Large variability in synaptic N-methyl-D-aspartate receptor density on interneurons and a comparison with pyramidal-cell spines in the rat hippocampus

Pyramidal cells receive input from several types of GABA-releasing interneurons and innervate them reciprocally. Glutamatergic activation of interneurons involves both alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) type glutamate receptors expressed in type I synapses, mostly on their dendritic shafts. On average, the synaptic AMPA receptor content is several times higher on interneurons than in the spines of pyramidal cells. To compare the NMDA receptor content of synapses, we used a quantitative postembedding immunogold technique on serial electron microscopic sections, and analysed the synapses on interneuron dendrites and pyramidal cell spines in the CA1 area. Because all NMDA receptors contain the obligatory NR1 subunit, receptor localisation was carried out using antibodies recognising all splice variants of the NR1 subunit. Four populations of synapse were examined: i). on spines of pyramidal cells in stratum (str.) radiatum and str. oriens; ii). on parvalbumin-positive interneuronal dendritic shafts in str. radiatum; iii). on randomly found dendritic shafts in str. oriens and iv). on somatostatin-positive interneuronal dendritic shafts and somata in str. oriens. On average, the size of the synapses on spines was about half of those on interneurons. The four populations of synapse significantly differed in labelling for the NR1 subunit. The median density of NR1 subunit labelling was highest on pyramidal cell spines. It was lowest in the synapses on parvalbumin-positive dendrites in str. radiatum, where more than half of these synapses were immunonegative. In str. oriens, synapses on interneurons had a high variability of receptor content; some dendrites were similar to those in str. radiatum, including the proximal synapses of somatostatin-positive cells, whereas others had immunoreactivity for the NR1 subunit similar to or higher than synapses on pyramidal cell spines.These results show that synaptic NMDA receptor density differs between pyramidal cells and interneurons. Some interneurons may have a high NMDA receptor content, whereas others, like some parvalbumin-expressing cells, a particularly low synaptic NMDA receptor content. Consequently, fast glutamatergic activation of interneurons is expected to show cell type-specific time course and state-dependent dynamics.     

Nitric oxide-containing pyramidal neurons of the subiculum innervate the CA1 area

Neurons and axon terminals containing neuron-specific nitric oxide synthase (nNOS) were examined in the rat subiculum and CA1 area of Ammon's horn. In the subiculum, a large subpopulation of the pyramidal neurons and non-pyramidal cells are immunoreactive for nNOS, whereas in the neighbouring CA1 area of Ammon's horn only non-pyramidal neurons are labelled with the antibody against nNOS. In the pyramidal layer of the subiculum, nNOS-positive axon terminals form both asymmetric and symmetric synapses. In the adjacent CA1 area the nNOS-positive terminals that form symmetric synapses are found in all layers, whereas those terminals that form asymmetric synapses are only in strata radiatum and oriens, but not in stratum lacunosum-moleculare. In both the subiculum and CA1 area, labelled terminals make symmetric synapses only on dendritic shafts, whereas asymmetric synapses are exclusively on dendritic spines. Previous observations demonstrated that all nNOS-positive non-pyramidal cells are GABAergic local circuit neurons, which form exclusively symmetric synapses. We suggest that nNOS-immunoreactive pyramidal cells of the subiculum may innervate neighbouring subicular pyramidal cells and, to a smaller extent, pyramidal cells of the adjacent CA1 area, forming a backward projection between the subicular and hippocampal principal neurons. Electronic Publication     

Apical dendritic depolarizations and field interactions evoked by stimulation of afferent inputs to rat hippocampal CA1 pyramidal cells

