Selective degeneration of CA1 pyramidal cells by chronic application of bismuth

The heavy metal bismuth induces a new type of selective neuronal degeneration that shares some common aspects with that seen following hypoxia and ischemia. Continuous application of 3 μm bismuth to organotypic cultures of rat hippocampus resulted after 2–3 weeks in selective degeneration of CA1 pyramidal cells, while CA3 pyramidal cells, dentate granule cells, and subicular neurons were resistant. With 10 μm MK-801, a noncompetitive NMDA-antagonist, during the entire culturing period failed to prevent neuronal degeneration induced by 3 μm bismuth. GABA-immunoreactive interneurons were also affected by bismuth, but were generally less sensitive than CA1 pyramidal cells. Acute application of up to 100 μm bismuth did not change the electrophysiological properties of CA1 pyramidal cells. © 1994 Wiley-Liss, Inc.     

Non-phosphorylated neurofilament protein immunoreactivity in adult and developing rat hippocampus: specificity and application in grafting studies

Neurofilament proteins are critical to the development and maintenance of neuronal shape in the nervous system. These proteins are developmentally regulated and several transition forms are expressed, prior to full neuronal stabilization. We have studied the spatial distribution and time course of expression of non-phosphorylated neurofilament protein (NPNFP) immunoreactivity in several preparations of rat hippocampus, using a mixture (SMI 311) of several monoclonal antibodies directed against NPNFP epitopes. Differential staining was observed in young and adult hippocampus. Large pyramidal neurons in CA3 and CA4 subfields were strongly immunoreactive in adult hippocampus whereas the smaller CA1 pyramidal neurons, most interneurons and dentate granule cells were immunonegative. SMI 311 staining initially appeared at postnatal day (P) 5 with positive staining in apical dendrites and soma in a few pyramidal neurons in CA3, but almost reached the adult pattern by P10. Compared to adult hippocampus, the number of immunoreactive interneurons in all subfields appeared increased at P10 and P15. In cultures of embryonic hippocampus, all neurons, regardless of their morphology, were SMI 311 positive, suggesting loss of differential expression in tissue culture conditions. However, SMI 311 expression in fetal hippocampal neurons grafted to adult hippocampus was similar to hippocampal neurons which had developed in situ. These results suggest that SMI 311 antibody identifies a distinct group of primarily CA3 and CA4 pyramidal cells in adult hippocampus. The application of SMI 311 immunostaining appears suitable for identification of large CA3 and CA4 pyramidal neurons within hippocampal transplants grafted to adult CNS but not in tissue culture.     

Cellular and connective organization of slice cultures of the rat hippocampus and fascia dentata

This study examined the cellular and connective organization of hippocampal tissue taken from 6-8-day-old rats and cultured by the roller tube technique for 3-6 weeks. In the cultures containing the fascia dentata and the hippocampus proper (CA1, CA3, CA4) the main cell and neuropil layers were organotypically organized when observed in ordinary cell stains. The normal distribution of smaller cell populations of AChE-positive neurons and somatostatin-reactive neurons was demonstrated by histochemical and immunohistochemical methods. Both cell types were mainly confined to str. oriens of CA3 and CA1 and the dentate hilus (CA4). Individual dentate granule cells and hippocampal pyramidal cells were injected with lucifer yellow and HRP, revealing great stability of the dendritic patterns of these cells in the culture condition. The same was found for the axonal branching and termination of HRP-filled mossy fibers arising from an HRP-injected granule cell. The preservation of organotypic afferent patterns in the cultures was also shown by Timm staining of the terminal distribution of the mossy fiber system. Mossy fiber terminals, with characteristic ultrastructural features verified in the electron microscope, were thus found in the hilus (CA4) and along the CA3 pyramidal cell layer onto the CA3-CA1 transition. Depending on the amount of dentate tissue relative to CA3 the terminals could stop before reaching CA1 (small fascia dentata) or take up additional intra and infrapyramidal locations along CA3 (small CA3). In cultures with a gap in the CA3 pyramidal cell layer some mossy fiber terminals were found in contact with the CA3 pyramidal cells beyond the gap. In all cultures there was an aberrant projection of supragranular mossy fibers. This projection is analogous to the one known from lesion and transplant studies to form in the absence of the entorhinal perforant path input to the dentate molecular layer. Also, in accordance with these studies the Timm staining pattern of the outer parts of the dentate molecular layer and the entire molecular layer of the hippocampus was altered corresponding to the spread of afferents normally confined to the inner zone of the dentate and str. radiatum of CA3 and CA1. Possibly as a consequence of the lack of normal targets for projections from CA1, this subfield contained an unusually dense Timm staining suggestive of autoinnervation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Complement C1q expression induced by Abeta in rat hippocampal organotypic slice cultures

