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)     

Cells of origin of entorhinal cortical afferents to the hippocampus and fascia dentata of the rat

The pathway from the entorhinal cortical region to the hippocampal formation has previously been shown to be comprised of two sub-systems, one of which projects predominantly to the ipsilateral fascia dentata and regio inferior of the hippocampus proper, and a second which projects bilaterally to regio superior. The goal of the present investigation was to determine if these two pathways might originate from different cell populations within the entorhinal area. The cells of origin of these entorhinal pathways were identified by retrograde labeling with horseradish peroxidase (HRP). Injections which labeled the entorhinal terminal fields in both the fascia dentata and regio superior resulted in the retrograde labeling of two populations of cells in the entorhinal area. Ipsilateral to the injection, HRP reaction product was found in the cells of layer II (predominantly stellate cells) and the cells of layer III (predominantly pyramidal cells). Contralateral to the injections, however, the reaction product was found almost exclusively in the cells of layer III. With selective injections of the entorhinal terminal field in regio superior, only the cells of layer III were labeled, but these were labeled bilaterally. Selective injection of the entorhinal terminal field in the fascia dentata, however, resulted in the labeling of cells of layer II, but not of layer III, and these cells of layer II were labeled almost exclusively ipsilaterally. A very small number of labeled cells in layer II were, however, found contralateral to the injection as well. No labeled cells were found either in the presubiculum or parasubiculum following injections of the hippocampal formation. These cell populations were found capable of retrograde transport of HRP, however, since cells in both presubiculum and parasubiculum were labeled following HRP injections into the contralateral entorhinal area. These results suggest that the projections to the fascia dentata originate from the cells of layer II, while the projections to regio superior originate from the cells of layer III of the entorhinal region proper. The very slight crossed projection from the entorhinal area to the contralateral area dentata probably originates from the small population of cells in layer II which are labeled following HRP injections in the contralateral area dentata.     

Ultrastructural organization of normal and transplanted rat fascia dentata: II. A quantitative analysis of the synaptic organization of intracerebral and intraocular grafts

As part of an ultrastructural analysis of the normal rat fascia dentata and intracerebral and intraocular dentate transplants the synapses in the dentate molecular layer were quantified. Hippocampal and dentate tissue from 21-day-old rat embryos were grafted into the brain of developing and adult rats and to the anterior eye chamber of adult rats. After 100 or 200 days of survival the recipient rat brains and the recipient eyes were processed for electron microscopy, and the graft dentate molecular layer with the adjacent granule cell layer selected for ultrastructural analysis. Tissue from the dentate molecular layer of normal adult rats served as controls. The dentate synapses were classified as asymmetric (Gray's type 1) or symmetric (Gray's type 2), and according to the postsynaptic element (cell body, dendritic shaft, dendritic spine). The spine synapses were further classified into simple and complex types according to the spine-terminal configuration. Also, the length of synaptic contacts of the individual synaptic types was measured in some grafts, just as the percentage of the cross sectional area of the neuropil covered by blood vessels. The results showed that the synaptic density, expressed as number per unit area of neuropil, to a large extent was the same within the different parts of the normal dentate molecular layer. Compared with this the synaptic density was reduced with 16.4% in dentate molecular layer of the intracerebral graft, primarily because of a 17.6% reduction of simple synapses on dendritic spines and almost halving of the symmetric synapses on dendritic shafts. The synaptic density was independent of the age of the recipient, the intracerebral location of the graft, and the survival time. Although the synaptic length of some of the individual synaptic types increased, this did not compensate for the loss of synapses. In the intraocular grafts the synaptic density was lower than in the intracerebral grafts. Despite the reduced synaptic density, which mainly involved two synaptic types, we conclude that grafted dentate granule cells can develop a remarkably normal, ultrastructural synaptic organization even in the absence of major afferent inputs. This outcome must accordingly be achieved by reorganization of the available intrinsic afferents.     

Ultrastructural organization of normal and transplanted rat fascia dentata: I. A qualitative analysis of intracerebral and intraocular grafts