The relationship between orthodromic extracellular field potentials and intradendritic depolarizations in apical dendrites of CA1 pyramidal neurons was investigated using the in vitro slice preparation of rat hippocampus. Orthodromic synaptic field potentials evoked by stimulation of afferent inputs in stratum radiatum or stratum oriens were used to measure extracellular voltage gradients generated over the pyramidal cell axis. Extracellular gradients were of opposite polarity over the region of pyramidal cell apical dendrites in stratum radiatum. The stratum radiatum-evoked gradient was negative towards the apical dendrites and the stratum oriens-evoked gradient negative towards the cell body layer, with gradients reaching values of up to 50 mV/mm over the apical dendritic axis. Intradendritic recordings obtained greater than 150 microns from stratum pyramidale directly measured the subthreshold apical dendritic excitatory postsynaptic potentials evoked by stratum radiatum or stratum oriens stimulation. These ground-referenced recordings were then compared to the transmembrane potential calculated by subtraction of the corresponding extradendritic field potential. Both stratum radiatum and stratum oriens stimulation evoked graded excitatory postsynaptic potentials that could be recorded in apical dendritic impalements up to 265 microns from stratum pyramidale. The calculated transmembrane potential of the stratum radiatum-evoked excitatory postsynaptic potential had a significantly greater rate of rise, peak amplitude, and rate of decay than that of the ground-referenced excitatory postsynaptic potential. In contrast, the rates of rise and decay of the transmembrane potential of the stratum oriens-evoked excitatory postsynaptic potential were reduced with respect to the ground-referenced recording. The peak amplitude of the stratum oriens-evoked transmembrane potential, however, varied according to the polarity of the corresponding extradendritic population spike response recorded in stratum radiatum. These data reveal that synaptic activation of either basal or apical dendrites of CA1 pyramidal cells evokes a depolarization that can be recorded over a substantial region of the apical dendritic arbor. Furthermore, extradendritic field potentials evoked by stimulation of these inputs produce opposite effects on the transmembrane potential of apical dendrites. The magnitude of the accompanying extracellular voltage gradients suggest that these shifts in transmembrane potential reflect ephaptic interactions at the apical dendritic level of pyramidal cells.     

Agmatine containing axon terminals in rat hippocampus form synapses on pyramidal cells

We examined the cellular and subcellular localization of agmatine in the hippocampal CA1 region by immunocytochemistry. By light microscopy, agmatine-like immunoreactivity (agmatine-LI) was found primarily in the perikarya and dendritic profiles of pyramidal cells and in punctate processes preponderantly in stratum radiatum. Electron microscopy revealed that agmatine-LI was cytoplasmic and concentrated in ‘clusters' associated with mitochondria and tubular vesicles. In stratum radiatum, agmatine-LI was primarily in axons and axon terminals associated with small, synaptic vesicles. The terminals almost exclusively formed asymmetric synapses on the spines of dendrites, many of which originated from pyramidal cells. Some agmatine-LI also was present in shafts and spines of pyramidal cell dendrites and in astrocytic processes. The results demonstrate that agmatine in the hippocampus is found primarily in terminals forming excitatory (asymmetric) synapses on pyramidal cells, some of which contain agmatine-LI. These findings further implicate agmatine as an endogenous neurotransmitter which may be co-stored with -glutamate and may act in part in the rat hippocampus as a blocker of the N-methyl- -aspartate receptor and nitric oxide synthase.     

Different modes of synaptic and extrasynaptic NMDA receptor alteration in the hippocampus of P301S tau transgenic mice

N-methyl-d -aspartate receptors (NMDARs) are pivotal players in the synaptic transmission and synaptic plasticity underlying learning and memory. Accordingly, dysfunction of NMDARs has been implicated in the pathophysiology of Alzheimer disease (AD). Here, we used histoblot and sodium dodecylsulphate-digested freeze-fracture replica labelling (SDS-FRL) techniques to investigate the expression and subcellular localisation of GluN1, the obligatory subunit of NMDARs, in the hippocampus of P301S mice. Histoblots showed that GluN1 expression was significantly reduced in the hippocampus of P301S mice in a laminar-specific manner at 10 months of age but was unaltered at 3 months. Using the SDS-FRL technique, excitatory synapses and extrasynaptic sites on spines of pyramidal cells and interneuron dendrites were analysed throughout all dendritic layers in the CA1 field. Our ultrastructural approach revealed a high density of GluN1 in synaptic sites and a substantially lower density at extrasynaptic sites. Labelling density for GluN1 in excitatory synapses established on spines was significantly reduced in P301S mice, compared with age-matched wild-type mice, in the stratum oriens (so), stratum radiatum (sr) and stratum lacunosum-moleculare (slm). Density for synaptic GluN1 on interneuron dendrites was significantly reduced in P301S mice in the so and sr but unaltered in the slm. Labelling density for GluN1 at extrasynaptic sites showed no significant differences in pyramidal cells, and only increased density in the interneuron dendrites of the sr. This differential alteration of synaptic versus extrasynaptic NMDARs supports the notion that the progressive accumulation of phospho-tau is associated with changes in NMDARs, in the absence of amyloid-β pathology, and may be involved in the mechanisms causing abnormal network activity of the hippocampal circuit.     