Amyloid beta peptide (Abeta) is a major component of senile plaques, one of the principle pathological features in Alzheimer's disease (AD) brains. Fibrillar Abeta has been shown to bind C1 via C1q, the recognition component of the classical complement pathway, resulting in the activation of the complement pathway, thereby initiating an inflammatory cascade in the brain. C1q has also been shown to enhance phagocytic activities of microglia, which could benefit in clearance of apoptotic cells or cellular debris. To begin to define the role of C1q in tissue injury mediated by Abeta, we assessed the appearance of C1q in hippocampal slice cultures treated with freshly solubilized or fibrillar Abeta 1-42. Here we demonstrate a dose- and time-dependent uptake of exogenously applied Abeta by pyramidal neurons in organotypic slice cultures from rat hippocampus. Importantly, when slices were immunostained with antibody against rat C1q, a distinct reactivity for C1q in cells within the neuronal cell layer of cornu ammonis (CA) of hippocampus, primarily the CA1/CA2, was observed in the Abeta-treated slices. No such immunoreactivity was detected in untreated cultures or upon addition of control peptides. ELISA assays also showed an increase in C1q in tissue extracts from slices of the treated group. Similarly, the mRNA level of C1q in slices was increased within 24 h after Abeta treatment. These data demonstrate that upon exposure to Abeta, C1q is expressed in neurons in this organotypic system. The induction of C1q may be an early, perhaps beneficial, tissue or cellular response to injury triggered by particular pathogenic stimuli.     

Development and selective neurodegeneration in cell cultures from different hippocampal regions

Previous studies have shown that pyramidal neurons in hippocampal regions CA1 and CA3 are selectively vulnerable in several neurodegenerative disorders and that a subpopulation of pyramidal neurons in cell cultures of embryonic hippocampus are sensitive to glutamate neurotoxicity. In order to determine whether the patterns of cell loss seen in situ correlate with intrinsic differences in neuronal sensitivities to glutamate-induced degeneration acquired during development, we characterized cultures established from different regions of postnatal rat hippocampus and then examined neuronal sensitivity to glutamate. Tissue corresponding to the dentate gyrus (DG) and regions CA1, CA2 and CA3 of Ammon's horn was removed by microdissection from transverse hippocampal slices and was used to establish cultures of dissociated cells. Cultures from all 4 regions contained 3 major morphological classes of neurons; pyramidal-like, bipolar and stellate. Pyramidal-like neurons comprised the majority of neurons in all cultures; these neurons extended one long and branching axon, and one or more short dendrites. Immunocytochemistry showed that all neurons possessed high levels of glutamate-like and gamma-aminobutyric acid (GABA)-like immunoreactivity when grown in isolation. In contrast, when bipolar and pyramidal neurons were cultured in contact with glial cells, glutamate and GABA immunoreactivity were selectively reduced in the bipolar and pyramidal cells, respectively, suggesting that cell interactions influence neurotransmitter phenotype. Subpopulations of hippocampal neurons from each hippocampal region were vulnerable to glutamate-induced neurotoxicity. Bipolar and stellate cells were resistant to glutamate, while pyramidal-like neurons showed varying degrees of sensitivity to glutamate depending upon which region they were taken from. Experiments with specific glutamate receptor agonists and antagonists demonstrated that both non N-methyl-D-aspartic acid (NMDA) receptors and NMDA receptors mediated glutamate-induced degeneration. There were clear differences in the vulnerability of the pyramidal-like neuron populations in cultures from the different hippocampal regions. The rank order of the vulnerability of pyramidal-like neurons to glutamate-induced neurodegeneration between regions in culture was: DG less than CA2 less than CA3 less than CA1. This pattern of selective vulnerability in cell culture corresponds directly to the pattern of selective cell loss seen in situ in Alzheimer's disease, epilepsy, and stroke suggesting that intrinsic neuronal differences in glutamate sensitivity may be involved in these disorders.     

Calbindin-D28K and ischemic damage of pyramidal cells in rat hippocampus.

