spines may serve primarily to increase the available synaptic surface area. The more distal portion of the dendritic field, where cerebellar afferents do not make synaptic contacts, exhibited a dramatically different appearance. The dendrites were much more branched, and exhibited many and varied dendritic appendages. The appendages were of three general types. One was a large protrusion with a cup-shaped head that formed the principal postsynaptic component of a glomerular arrangement also involving an axon terminal and usually a presynaptic dendrite. A second was a long thin filiform process that usually occurred around the glomeruli. This appendage was occasionally postsynaptic. The third was a spherical appendage containing many lysosomal organelles resembling residual bodies. The glomerular dendritic protrusions were very common in the distal portion of the dendritic field, numbering at least 1000 per cell. At least some of the glomeruli are specialized for receipt of synaptic input from the corticorubral pathway, since lesions of sensorimotor cortex resulted in degeneration of the central synaptic terminal in some glomeruli on horseradish peroxidase-injected rubrospinal neurons. These specializations of dendritic structure may contribute to the differences in excitatory postsynaptic potential wave shape between cortical and cerebellar inputs, and they may play a role in the changes in the cortical excitatory postsynaptic potential that develop after lesions of cerebellar inputs.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nucleus accumbens nitric oxide immunoreactive interneurons receive nitric oxide and ventral subicular afferents in rats

The nitric oxide generating neurons of the nucleus accumbens exert a powerful influence over striatal function, in addition, these nitrergic inputs are in a position to regulate the dopaminergic and glutamatergic inputs on striatal projection neurons. It was the aim of this study to establish the source of the glutamatergic drive to nitric oxide synthase interneurons of the nucleus accumbens. The nucleus accumbens nitric oxide-generating neurons receive asymmetrical, excitatory, presumably glutamatergic inputs. Possible sources of these inputs could be the limbic and cortical regions known to project to this area. To identify sources of the excitatory inputs to the nitric oxide synthase-containing interneurons of the nucleus accumbens in the rat we first examined the ultrastructural morphology of asymmetrical synaptic specializations contacting nitric oxide synthase-immunohistochemically labeled interneurons in the nucleus accumbens. Neurons were selected from different regions of the nucleus accumbens, drawn using camera lucida, processed for electron microscopic analysis, and the boutons contacting nitric oxide synthase-labeled dendrites were photographed and correlated to the drawings. Using vesicle size as the criterion the source was predicted to be either the prefrontal cortex or the ventral subiculum of the hippocampus. To examine this prediction, a further study used anterograde tracing from both the prefrontal cortex and the ventral subiculum, and nitric oxide synthase immunohistochemistry with correlated light and electron microscopy. Based on appositions by anterogradely labeled fibers, selected nitric oxide synthase-labeled neurons within the nucleus accumbens, were examined with electron microscopic analysis. With this technique we confirmed the prediction that subicular afferent boutons make synaptic contact with nitric oxide synthase interneurons, and demonstrated anatomically that nitric oxide synthase boutons make synaptic contact with the dendritic arbors of nitric oxide synthase interneurons. We suggest that the subicular input may excite the nitric oxide synthase neurons synaptically, while the nitric oxide synthase-nitric oxide synthase interactions underlie a nitric oxide signaling network which propagates hippocampal information, and expands the hippocampus's influence on 'gating' information flow across the nucleus accumbens.     

Increased gabaergic input to ventral tegmental area dopaminergic neurons associated with decreased cocaine reinforcement in mu-opioid receptor knockout mice