The termination of callosal fibres in the auditory cortex of the rat. A combined Golgi--electron microscope and degeneration study

When the corpus callosum of the rat is sectioned, the callosal fibres in the cerebral cortex undergo degeneration. In the auditory cortex (area 41) the degenerating axon terminals form asymmetric synapses, and the vast majority of them synapse with dendritic spines. Some other synapse with the shafts of both spiny and smooth dendrites, and a few with the perikarya of non-pyramidal cells. The degenerating axon terminals are contained principally within layer II/III, in which they aggregate in patches. Using a technique in which neurons within the cortex are Golgi-impregnated, then gold-toned and examined in the electron microscope, it has been shown that the dendritic spines of pyramidal neurons with cell bodies in different layers receive the degenerating callosal afferents. The spines arise from the main apical dendritic shafts and their branches, from the dendrites of the apical tufts, and in some cases from the basal dendrites of the pyramidal neurons. The shafts of some pyramidal cell apical dendrites also form asymmetric synapses with callosal afferents. Since we have encountered no spiny non-pyramidal neurons in Golgi preparations of rat auditory cortex, and because other types of non-pyramidal cells have few dendritic spines, it is concluded that practically all of the dendritic spines synapsing with callosal afferents originate from pyramidal neurons.     

Alterations of perisomatic GABA synapses on hippocampal CA1 inhibitory interneurons and pyramidal cells in the kainate model of epilepsy.

In the kainate model of epilepsy, electrophysiological and anatomical modifications occur in inhibitory circuits of the CA1 region of the rat hippocampus. Using postembedding GABA immunocytochemistry and electron microscopy, we characterized perisomatic GABA and non-GABA synaptic contacts in CA pyramidal cells, and GABAergic interneurons of stratum oriens/alveus and stratum lacunosum-moleculare, and examined if changes occurred at these synapses at two weeks post-kainate treatment. We found that, in control rats, the number and total length of perisomatic GABA synapses were significantly smaller (approximately 40-50%) in lacunosum-moleculare interneurons than in oriens/alveus interneurons and pyramidal cells. Additionally, the number and total length of perisomatic non-GABA synapses were different among all cell types, with these parameters increasing significantly in the following order: pyramidal cells相似文献   

Changes in excitatory and inhibitory circuits of the rat hippocampus 12-14 months after complete forebrain ischemia.

Changes in interneuron distribution and excitatory connectivity have been investigated in animals which had survived 12-14 months after complete forebrain ischemia, induced by four-vessel occlusion. Anterograde tracing with Phaseolus vulgaris leucoagglutinin revealed massive Schaffer collateral input even to those regions of the CA1 subfield where hardly any surviving pyramidal cells were found. Boutons of these Schaffer collaterals formed conventional synaptic contacts on dendritic spines and shafts, many of which likely belong to interneurons. Mossy fibres survived the ischemic challenge, however, large mossy terminals showed altered morphology, namely, the number of filopodiae on these terminals decreased significantly. The entorhinal input to the hippocampus did not show any morphological alterations. The distribution of interneurons was investigated by neurochemical markers known to label functionally distinct GABAergic cell populations. In the hilus, spiny interneurons showed a profound decrease in number. This phenomenon was not as obvious in CA3, but the spiny metabotropic glutamate receptor 1alpha-positive non-pyramidal cells, some of which contain calretinin or substance P receptor, disappeared from stratum lucidum of this area. In the CA1 region, somatostatin immunoreactivity disappeared from stratum oriens/lacunosum-moleculare-associated cells, while in metabotropic glutamate receptor 1alpha-stained sections these cells seemed unaffected in number. Other interneurons did not show an obvious decrease in number. In stratum radiatum of the CA1 subfield, some interneuron types had altered morphology: the substance P receptor-positive dendrites lost their characteristic radial orientation, and the metabotropic glutamate receptor 1alpha-expressing cells became extremely spiny. The loss of inhibitory interneurons at the first two stages of the trisynaptic loop coupled with a well-preserved excitatory connectivity among the subfields suggests that hyperexcitability in the surviving dentate gyrus and CA3 may persist even a year after the ischemic impact. The dorsal CA1 region is lost; nevertheless hyperactivity, if it occurs, may have a route to leave the hippocampus via the longitudinally extensive axon collaterals of CA3 pyramidal cells, which may activate the subiculum and entorhinal cortex with a relay in the surviving ventral hippocampal CA1 region.     