An antibody against rat calbindin-D28K, a calcium-binding protein present at high concentration in certain neurons of the central and peripheral nervous systems, was used to determine the progression of the pathological events in the rat hippocampus following experimental cerebral ischemia. Calbindin-D28K immunoreactivity is present in dentate granule cells and in the CA1-CA2 pyramidal cells. CA1 subfield contains a higher proportion of calbindin-D28K-positive pyramidal cells than does the CA2 subfield and CA1 cells are more immunoreactive than the CA2 cells. The pyramidal cells of the CA1 and CA2 subfields are vulnerable to ischemia. The cells in the CA1 became necrotic within 3-4 days after ischemia while those of the CA2 became necrotic within 2 days. There was a concomitant decrease in calbindin-D28K immunoreactivity in the whole hippocampal regio superior after ischemia which peaked 3 days postischemia. The difference in CA2 and CA1 vulnerability seemed to be inversely correlated with the calbindin-D28K contents of the CA2 and CA1 pyramidal cells. The decrease in the calbindin-D28K contents of these neurons was accompanied by cell damage. We therefore suggest that calbindin-D28K is an important factor for the survival of pyramidal cells in the hippocampal formation after ischemia.     

Local circuit synaptic interactions between CA1 pyramidal cells and interneurons in the kainate-lesioned hyperexcitable hippocampus.

Following kainate (KA)-induced lesions of subfield CA3--a lesion relevant to human temporal lobe epilepsy--remaining pyramidal cells in CA1 display synchronous hyperexcitability associated with a loss of synaptic inhibition. Despite this loss, inhibitory interneurons in CA1 remain viable, and the density and function of GABAergic receptors on the CA1 pyramidal cells are maintained at approximately normal levels. To further evaluate inhibition in this system, the authors examined interactions between pyramidal cells and inhibitory interneurons in paired intracellular recordings. Recordings were carried out in rat hippocampal slices 2-4 weeks following bilateral intraventricular KA injections. The frequency of synaptic interactions between CA1 basket cells and pyramidal cells was lower in hyperexcitable slices than in controls; both synapses in the recurrent inhibitory circuit appeared to be involved. No recurrent excitatory interactions were seen between pyramidal cell pairs in lesioned or normal slices. The weakened interconnections between pyramidal cells and interneurons are consistent with the decreased inhibition previously found in this model. Unexpectedly, strong stimulation, which may directly activate local inhibitory circuitry, was effective in reducing hyperexcitability in KA-lesioned slices. These data suggest that development of recurrent excitatory connections among CA1 hippocampal pyramidal cells contribute little to tissue excitability, and support the hypothesis that a functional uncoupling between inhibitory interneurons and CA1 pyramidal cells is responsible for the seizure-like activity typical of KA-lesioned hippocampus. The data are also consistent with the hypothesis that in the KA model, the structural circuitry needed for inhibition in CA1 is maintained, and can be functionally activated by appropriate stimuli.     

Effects of GABA and baclofen on pyramidal cells in the developing rabbit hippocampus: an 'in vitro' study

Using the in vitro hippocampal slice preparation, we have investigated the effects of gamma-aminobutyric acid (GABA) and its analogue beta-(p-chlorophenyl)-GABA (baclofen) on CA1 and CA3 pyramidal cells in the developing rabbit hippocampus. Somatic applications: both GABA and baclofen, when applied to CA1 pyramidal cells from immature tissue, led to cell depolarization from resting membrane potential; this baclofen depolarization may be indirectly mediated. In contrast, CA3 pyramidal cells at the same age were primarily hyperpolarized by both drugs. In mature tissue, both GABA and baclofen applied at the soma induce cell hyperpolarizations. Dendritic applications: immature CA1 cells responded to dendritic GABA and baclofen application with depolarizations associated with increased cell excitability; here, too, the baclofen depolarization may be due to indirect 'disinhibition'. Both depolarizing and hyperpolarizing responses were recorded in immature tissue when GABA was applied to CA3 pyramidal cell dendrites: baclofen produced only hyperpolarizations. In mature CA1 cells, dendritic GABA application produced membrane depolarization, but dendritic baclofen application produced hyperpolarizations. In mature CA3 cells, dendritic GABA and baclofen application produced predominant hyperpolarizations. Mature CA1 pyramidal cells appear to retain some of the GABA-induced depolarizations characteristic of immature tissue. In contrast, mature CA3 neurons show only hyperpolarizing responses to GABA and baclofen application. In all cases, responses to GABA and baclofen are associated with a decrease in cell input resistance. We conclude that the GABAergic receptor/channel complexes mature differently in the CA1 and CA3 regions of the hippocampus.     