Few studies have dealt with the general ultrastructure and synaptic organization of grafted brain tissue. This study was therefore performed to extend current light microscopic observations on intracerebral and intraocular grafts of hippocampal tissue to the ultrastructural level. Blocks of tissue containing the hippocampus and fascia dentata from day 21 embryonic rats were grafted into the brain of developing and adult rats and to the anterior eye chamber of adult rats. After 100 or 200 days of survival the recipient rat brains or eyes were processed for electron microscopy. Tissue containing the graft dentate molecular layer and adjacent granule cell layer was selected for ultrastructural analysis, together with a few samples of the hilus and CA3. Normal dentate tissue was analyzed as control. At the light microscopic level most intracerebral and intraocular grafts displayed an organotypic organization with clearly recognizable cell and neuropil layers. Under the electron microscope the grafted granule cells had normal-appearing dendrites bearing the normal types of spines and forming the normal types of synapses. This was the case even in the absence of the normal major extrinsic afferents like the perforant path. The graft dentate granule cells formed axons and terminals with characteristics of the normal mossy fiber system in the hilus and CA3, in addition to aberrant supragranular mossy fiber terminals known from light microscopic studies of dentate transplants. Abnormal structures included a few dendritic growth cones and an increased occurrence of polyribosomes in spines. Their occurrence indicates ongoing dendritic plasticity even 100 days after transplantation. There was also an increased density of glial elements, particularly in the intraocular grafts. In some of these grafts the granule cells displayed immature traits in terms of nuclear indentations. Dentate interneurons of the basket cell type were present in both the intracerebral and the intraocular grafts. We conclude that grafted dentate granule cells, in different surroundings and without the normal, major perforant path input, can develop a basically normal cellular morphology, which includes the normal ultrastructural characteristics of the dendrites with spines and synapses, and the mossy fibers and its terminals.     

Neurturin, persephin, and artemin in the human pre- and full-term newborn and adult hippocampus and fascia dentata

The immunochemical occurrence and localization of the Glial cell line-derived neurotrophic factor (GDNF) family ligands neurturin (NTN), persephin (PSP), and artemin (ART) is described in the human postmortem hippocampus and fascia dentata from subjects aged 21 weeks of gestation to 88 years. The detectability of NTN, PSP, and ART is shown in the rat by Western blot and immunohistochemistry up to 70 h postmortem. In the human tissue, labeled neuronal perikarya were detectable for each trophin at all examined ages, with prevalent localization in the pyramidal layer of the Ammon's horn and hilus and granular layer of the fascia dentata. In the adult subjects, punctate elements were also present. Comparison of the pattern of immunoreactive structures among young and adult subjects suggests that intracellular distribution and/or trafficking of the GDNF family ligands may undergo age-related changes. Labeled glial elements were also identifiable. Western blot analysis indicates that the availability of the dimeric and monomeric forms of the trophins may vary with age and postmortem delay. The results obtained suggest the involvement of NTN, PSP, and ART in processes subserving both the organization of this cortical region during development and the functional activity and maintenance of the mature human hippocampal neurons.     

Projection of the lateral part of the entorhinal area to the hippocampus and fascia dentata

The hippocampus and fascia dentata receive their major extrinsic input from the entorhinal area through the so-called perforant path. This pathway is now shown to be composed of at least two distinct fiber systems: (1) A medial perforant path coming from the medial part of the entorhinal area and terminating in the middle of the dentate molecular layer and in the deep half of the stratum lacunosum-moleculare of the hippocampal subfield CA3. (2) A lateral perforant path from the lateral part of the entorhinal area to a superficial zone in the dentate molecular layer and to the superfcial part of the stratum lacunosum-moleculare of CA3. This paper deals specifically with the lateral perforant path. A third group of perforant fibers, bing intermediate to the others with regard to both origin and termination has been noticed in one animal. The fiber-course of the lateral perforant path is found to be identical to that previously described for the medial path. The terminal field is present along the whole axial extent of the hippocampus and fascia dentata, i.e., from the temporal tip to the subsplenial portion. No sings of degeneration corresponding to the so-called alvear path were observed following lesions of either the medial or the lateral part of the entorhinal cortex. Terminal degeneration appeared in the molecular layer of the subiculum and CA1 and in the anterior continuation of the hippocampal formation subsequent to lesions including the prepyriform cortex.     

Mossy cells of the rat fascia dentata are glutamate-immunoreactive

The mossy cells represent a prominent cell type of the hilar region. Whereas the morphology of these neurons, their synaptic connections, and physiological characteristics have been described in some detail, information about their neurotransmitter is still lacking. Using immunocytochemistry in combination with Golgi impregnation, the authors demonstrate that identified mossy cells are GABA-immunonegative but stain for glutamate. These results do not prove that these cells use glutamate as a transmitter, since glutamate is a ubiquitous metabolite. However, together with the lack of GABA staining and a recent report on asymmetric spine synapses formed by identified mossy cell axons, the present results support an excitatory nature of these neurons.     

Susceptibility of immature and adult brains to seizure effects

The extent that status epilepticus (SE), but also brief seizures, affects neuronal structure and function has been the subject of much clinical and experimental research. There is a reliance on findings from animal research because there have been few prospective clinical studies. This review suggests that the features of seizure-induced injury in the immature brain compared with the adult brain are different and that duration of seizures (SE versus brief), number of seizures, cause of seizures, presence of pre-existing abnormalities, and genetics affect the injury. Increased awareness of age-specific injuries from seizure has promoted research to determine the circumstances under which seizures may produce permanent detrimental effects. Together with recent advances in functional neuroimaging, genomic investigation, and prospective human data, these studies are likely to substantially increase our knowledge of seizure-induced injury, leading to the development of improved algorithms for prevention and treatment of epilepsy.     