Ultrastructure and synaptic connections of vasoactive intestinal polypeptide-like immunoreactive non-pyramidal neurons and axon terminals in the rat hippocampus

In the hippocampus, antibody raised against vasoactive intestinal polypeptide (VIP) labeled perikarya and processes of non-pyramidal neurons whereas these structures remained unlabeled in pyramidal cells and granule cells. In the present study, VIP-immunostaining was used to investigate the fine structure and synaptic connections of identified non-pyramidal neurons and of imrnunoreactive axon terminals in the CA1 region of the rat hippocampus by means of electron microscopic immunocytochemistry.From a number of cells studied, two VIP-like imrnunoreactive non-pyramidal neurons in the regio superior were selected for an electron microscopic analysis of serial thin sections. These cells were different with regard to the location of their cell bodies and the orientation of their dendrites. One cell was located in the stratum lacunosum-moleculare with dendritic processes oriented parallel to the hippocampal fissure. The second neuron was found in the inner one-third of the stratum radiatum. The dendrites of this cell ran nearly parallel to the ascending apical dendrites of the pyramidal cells. Both cells had a round or ovoid perikaryon and an infolded nucleus. The aspinous dendrites of both neurons were densely covered with synaptic boutons. These terminals were small, filled with spherical vesicles and established asymmetric synaptic contacts. No variations in the fine structure of the presynaptic boutons were found along the course of the labeled dendrites through the various hippocampal layers, although different afferents are known to terminate in these layers.Vasoactive intestinal polypeptide-like immunopositive axon terminals course through all layers of the hippocampus. In the stratum pyramidale they established symmetric synaptic contacts with the perikarya of pyramidal cells. In the stratum radiatum they made symmetric contacts with the shafts of apical dendrites of pyramidal cells but never contacted dendritic spines.The symmetric contacts with pyramidal cell perikarya suggest an involvement of the VIP-like immunoreactive axon terminals in pyramidal cell inhibition.     

Distribution of synapses on an intracellularly labeled small pyramidal neuron in the cat motor cortex

Summary The morphological characteristics and distribution of synapses on a small pyramidal neuron in layer III of the cat motor cortex have been studied by combining intracellular HRP staining and electron microscopic examination. The stained neuron showed spiny apical and basal dendritic profiles under the light microscope, and exhibited the morphological features of a pyramidal neuron. Ultrastructural analysis indicated that about 80% of the presynaptic terminals formed asymmetrical synapses with spines of distal apical and basal dendrites. On proximal apical dendrites, 64% of the synapses were found to make contact with spines, and 16.7% of the synapses were of symmetrical type and formed with dendritic shafts. Two types of terminal could be identified on the soma; they were alternately located and established symmetrical and asymmetrical synaptic contacts respectively. Possible functional implications are discussed.This paper is dedicated to Professor Fred Walberg on the occasion of his 70th birthday.     

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.     

Cholecystokinin-immunopositive basket and Schaffer collateral-associated interneurones target different domains of pyramidal cells in the CA1 area of the rat hippocampus.