Autoradiographic demonstration of glutamate structures after stereotaxic injection of kainic acid in rat hippocampus

Autoradiography of high affinity uptake ofl-glutamate in rat hippocampus after kainic acid lesion of CA3 and CA4 pyramidal cells has been used to identify the neurotransmitter of their ipsilateral connections. Glutamate or aspartate has tentatively been identified as the neurotransmitter of the projection from CA3 pyramidal cells to strata radiatum and oriens of CA1 and the projection from CA4 pyramidal cells to the inner parts of the molecular layer of area dentata.     

The comparative immunocytochemical distribution of 28 kDa cholecalcin (CaBP) in the hippocampus of rat, guinea pig and hedgehog

The distribution of 28 kDa cholecalcin (calcium-binding protein, CaBP) in the hippocampal formation of the rat, guinea pig and European hedgehog was examined by immunocytochemistry. The extension of the mossy fibers (the axons of the granule cells of the dentate gyrus) was also studied using the Timm's sulfide-silver method. Cholecalcin was present in all mossy fibers. In the rat, only those pyramidal cells not reached by the labeled mossy fibers displayed cholecalcin immunoreactivity. Immunocytochemical staining of the hedgehog hippocampus showed that contacts between cholecalcin-containing mossy fibers and cholecalcin-containing pyramidal cells are possible. Consequently, the protein is probably not involved in the control of mossy fiber extension. Strikingly, no guinea pig pyramidal cells showed cholecalcin immunoreactivity. The possible involvement of cholecalcin in the differential excitability of pyramidal cells in the CA3 and CA1 areas of the hippocampus could therefore be tested in a comparative study of rat, guinea pig and hedgehog.     

Selective kainic acid lesions in cultured explants of rat hippocampus

Summary The influence of the excitotoxin kainic acid (KA) on cultivated explants of rat hippocampus was investigated. Addition of 3 M KA to the culture medium over 24–48 h induced a destruction of the pyramidal cells in the CA3 region, whereas the CA1 pyramidal cells and the granule cells were left undamaged. Higher concentrations (10–100 M) of KA destroyed also the latter cell groups. The selectivity of the KA lesion at 3 M was further indicated by the fact that the acetylcholinesterase-positive neurons in the hippocampus were not destroyed through KA administration and that the stereoisomer dihydrokainic acid was ineffective in inducing lesions. Application of tetrodotoxin did not protect the CA3 pyramidal cells from KA lesion, whereas -glutamylaminomethylsulphonic acid (GAMS) only offered a very small, statistically not significant, protection. Baclofen protected the cultures slightly from KA lesions but not when added together with GAMS. Possible mechanisms responsible for the KA lesions in these cultures are discussed.Supported in part by a grant from the Swiss National Foundation for Scientific Research (No. 3.528.-0.83)     

Restoration of calbindin after fetal hippocampal CA3 cell grafting into the injured hippocampus in a rat model of temporal lobe epilepsy

Degeneration of the CA3 pyramidal and dentate hilar neurons in the adult rat hippocampus after an intracerebroventricular kainic acid (KA) administration, a model of temporal lobe epilepsy, leads to permanent loss of the calcium binding protein calbindin in major fractions of dentate granule cells and CA1 pyramidal neurons. We hypothesize that the enduring loss of calbindin in the dentate gyrus and the CA1 subfield after CA3-lesion is due to disruption of the hippocampal circuitry leading to hyperexcitability in these regions; therefore, specific cell grafts that are capable of both reconstructing the disrupted circuitry and suppressing hyperexcitability in the injured hippocampus can restore calbindin. We compared the effects of fetal CA3 or CA1 cell grafting into the injured CA3 region of adult rats at 45 days after KA-induced injury on the hippocampal calbindin. The calbindin immunoreactivity in the dentate granule cells and the CA1 pyramidal neurons of grafted animals was evaluated at 6 months after injury (i.e. at 4.5 months post-grafting). Compared with the intact hippocampus, the calbindin in "lesion-only" hippocampus was dramatically reduced at 6 months post-lesion. However, calbindin expression was restored in the lesioned hippocampus receiving CA3 cell grafts. In contrast, in the lesioned hippocampus receiving CA1 cell grafts, calbindin expression remained less than the intact hippocampus. Thus, specific cell grafting restores the injury-induced loss of calbindin in the adult hippocampus, likely via restitution of the disrupted circuitry. Since loss of calbindin after hippocampal injury is linked to hyperexcitability, re-expression of calbindin in both dentate gyrus and CA1 subfield following CA3 cell grafting may suggest that specific cell grafting is efficacious for ameliorating injury-induced hyperexcitability in the adult hippocampus. However, electrophysiological studies of KA-lesioned hippocampus receiving CA3 cell grafts are required in future to validate this possibility.     