There is general agreement that dopaminergic neurons projecting from the ventral tegmental area (VTA) to the nucleus accumbens and prefrontal cortex play a key role in drug reinforcement. The activity of these neurons is strongly modulated by the inhibitory and excitatory input they receive. Activation of mu-opioid receptors, located on GABAergic neurons in the VTA, causes hyperpolarization of these GABAergic neurons, thereby causing a disinhibition of VTA dopaminergic neurons. This effect of mu-opioid receptors upon GABA neurotransmission is a likely mechanism for mu-opioid receptor modulation of drug reinforcement. We studied mu-opioid receptor signaling in relation to cocaine reinforcement in wild-type and mu-opioid receptor knockout mice using a cocaine self-administration paradigm and in vitro electrophysiology. Cocaine self-administration was reduced in mu-opioid receptor knockout mice, suggesting a critical role of mu-opioid receptors in cocaine reinforcement. The frequency of spontaneous inhibitory post-synaptic currents onto dopaminergic neurons in the ventral tegmental area was increased in mu-opioid receptor knockout mice compared with wild-type controls, while the frequency of spontaneous excitatory post-synaptic currents was unaltered. The reduced cocaine self-administration and increased GABAergic input to VTA dopaminergic neurons in mu-opioid receptor knockout mice supports the notion that suppression of GABAergic input onto dopaminergic neurons in the VTA contributes to mu-opioid receptor modulation of cocaine reinforcement.     

Synaptic organization of intracellularly stained CA3 pyramidal neurons in slice cultures of rat hippocampus

Pyramidal cells of regio inferior in slice cultures of the rat hippocampus were impaled and intracellularly stained with horseradish peroxidase. A correlated light- and electron-microscopic analysis was then performed to study the properties of these neurons under culture conditions with particular emphasis on input synapses onto these cells. Like pyramidal cells in situ, CA3 pyramidal neurons in slice cultures had a triangular cell body with an apical stem dendrite emerging from it. Several basal dendrites and the axon arose from the basal pole of the cell body. The peripheral thin branches of both apical and basal dendrites were covered with small spines, whereas proximal thick dendritic segments and portions of the cell body exhibited large spines or excrescences. The axon gave off numerous fine varicose collaterals which projected to stratum radiatum of CA1 (Schaffer collaterals), to the alveus and to the hilar region. In one case a collateral could be followed to stratum moleculare of the fascia dentata. Electron-microscopic analysis of the injected pyramidal neurons revealed that their cell bodies, dendritic shafts and spines formed synaptic contacts with presynaptic terminals. Mossy fiber endings were identified by their large size and their numerous clear synaptic vesicles with some dense-core vesicles intermingled, and were observed to form synaptic contacts on the large spines or excrescences. Since extrinsic afferents degenerate in slice cultures, the numerous synaptic boutons on the identified pyramidal neurons probably arise from axons of intrinsic neurons that have sprouted in response to deafferentation. This assumption is supported by the finding that collaterals of the injected neurons formed abundant synaptic contacts on dendritic shafts and spines of other cells. These results suggest that, although pyramidal cells under culture conditions retain a remarkable number of their normal characteristics, considerable synaptic reorganization does take place.     

The postsynaptic targets of substance P-immunoreactive terminals in the rat neostriatum with particular reference to identified spiny striatonigral neurons

Summary Substance P-immunoreactive boutons were examined in the electron microscope in sections of the rat neostriatum that contained retrogradely labelled striatonigral neurons and/or Golgi-impregnated medium-size densely spiny neurons. The postsynaptic targets of the immunoreactive boutons were characterized on the basis of ultrastructural features, their projection to the substantia nigra and/or their somato-dendritic morphology. Substance P-immunoreactive axonal boutons formed symmetrical synaptic specializations. Of a total of 233 randomly identified synaptic boutons 72.5% made contact with dendritic shafts, 15% with dendritic spines and 10.7% with perikarya. The ultrastructural characteristics of some of the postsynaptic neuronal perikarya were consistent with their identification as striatal interneurons. Similarly, the observation of some of the substance P-containing terminals in contact with spines, spine-bearing dendritic shafts and perikarya with the ultrastructural characteristics of medium-size densely spiny neurons suggested that one of the targets of substance P-positive terminals are striatal projection neurons. Direct evidence for this was obtained in sections from rats that had received injections of horseradish peroxidase conjugated with wheatgerm agglutinin in the substantia nigra. The perikarya of retrogradely labeled striatonigral neurons were found to receive symmetrical synaptic input from substance P-positive boutons. Ultrastructural analysis of Golgi-impregnated medium-size densely spiny neurons, some of which were also retrogradely labeled from the substantia nigra, demonstrated directly that this class of neuron was postsynaptic to the substance P-immunoreactive boutons. The combination of Golgi-impregnation with substance P-immunocytochemistry made it possible to study the pattern or topography of the substance P-positive input to medium size densely spiny neurons. The substance P-containing boutons made contact predominantly with perikarya and dendritic shafts. This pattern of input is markedly different from that of other identified inputs to medium-size densely spiny neurons.     