Two types of GABAergic interneurone are known to express cholecystokinin-related peptides in the isocortex: basket cells, which preferentially innervate the somata and proximal dendrites of pyramidal cells; and double bouquet cells, which innervate distal dendrites and dendritic spines. In the hippocampus, cholecystokinin immunoreactivity has only been reported in basket cells. However, at least eight distinct GABAergic interneurone types terminate in the dendritic domain of CA1 pyramidal cells, some of them with as yet undetermined neurochemical characteristics. In order to establish whether more than one population of cholecystokinin-expressing interneurone exist in the hippocampus, we have performed whole-cell current clamp recordings from interneurones located in the stratum radiatum of the hippocampal CA1 region of developing rats. Recorded neurones were filled with biocytin to reveal their axonal targets, and were tested for the presence of pro-cholecystokinin immunoreactivity.The results show that two populations of cholecystokinin-immunoreactive interneurones exist in the CA1 area (n=15 positive cells). Cholecystokinin-positive basket cells (53%) preferentially innervate stratum pyramidale and adjacent strata oriens and radiatum. A second population of cholecystokinin-positive cells, previously described as Schaffer collateral-associated interneurones [Vida et al. (1998) J. Physiol. 506, 755-773], have axons that ramify almost exclusively in strata radiatum and oriens, overlapping with the Schaffer collateral/commissural pathway originating from CA3 pyramidal cells. Two of seven of the Schaffer collateral-associated cells were also immunopositive for calbindin. Soma position and orientation in stratum radiatum, the number and orientation of dendrites, and the passive and active membrane properties of the two cell populations are only slightly different. In addition, in stratum radiatum and its border with lacunosum of perfusion-fixed hippocampi, 31.6+/-3.8% (adult) or 26.8+/-2.9% (postnatal day 17-20) of cholecystokinin-positive cells were also immunoreactive for calbindin.Therefore, at least two populations of pro-cholecystokinin-immunopositive interneurones, basket and Schaffer collateral-associated cells, exist in the CA1 area of the hippocampus, and are probably homologous to cholecystokinin-immunopositive basket and double bouquet cells in the isocortex. It is not known if the GABAergic terminals of double bouquet cells are co-aligned with specific glutamatergic inputs. However, in the hippocampal CA1 area, it is clear that the terminals of Schaffer collateral-associated cells are co-stratified with the glutamatergic input from the CA3 area, with as yet unknown functional consequences. The division of the postsynaptic neuronal surface by two classes of GABAergic cell expressing cholecystokinin in both the hippocampus and isocortex provides further evidence for the uniform synaptic organisation of the cerebral cortex.     

Septohippocampal connections to field CA1 of the rat identified with field potential analysis and retrograde labeling by horseradish peroxidase

Electrical stimulation of the medial septal nucleus produced field potentials in the hippocampal CA1 region of the rat. The laminar field-potential analysis suggested that the electromotive force of the septum-induced responses might be attributable mainly to excitatory postsynaptic potential currents generated in the stratum oriens (layer of distribution of basal dendrites of hippocampal pyramidal cells). Neural cell bodies in the medial septum-diagonal band complex were retrogradely labeled with horseradish peroxidase (HRP) injected into the stratum oriens of CA1, but not with HRP injected into other CA1 strata. Thus the medial septal nucleus was indicated to send excitatory inputs to basal dendrites of CA1 pyramidal cells.     

Synaptic reorganization in the rabbit hippocampus after lesion of commissural afferents

Summary The degeneration of commissural afferents to the hippocampus in the rabbit was studied by using the Fink-Heimer degeneration method, electron microscopy, and the combined Golgi/EM technique. The stratum oriens (CA3) was selected for quantitative electron microscopic evaluation of postlesional changes since the degeneration of commissural fibers as seen in Fink-Heimer preparations was dense throughout the width of that layer. Accordingly, in electron micrographs of stratum oriens many electron-dense degenerating boutons were found after short survival times (3 and 6 days, respectively), most of them (96%) in synaptic contact with dendritic spines. In the fine structural analysis of Golgi-impregnated CA3 pyramidal cells, spines of basal dendrites were identified as postsynaptic elements of degenerating commissural afferents in stratum oriens.Three days after the lesion, the number of intact synapses/unit area was reduced in stratum oriens of CA3 to 64% of the control; 20% of the synapses were degenerating. Thus, part of the degenerated synapses had disappeared. Evidence is provided that phagocytosis of degenerated boutons still attached to fragments of dendritic spines played a role in this process.Seven weeks after the lesion, the number of intact synapses had returned to control level, suggesting reactive growth of synaptic structures. When the ratio of spine synapses versus shaft synapses was compared with controls, no change had occurred. Thus, after an initial loss of spine synapses after short survival times, new spines have been formed in parallel with ingrowth (sprouting) of neighbouring nonlesioned afferents.     