Disposition of the slab-like modules formed by axon branches originating from single CA1 pyramidal neurons in the rat hippocampus

The hippocampus is thought to be an area where the neuronal circuits for short-term memory or the cognitive map may reside. In order to advance theoretical studies and neuronal model simulations of such circuits, the projection of the CA1 pyramidal neurons in the rat dorsal hippocampus, especially in the subiculum, was studied by means of intracellular and extracellular HRP injection. The CA1 pyramidal neurons project principally to the subiculum where each forms a slab-like axonal field 2 mm long along the septotemporal axis, which may be regarded as a module for columnar organization, at a specific rostrocaudal level of the subiculum. The modules of the CA1a pyramidal neurons are disposed in the rostral part of the subiculum, those of the CA1c pyramidal neurons in the caudal part, and those of the CA1b pyramidal neurons in the middle part of the subiculum. The CA1 pyramidal neurons also participate in the construction of the lamellar organization in the hippocampus in that their axon branches run rostrocaudally following the stream of the alvear fibers. The CA1 pyramidal neurons in the dorsal rat hippocampus transfer the topographic map from field CA1 to the subiculum with reversed order in the lamellar direction. The topographical relationship is composed of partially shifted, overlapping slab-like modules. As a result, information conveyed through a lamella will diverge into the subiculum approximately 2 mm wide, and information through a group of lamellae 2 mm wide will converge upon single subicular neurons.     

Temporal profiles of the in vitro phosphorylation rate and immunocontent of glial fibrillary acidic protein (GFAP) after kainic acid-induced lesions in area CA1 of the rat hippocampus: demonstration of a novel phosphoprotein associated with gliosis

The in vitro phosphorylation rate and immunocontent of glial fibrillary acidic protein was studied in slices of area CA1 of the rat hippocampus after stereotaxic injection of 1 nmol of kainic acid. For controls the contralateral hippocampus was injected with saline. Hippocampal tissue was incubated with [³²P]phosphate and analysed by two-dimensional electrophoresis for phosphorylation rate and by immunoblotting for immunocontent. Both these parameters decreased during the first 4 days after injection and then started to increase at 10 days and continued to increase until at least 84 days. Except for a small excess of phosphorylation rate at 28 days, the relationship between immunocontent and in vitro phosphorylation rate of glial fibrillary acidic protein remained constant, indicating that the reactive gliosis was not associated with hypo- or a major hyperphosphorylation of this protein. Histology showed a pronounced loss of CA1 pyramidal cells 1 day after injection. At 28 days after injection the pyramidal cells had disappeared and only a few abnormal neurones were present. In contrast, immunocytochemistry after 28 days showed a marked increase in astrocytes reacting positive to the antibody in the strata radiatum and lacunosum moleculare. Besides glial fibrillary acidic protein the expression of several other proteins was upregulated as a result of the injection of kainic acid. These included phosphovimentin and an unknown phosphoprotein designated pp25 which co-migrated on 2-D gels with a prominent phosphoprotein expressed in primary cultures of astrocytes. Pp25 was expressed in lesioned tissue more frequently than phosphovimentin and with a time course that started earlier. Of particular interest was the expression of pp25 in the contralateral saline-injected hippocampus 1 day after injection of kainic acid. It is possible that pp25 will prove to be a sensitive marker of gliosis.     