Descriptive findings on the morphology of dendritic spines in the rat medial amygdala

The rat posterodorsal medial amygdala (MePD) is a brain area in which gonadal hormones induce notable plastic effects in the density of dendritic spines. Dendritic spines are post-synaptic specializations whose shape and spacing change neuronal excitability. Our aim was to obtain new data on the dendritic spines morphology and density from MePD neurons using the carbocyanine dye DiI under confocal microscopy. In adult male rats, the dendritic spine density of the medial branches of the left MePD (mean ± SD) was 1.15 ± 0.67 spines/dendritic μm. From the total sampled, approximately 53% of the spines were classified as thin, 22.5% as “mushroom-like”, and 21.5% as stubby/wide. Other spine shapes (3%) included those ramified, with a filopodium-like or a gemule appearance, and others with a protruding spinule. Additional experiment joining DiI and synaptophysin (a pre-synaptic protein) labeling suggested synaptic sites on dendritic shafts and spines. Dendritic spines showed synaptophysin puncta close to their head and neck, although some spines had no evident labeled puncta on them or, conversely, multiple puncta appeared upon one spine. These results advance previous light microscopy results by revealing features and complexities of the dendritic spines at the same time that give new insight on the possible synaptic organization of the adult rat MePD.     

Kappa opioid receptor inhibition of glutamatergic transmission in the nucleus accumbens shell

Microinjection of kappa-opioid receptor agonists into the nucleus accumbens produces conditioned place aversion. While attention has focused primarily on the inhibition of dopamine release by kappa-receptor agonists as the synaptic mechanism underlying this effect, recent anatomical studies have raised the possibility that regulation of noncatecholaminergic transmission also contribute. We have investigated the effects of kappa-receptor activation on fast excitatory synaptic transmission in an in vitro slice preparation using whole cell voltage-clamp or extracellular field recordings in the shell region of the nucleus accumbens. The kappa-receptor agonist U69593 produces a pronounced, dose-dependent inhibition of glutamatergic excitatory postsynaptic currents (EPSCs) that can be reversed by 100 nM nor-BNI. Furthermore, U69593 causes an increase in the paired-pulse ratio as well as a decrease in the frequency of spontaneous miniature events, suggesting a presynaptic site of action. Despite anatomical evidence for kappa-receptor localization on dendritic spines of nucleus accumbens neurons, no electrophysiological evidence of a postsynaptic effect was found. This presynaptic inhibition of excitatory synaptic transmission in the nucleus accumbens shell provides a novel mechanism that may contribute to the kappa-receptor-mediated aversion observed in intact animals.     

Electron-microscopic study of dopaminergic structures in the medial subdivision of the monkey nucleus accumbens

The medial subdivision of the monkey nucleus accumbens (NAC) is rich in dopamine (DA) and peptides. In the present investigation the mode of DA transmission in the medial subdivision was studied morphologically by light- and electron-microscopic immunocytochemistry using a monoclonal antibody raised against dopamine. The medial subdivision showed extremely dense accumulation of thick DA-immunoreactive varicose fibers. Electron-microscopic observation of single sections revealed that DA afferents had a relatively high incidence (33.2%) of asymmetric junctions in this area. Approximately 50% of the targets were dendritic shafts, 44.2% dendritic spines, and 5.1% somata. Some DA axons showed terminal profiles en passant within the synaptic complex, some of which showed synaptic triads. The unique ultrastructural features of DA terminals in the medial NAC indicate the existence of specific styles of DA transmission in the limbic structure.     