Development of synapses on pyramidal and multipolar non-pyramidal neurons in the visual cortex of rabbits. A combined Golgi-electron microscope study

A combined Golgi-electron microscope method was used to study the ultrastructural maturation of synapses on identified pyramidal and multipolar non-pyramidal neurons in the visual cortex of young and adult rabbits. In samples of 10 (time of eye opening), 14, 20 day old and 7 month old animals, fully impregnated pyramidal neurons within the layers II-V and multipolar non-pyramidal neurons mainly located in lower layer III and layer IV was studied. We found that synapses in 10 and 14 day old animals were occasionally immature in appearance. They were characterized by either a poorly defined postsynaptic band or equal rims of pre- and postsynaptic electron-dense material and could therefore not be classified as Gray type I or II. The distinction between both types of synapses was easier at day 20 and in the adults when the postsynaptic band of the asymmetrical (type I) synapses had become remarkably thicker. In pyramidal neurons the cytoplasmic organelles increased in number during development. Although a few symmetrical synapses were present on dendritic spines of pyramidal neurons in 14 and 20 day old animals, all pyramidal neurons exhibited the same types of synapses on specific sites of their neuronal surface. They received exclusively type II synapses on their somata, type I synapses on their dendritic spines and both types of synapses on their dendritic shafts. However, in the adult animals the frequency of occurrence of type II synapses, especially on basal dendritic shafts, had increased. In some cases only type II and no type I synapses were present. A striking finding in all young and adult animals was that synapses at the borderline between somata and apical dendritic shafts as well as on dendritic spines were frequently complex or interrupted. The characteristic ultrastructural features of adult spine-free and sparsely spiny multipolar non-pyramidal neurons e.g. the many cytoplasmic organelles and type I and II synapses on somata and on dendrites were already present at day 10. After day 10 the number of organelles and synapses increased prominently and in adult animals the different types of synapses on dendrites were located at relatively short intervals of about 4 microns. In contrast with the dendritic shafts of pyramidal neurons many asymmetrical synapses were observed on dendritic shafts of the non-pyramidal neurons analysed in the adult animals. Furthermore, it appeared that the number of synapses on these non-pyramidal neurons is about twice that on pyramidal neurons in day 20 old animals and about four times in adult animals.(ABSTRACT TRUNCATED AT 400 WORDS)     

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.     

Postsynaptic targets of somatostatin-immunoreactive interneurons in the rat hippocampus

Two characteristic interneuron types in the hippocampus, the so-called hilar perforant path-associated cells in the dentate gyrus and stratum oriens/lacunosum-moleculare neurons in the CA3 and CA1 regions, were suggested to be involved in feedback circuits. In the present study, interneurons identical to these cell populations were visualized by somatostatin-immunostaining, then reconstructed, and processed for double-immunostaining and electron microscopy to establish their postsynaptic target selectivity. A combination of somatostatin-immunostaining with immunostaining for GABA or other interneuron markers revealed a quasi-random termination pattern. The vast majority of postsynaptic targets were GABA-negative dendritic shafts and spines of principal cells (76%), whereas other target elements contained GABA (8%). All of the examined neurochemically defined interneuron types (parvalbumin-, calretinin-, vasoactive intestinal polypeptide-, cholecystokinin-, substance P receptor-immunoreactive neurons) received innervation from somatostatin-positive boutons. Recent anatomical and electrophysiological data showed that the main excitatory inputs of somatostatin-positive interneurons originate from local principal cells. The present data revealed a massive GABAergic innervation of distal dendrites of local principal cells by these feedback driven neurons, which are proposed to control the efficacy and plasticity of entorhinal synaptic input as a function of local principal cell activity and synchrony.     