The expression of somatostatin receptors in the hippocampus of pilocarpine-induced rat epilepsy model

During the course of this study, we sought examine whether the expression of somatostatin receptors (SSTRs) is altered in the hippocampus following pilocarpine-induced status epilepticus (SE) in order to understand the role/function of SSTRs in the hippocampus after epileptogenic insults. SSTR1 and SSTR4 immunoreactivities were increased in the hippocampus at 1 week after SE. At 4 weeks after SE, SRIF1-family (SSTR 2A, SSTR2B, and SSTR5) immunoreactivity was increased only in neuropil. Both SSTR2A and 2B immunoreactivities were increased in CA2-3 pyramidal cells. However, SSTR3 and SSTR4 immunoreactivities were reduced in the CA1 pyramidal cells of epileptic rat due to neuronal loss. In addition, SSTR5 immunoreactivity was reduced in CA2 pyramidal cells and various interneurons. Both SSTR2B and SSTR4 immunoreactivities were increased within microglia following SE. Our findings suggest that increases in neuron-glial SSTR expressions may be closely related to the enhanced inhibition of the dentate gyrus and regulation of reactive microgliosis in the hippocampus of a pilocarpine model of temporal lobe epilepsy.     

Spatial and temporal patterns of morphogenesis of hippocampal pyramidal cells: study in the early postnatal rat

Several studies have dealt with the morphogenesis of the rat hippocampal pyramidal cell, but little is known about how the different pyramidal cell shapes of CA fields differentiate from neuroepithelial cells, or about how the field morphological identity emerges. From our studies of pyramidal cell shapes in the CA1, CA3, and CA4 fields of hippocampi at postnatal developmental stages between P0 and P12, using fresh semidissociated slices and acutely dissociated cells, we identified the sequence of cell shape transformation by which they differentiate from simple bipolar to complex shapes characteristic of adult pyramidal cells of CA1 and CA3. Pyramidal cell morphogenesis does not occur synchronously throughout the CA hippocampus fields, but cells in the CA4 field undergo morphological differentiation at earlier stages, prenatally, than CA3 cells, and these in turn earlier than CA1 cells. Thus, during the P1-P6 stages, a gradient of shapes from less to greater differentiation was clearly observed from CA1 to CA4 in a single slice. Furthermore, a mixture of cells at different degrees of differentiation is observed in CA1 from P1 to P10, and in CA3 from P1 to P5. A gradient of shapes from more to less differentiation was observed at stages P5-P6 from septal to temporal. We describe two processes in the pyramidal cell morphogenesis in the CA1 and CA3 fields, the approximation to the soma of the point of bifurcation of the main apical process, and the acquisition of triangular shape of the soma, showing by a quantitative study of both processes that they occur earlier in CA3 than in CA1. Our study, therefore, provides new insight into rat pyramidal cell morphogenesis, and indicates that this process might be differently regulated in the various CA fields. Hippocampus     

Exposure to excess glucocorticoids alters dendritic morphology of adult hippocampal pyramidal neurons.

We have used Golgi-impregnated tissue to demonstrate that exposure to excess glucocorticoids alters dendritic morphology in a specific population of neurons in the adult rat hippocampus. Daily injection of 10 mg of corticosterone for 21 days resulted in decreased numbers of apical dendritic branch points and decreased total apical dendritic length measured in a 100-microns-thick section in CA3 pyramidal cells compared to sham-injected and non-injected controls. In contrast, no changes were observed in CA3 pyramidal cell basal dendritic morphology. Furthermore, no changes were observed in the dendritic morphology of CA1 pyramidal cells or granule cells of the dentate gyrus. Cross-sectional cell body area of any of the 3 cell types examined in this study was unaffected by corticosterone treatment. Finally, qualitative analysis of Nissl-stained tissue from the same brains revealed increased numbers of darkly staining, apparently shrunken CA3 pyramidal cells in corticosterone treated compared to control brains. The changes in dendritic morphology we have observed may be indicative of neurons in the early stages of degeneration, as prolonged exposure to high levels of corticosterone has been shown by others to result in a loss of CA3 pyramidal cells. Additionally, these results suggest possible structural alterations which may occur under physiological conditions in which corticosterone levels are chronically elevated such as in aged animals.     