Differentiation of granule cells in relation to GABAergic neurons in the rat fascia dentata

Summary Golgi impregnation was used to study the dendritic differentiation of granule cells in the rat fascia dentata. The impregnated granule cells were gold-toned allowing for a fine structural study of the same identified neurons and of the input synapses onto their cell bodies and dendrites. Due to the long postnatal formation of these cells it was possible to describe a sequence of maturational stages coexisting on the same postnatal day (P5). Characteristic features of the dendritic development of granule cells were i) occurrence of varicose swellings along the dendrites, ii) growth cones on dendritic tips, iii) transient formation of basal dendrites, and iv) progressive development of dendritic spines. Incoming synapses on the differentiating granule cells were mainly found on dendritic shafts. Their membrane specializations were symmetric. At least some of these symmetric synapses were GABAergic because immunostaining of Vibratome sections from the same postnatal stage (P5) demonstrated a well-developed GABAergic axon plexus in the fascia dentata (antibodies against glutamate decarboxylase (GAD), the GABA synthesizing enzyme). Electron microscopy of the immunostained axon plexus revealed numerous GABAergic terminals that formed symmetric synaptic contacts, mainly on shafts of differentiating dendrites but also on cell bodies of granule cells. Our results thus indicate that the plexus of inhibitory GABAergic axons is already well developed at a stage when the target neurons, the granule cells, are still being formed.     

Synaptic relationships of a type of GABA-immunoreactive neuron (clutch cell), spiny stellate cells and lateral geniculate nucleus afferents in layer IVC of the monkey striate cortex

The precise stimulus specificity of striate cortical neurons is strongly influenced by processes involving gamma-aminobutyric acid (GABA). In the visual cortex of the monkey most afferents from the lateral geniculate nucleus terminate in layer IVC. We identified a type of smooth dendritic neuron (clutch cell) that was immunoreactive for GABA, and whose Golgi-impregnated dendrites and axon were largely restricted to layer IVC beta. The slightly ovoid somata were 8-12 micron by 12-15 micron and the dendritic field was often elongated, extending 80-200 micron in one dimension. The axonal field was 100-150 micron in diameter and densely packed with large bulbous boutons. Although mainly located in IVC beta both the dendritic and axonal processes entered IVC alpha. Fine structural features of GABA-immunoreactive and-impregnated clutch cells and impregnated spiny stellate cells were compared. Clutch cells had more cytoplasm, more densely packed mitochondria and endoplasmic reticulum, and made type II as opposed to type I synapses. A random sample of 159 elements postsynaptic to three clutch cells showed that they mainly terminated on dendritic shafts (43.8-58.5%) and spines (20.8-46.3%), rather than somata (10-17%). The majority of the postsynaptic targets were characteristic of spiny stellate cells. This was directly demonstrated by studying synaptic contacts between an identified GABA positive clutch cell and the dendrites and soma of an identified spiny stellate cell. The termination of clutch cells mainly on dendrites and spines of spiny stellate cells suggests that they interact with other inputs to the same cells. Following an electrolytic lesion in the ipsilateral lateral geniculate nucleus we examined the distribution of degenerating terminals on three identified spiny stellate neurons in layer IVC beta. Out of eight synapses from the lateral geniculate nucleus one was on a dendritic shaft, the rest on spines. Only a small fraction of all synapses on the cells were from degenerating boutons. A clutch cell within the area of dense terminal degeneration was not contacted by terminals from the lateral geniculate nucleus. The results show that layer IVC in the monkey has a specialized GABAergic neuron that terminates on spiny stellate cells monosynaptically innervated from the lateral geniculate nucleus. Possible functions of clutch cells may include inhibitory gating of geniculate input to cortex; maintenance of the antagonistic subregions in the receptive fields; and the creation from single opponent of double colour opponent receptive fields.     

Synaptic modifications at the CA3–CA1 synapse after chronic AMPA receptor blockade in rat hippocampal slices

Maintenance of dendritic spines, the postsynaptic elements of most glutamatergic synapses in the central nervous system, requires continued activation of AMPA receptors. In organotypic hippocampal slice cultures, chronic blockade of AMPA receptors for 14 days induces a substantial loss of dendritic spines on CA1 pyramidal neurons. Here, using serial section electron microscopy, we show that loss of dendritic spines is paralleled by a significant reduction in synapse density. In contrast, we observed an increased number of asymmetric synapses onto the dendritic shaft, suggesting that spine retraction does not inevitably lead to synapse elimination. Functional analysis of the remaining synapses revealed that hippocampal circuitry compensates for the anatomical loss of synapses by increasing synaptic efficacy. Moreover, we found that the observed morphological and functional changes were associated with altered bidirectional synaptic plasticity. We conclude that continued activation of AMPA receptors is necessary for maintaining structure and function of central glutamatergic synapses.     