AMPA receptor modulators have different impact on hippocampal pyramidal cells and interneurons

Positive modulators of AMPA receptors enhance synaptic plasticity and memory encoding. Facilitation of AMPA receptor currents not only results in enhanced activation of excitatory neurons but also increases the activity of inhibitory interneurons by up-modulating their excitatory input. However, little is known about the effects of these modulators on cells other than pyramidal neurons and about their impact on local microcircuits. This study examined the effects of members from three subfamilies of modulators (mainly CX516, CX546 and cyclothiazide) on excitatory synaptic responses in four classes of hippocampal CA1 neurons and on excitatory and disynaptically induced inhibitory field potentials in hippocampal slices. Effects on excitatory postsynaptic currents (EPSCs) were examined in pyramidal cells, in two types of inhibitory interneurons located in stratum radiatum and oriens, and in stratum radiatum giant cells, a novel type of excitatory neuron. With CX516, increases in EPSC amplitude in pyramidal cells were two to three times larger than in interneurons and six times larger than in radiatum giant cells. The effects of CX546 on response duration similarly were largest in pyramidal cells. However, this drug also strongly differentiated between stratum oriens and radiatum interneurons with increases being four times larger in the latter. In contrast, cyclothiazide had similar effects on response duration in all cell types. In field recordings, CX516 was several times more potent in enhancing excitatory postsynaptic potentials (EPSPs) than feedback or feedforward circuits, as expected from its larger influence on pyramidal cells. In contrast, BDP-20, a CX546 analog, was more potent in enhancing feedforward inhibition than either EPSPs or feedback inhibition. This preference for feedforward over feedback circuits is probably related to its higher potency in stratum radiatum versus oriens interneurons. Taken together, AMPA receptor modulators differ substantially in their potency and/or efficacy across major classes of neurons which is likely to have consequences with regard to their impact on circuits and behavior.     

Mu-opioid receptors facilitate the propagation of excitatory activity in rat hippocampal area CA1 by disinhibition of all anatomical layers

Hippocampal mu-opioid receptors (MORs) have been implicated in memory formation associated with opiate drug abuse. MORs modulate hippocampal synaptic plasticity acutely, when chronically activated, and during drug withdrawal. At the network level, MORs increase excitability in area CA1 by disinhibiting pyramidal cells. The precise inhibitory interneuron subtypes affected by MOR activation are unknown; however, not all subtypes are inhibited, and specific interneuron subtypes have been shown to preferentially express MORs. Here we investigate, using voltage-sensitive dye imaging in brain slices, the effect of MOR activation on the patterns of inhibition and on the propagation of excitatory activity in rat hippocampal CA1. MOR activation augments excitatory activity evoked by stimulating inputs in stratum oriens [i.e., Schaffer collateral and commissural pathway (SCC) and antidromic], stratum radiatum (i.e., SCC), and stratum lacunosum-moleculare (SLM; i.e., perforant path and thalamus). The augmented excitatory activity is further facilitated as it propagates through the CA1 network. This was observed as a proportionately larger increase in amplitudes of excitatory activity at sites distal from where the activity was evoked. This facilitation was observed for excitatory activity propagating from all three stimulation sites. The augmentation and facilitation were prevented by GABAA receptor antagonists (bicuculline, 30 microM), but not by GABAB receptor antagonists (CGP 55845, 10 microM). Furthermore, MOR activation inhibited IPSPs in all layers of area CA1. These findings suggest that MOR-induced suppression of GABA release onto GABAA receptors augments all inputs to CA1 pyramidal cells and facilitates the propagation of excitatory activity through the network of area CA1.     

Density and morphology of dendritic spines in mouse neocortex

Dendritic spines of pyramidal cells are the main postsynaptic targets of cortical excitatory synapses and as such, they are fundamental both in neuronal plasticity and for the integration of excitatory inputs to pyramidal neurons. There is significant variation in the number and density of dendritic spines among pyramidal cells located in different cortical areas and species, especially in primates. This variation is believed to contribute to functional differences reported among cortical areas. In this study, we analyzed the density of dendritic spines in the motor, somatosensory and visuo-temporal regions of the mouse cerebral cortex. Over 17,000 individual spines on the basal dendrites of layer III pyramidal neurons were drawn and their morphologies compared among these cortical regions. In contrast to previous observations in primates, there was no significant difference in the density of spines along the dendrites of neurons in the mouse. However, systematic differences in spine dimensions (spine head size and spine neck length) were detected, whereby the largest spines were found in the motor region, followed by those in the somatosensory region and those in visuo-temporal region.