Ultrastructural description of taurine-like immunoreactive cells and processes in the rat hippocampus

A monoclonal antibody against taurine conjugated to KLH was used to identify and describe taurine-like immunoreactive processes in the rat hippocampus. Tissue from perfused rats was processed for immunohistochemical visualization of taurine and embedded for electron microscopy. Representative tissue samples from three regions, the dentate gyrus, CA3, and CA1, were sectioned, examined, and photographed. In the dentate gyrus, both granule cells and pyramidal basket cells were taurine-like immunoreactive. Some axon terminals in the dentate gyrus molecular layer as well as some mossy fiber boutons in the hilus were also taurine-like immunoreactive. In the CA3 region both pyramidal neurons and glial cells were taurine-like immunoreactive A few small-diameter axon terminals in stratum radiatum and some mossy fiber boutons in stratum lucidum were taurine-like immunoreactive. In CA1, pyramidal neurons and some glia were intensely taurine-like immunoreactive. A few immunoreactive axon terminals were seen in stratum radiatum and stratum oriens. In all regions, dendritic staining predominated. Our results support the hypothesis that while taurine may act as a neurotransmitter in a small portion of hippocampal terminals, its main function is probably as a neuromodulator or ionic regulator.     

The hippocampal CA3 network: An in vivo intracellular labeling study

The intrahippocampal distribution of axon collaterals of individual CA3 pyramidal cells was investigated in the rat. Pyramidal cells in the CA3 region of the hippocampus were physiologically characterized and filled with biocytin in anesthetized animals. Their axonal trees were reconstructed with the aid of a drawing tube. Single CA3 pyramidal cells arborized most extensively in the CA1 region, covering approximately two-thirds of the longitudinal axis of the hippocampus. The total length of axon collaterals in the CA3 region was less than in CA1 and the axon branches tended to cluster in narrow bands (200–800 μm), usually several hundred microns anterior or posterior to the cell body. The majority of the recurrent collaterals of a given neuron remained in the same subfield (CA3a, b, or c) as the parent cell. CA3a neurons innervated predominantly the basal dendrites, whereas neurons located proximal to the hilus (CA3c) terminated predominantly on the apical dendrites of both CA1 and CA3 cells. Two cells, with horizontal dendrites and numerous thorny excrescences at the CA3c–hilus transitional zone, were also labeled and projected to both CA3 and CA1 regions. All CA3 neurons projected some collaterals to the hilar region. Proximal (CA3c) neurons had numerous collaterals in the hilus proper. One CA3c pyramidal cell in the dorsal hippocampus sent an axon collateral to the inner third of the molecular layer. CA3c pyramidal cells in the ventral hippocampus had extensive projections to the inner third of the dentate molecular layer, as well as numerous collaterals in the hilus, CA3, and CA1 areas, and several axon collaterals penetrated the subiculum. The total projected axon length of a single neuron ranged from 150 to 300 mm. On the basis of the projected axon length and bouton density (mean interbouton distance: 4.7 μm), we estimate that a single CA3 pyramidal cell can make synapses with 30,000–60,000 neurons in the ipsilateral hippocampus. The concentrated distribution of the axon collaterals (“patches”) indicates that subpopulations of neurons may receive disproportionately denser innervation, whereas innervation in the rest of the target zones is rather sparse. These observations offer new insights into the physiological organization of the CA3 pyramidal cell network. © 1994 Wiley-Liss, Inc.     

Estradiol induces a phasic Fos response in the hippocampal CA1 and CA3 regions of adult female rats

Previous studies have shown that estradiol induces structural and functional changes in hippocampal CA1 pyramidal cells of the adult female rat. Estradiol increases the density of dendritic spines and axospinous synapses on CA1 pyramidal cells, and increases these cells' sensitivity to NMDA receptor-mediated synaptic input. Curiously, while estradiol effects are observed in CA1 pyramidal cells, the majority of the evidence indicates that these cells lack genomic estradiol receptors. In contrast, genomic estradiol receptors are expressed in at least some hippocampal interneurons in CA1. The goal of the present study was to determine which hippocampal neuronal populations are activated by estradiol, as determined by induction of c-Fos immunoreactivity, as well as the time-course of this activation. We quantified c-Fos expression in each of the major subdivisions of the hippocampus in adult female rats at various time points during the same estradiol treatment regimen known to regulate dendritic spines and synapses on CA1 pyramidal cells. Our results show a phasic estradiol-induced c-Fos response in the pyramidal cell layers of both CA1 and CA3. c-Fos was induced within 2 h of treatment, decreased at 6 and 12 h, and subsequently increased again at 24 h after treatment with estradiol. Double labeling for c-Fos and GAD 65 or GAD 67 suggests that c-Fos is induced primarily in principal cells, though a small proportion of GABAergic cells is also labeled. These estradiol-induced changes in c-Fos expression may reflect phasic neuronal activation and coupling to gene expression, which could be involved in estradiol's effects on excitatory synaptic connectivity in the hippocampus.