A quantitative analysis of the laminar distribution of synaptic boutons in field CA3 of the rat hippocampus

We analyzed the laminar distribution of synaptic boutons in field CA3 of the rat hippocampus using a large montage electron micrograph. The size of boutons and synaptic vesicles was measured using a computer-assisted digitizing system. In all, 3353 synaptic boutons were observed in a 15 microm x 100 microm strip. Of these, 86.3% contained spherical vesicles (S-boutons), 12% contained flat vesicles (F-boutons), and 1.7% were mossy terminals (M-boutons). S-boutons were distributed widely in the strata moleculare (st. Mol), radiatum (st. Rad), and oriens (st. Ori), but there were only a few in the strata lucidum (st. Luc) and pyramidale (st. Pyr). The upper portions of both the st. Rad and Ori contained slightly fewer boutons. In terms of the location of synaptic contacts, 83% of all S-boutons were found on the dendritic spines and the rest were on the dendritic shafts. S-boutons on the dendritic shafts were observed more frequently in the st. Mol than in the other strata. According to the morphometry of the size of synaptic vesicles, S-boutons with small vesicles (mean vesicle area <1109 nm(2)) were located exclusively in the st. Mol, S-boutons with medium-sized vesicles (mean vesicle area 1109-1482 nm(2)) were observed in all strata, and S-boutons with large vesicles (mean vesicle area >1482 nm(2)) were distributed in the st. Luc and Ori, but not in the st. Mol. F-boutons were predominantly distributed in the upper half of the st. Mol and in the area around the st. Pyr, although they were observed in all strata. In the st. Mol, all the F-boutons were in contact with dendritic shafts, while near the st. Pyr, F-boutons were found exclusively on somata, the proximal parts of the dendritic shafts, and the initial segments of axons. The average F-bouton was smaller in the st. Mol (0.23 microm(2)) than near the st. Pyr (0.39 microm(2)). In this synapto-architectural study of the hippocampal CA3 region using large montage electron micrographs, we observed (1) an intimate relationship between synapse distribution and the dendritic structure of pyramidal neurons, (2) the distribution of different types of boutons containing vesicles of various size, and (3) two different plausible foci of postsynaptic inhibition where F-boutons were distributed densely, and (4) estimated the input ratios of pyramidal neurons.     

The thalamic projection to cat visual cortex: ultrastructure of neurons identified by Golgi impregnation or retrograde horseradish peroxidase transport

Specific afferents to the primary visual cortex of the cat were identified by electron microscopy after electrolytic lesions of the lateral geniculate nucleus. Postsynaptic target neurons were marked by either Golgi impregnation or horseradish peroxidase retrogradely transported from the opposite visual cortex. The type and the location of identified neurons were determined by light microscopy before further investigation by electron microscopy. Different pyramidal neurons have relatively homogeneous ultrastructural characteristics, but non-pyramidal neurons, divisible into large and small, spiny, sparsely-spiny and non-spiny, vary in their synaptology. Thalamo-cortical afferents have been found to all neuronal types examined in layer IV and lower layer III, including horseradish peroxidase-filled callosal cells. These include pyramidal cells of layer III which receive thalamo-cortical afferents mainly on spines of apical and basal dendrites, but also on basal dendritic shafts. Layer V pyramids receive thalamo-cortical input to apical dendritic spines. All types of layer IV non-pyramidal neurons studied are contacted by geniculate terminals on dendritic spines and shafts and directly on the cell body.We conclude that input to the visual cortex is organized on both hierarchical and parallel bases.     

A light and electron microscopic study of NADPH-diaphorase-, calretinin- and parvalbumin-containing neurons in the rat nucleus accumbens

The rat nucleus accumbens contains medium-sized, spiny projection neurons and intrinsic, local circuit neurons, or interneurons. Sub-classes of interneurons, revealed by calretinin (CR) or parvalbumin (PV) immunoreactivity or reduced nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, were compared in the nucleus accumbens core, shell and rostral pole. CR, PV and NADPH-diaphorase-containing neurons are shown to form three non-co-localising populations in these three areas. No significant differences in neuronal population densities were found between the subterritories. NADPH-diaphorase-containing neurons could be further separated morphologically into three sub-groups, but CR- and PV-immunoreactive neurons form homogeneous populations. Ultrastructurally, NADPH-diaphorase-, CR- and PV-containing neurons in the nucleus accumbens all possess nuclear indentations. These are deeper and fewer in neurons immunoreactive for PV than in CR- and NADPH-diaphorase-containing neurons. CR-immunoreactive boutons form asymmetrical and symmetrical synaptic specialisations on spines, dendrites and somata, while PV-immunoreactive boutons make only symmetrical synaptic specialisations. Both CR- and PV-immunoreactive boutons form symmetrical synaptic specialisations with medium-sized spiny neurons and contact other CR- and PV-immunoreactive somata, respectively. A novel non-carcinogenic substrate for the peroxidase reaction (Vector Slate Grey, SG) was found to be characteristically electron-dense and may be distinguishable from the diaminobenzidine reaction product. We conclude that the three markers used in this study are localised in distinct populations of nucleus accumbens interneurons. Our studies of their synaptic connections contribute to an increased understanding of the intrinsic circuitry of this area.     

Membrane structure at synaptic junctions in area CA1 of the rat hippocampus

In tissue from area CA1 of the rat hippocampus prepared for electron microscopic study by thin-sectioning, asymmetric synaptic junctions were found on dendritic spines, spiny dendritic shafts, and non-spiny dendritic shafts. In freeze-fractured preparations, aggregates of large particles were found on the extracellular half of the postsynaptic membrane at each of these synaptic junctions. Particle aggregate areas were measured and particle packing densities were computed at dendritic spine synapses and dendritic shaft synapses in area CA1, and compared to similar measures of particle aggregates on dendritic spines of cerebellar Purkinje cells. All of these CA1 and cerebellar synapses are excitatory and are thought to use glutamate as a neurotransmitter. There was a tendency for the dispersion of particles within individual aggregates to be less uniform in larger aggregates in both area CA1 and cerebellar cortex. Distinct particle-free zones could be distinguished in the center of particle aggregates on large "mushroom-shaped" spines in area CA1. There was no statistically significant difference between the particle densities at CA1 dendritic spines (2848 +/- 863 particles/micron2) and CA1 dendritic shafts (2707 +/- 718 particles/micron2). However, the average density of particles at cerebellar dendritic spine synapses (3614 +/- 1081 particles/micron2) was significantly greater than at dendritic spine or shaft synapses found in area CA1. Symmetric synaptic junctions were observed on the CA1 pyramidal cell somas and dendritic shafts in thin-sectioned preparations. These synapses typically exert an inhibitory action mediated by gamma-aminobutyric acid. In freeze-fracture preparations, large varicosities were found apposed to the pyramidal somal and dendritic membranes, but there were no specializations of particle distribution on either the extracellular or the cytoplasmic half of the fractured postsynaptic membranes. This finding parallels observations from freeze-fracture preparations of other GABAergic synapses in the central nervous system.     

Differential membrane properties and dopamine effects in the shell and core of the rat nucleus accumbens studied in vitro.

Electrophysiological differences between the shell and core of the rat nucleus accumbens were investigated by intracellular recordings from an in vitro slice preparation. The average input resistance of neurons recorded in the shell was larger than in the core. Neurons in the core were characterized by a more negative resting membrane potential than neurons in the shell. Furthermore, bath-applied dopamine attenuated synaptic responses recorded in the shell, but not in the core. Thus, the two main subregions of the nucleus accumbens differ both in basal membrane properties and in dopaminergic modulation of synaptic transmission.