Cholinergic innervation of mossy cells in the rat fascia dentata

Hilar mossy cells are the main cells of origin of the commissural/associational projection to the inner molecular layer of the rat fascia dentata. In order to analyze the cholinergic innervation of hilar mossy cells, a light and electron microscopic double‐labeling technique was used. Immunolabeling for calcitonin gene‐related peptide (CGRP) was employed to identify mossy cells and immunocytochemistry for choline acetyltransferase (ChAT) was used to label cholinergic septohippocampal fibers. Cholinergic boutons were abundant around mossy cell somata and on their proximal dendrites. Electron microscopy confirmed that many of these boutons formed synapses with the CGRP‐positive mossy cells. These data demonstrate a direct innervation of hilar mossy cells by cholinergic septohippocampal afferents. This connectivity could contribute to the electrophysiological behavior of mossy cells during theta oscillations. Hippocampus 1999;9:314–320. © 1999 Wiley‐Liss, Inc.     

Ret, GFRalpha-1, GFRalpha-2 and GFRalpha-3 receptors in the human hippocampus and fascia dentata

The immunohistochemical occurrence and localization of the receptor components of the glial cell line-derived neurotrophic factor (GDNF) family ligands, the Ret receptor tyrosine kinase and GDNF family receptor (GFR) alpha-1 to -3, is described in the human post-mortem hippocampal formation at pre- and full-term newborn, and adult age. Two different antibodies for each of the four-receptor molecules were used. Western blot analysis indicates that the availability of GFRalpha receptor proteins may vary with age and post-mortem delay. The immunohistochemical detectability of GFRalpha-1, GFRalpha-2, GFRalpha-3 and Ret receptor molecules is shown in the rat up to 72 h post-mortem. In the human specimens, labelled neuronal perikarya were detectable for each receptor protein at all examined ages, with prevalent localization in the pyramidal layer of the Ammon's horn and hilus and granular layer of the fascia dentata. In the adult subjects, abundant punctate-like structures were also present. Labelled glial elements were identifiable. Comparison of the pattern of immunoreactive elements among young and adult subjects suggests that the intracellular distribution of the GDNF family ligands may vary between pre- and perinatal life and adult age. The results obtained suggest the involvement of the Ret and GFRalpha receptors signalling in processes subserving both the organization of this cortical region during development and the functional activity and maintenance of the mature hippocampal neurons.     

Effects of baclofen and bicuculline on inhibition in the fascia dentata and hippocampus regio superior

The effects of microiontophoretically applied baclofen, bicuculline and phaclofen were studied on evoked field responses, paired-pulse (PP) plasticity and single-unit activity of dentate granule cells (DGCs) and CA1 pyramidal cells (PCs) in anesthetized rats. The GABAB agonist, baclofen, increased population spike (PS) amplitudes in the dentate evoked by perforant path stimulation but decreased PS amplitudes in CA1 evoked by Schaffer collateral stimulation, whereas the GABAA antagonist, bicuculline, increased PS amplitudes in both regions. Neither baclofen nor bicuculline had significant effects on dendritically recorded population excitatory postsynaptic potentials (EPSPs) in the dentate or CA1 evoked by stimulation of their respective afferents. Control PP curves in the dentate revealed a triphasic response of inhibition/potentiation/inhibition, whereas control PP curves in CA1 manifested a biphasic response of inhibition/potentiation of test/conditioned PS amplitudes. Baclofen and bicuculline reversed the early and late phases of PP inhibition in the dentate and the early phase of PP inhibition in CA1. The GABAB antagonist, phaclofen, selectively reversed the effects of baclofen on PP inhibition in both the dentate and CA1. Whereas baclofen had no effect, bicuculline incre sed and phaclofen decreased DGC single-unit spontaneous firing rate, while baclofen decreased and bicuculline and phaclofen increased PC firing rate. These results support and extend studies suggesting that GABAergic feedback inhibition of DGCs and PCs is mediated by postsynaptic GABAA receptors and feedback inhibition of PCs is mediated by postsynaptic GABAB receptors. Our results also provide significant new evidence suggesting that postsynaptic inhibition in the dentate is not regulated by GABAB receptors and that feedback and feedforward inhibition of DGCs and PCs is regulated by presynaptic GABAB receptors located on GABAergic interneurons.     

GABAergic nonpyramidal neurons in intracerebral transplants of the rat hippocampus and fascia dentata: a combined light and electron microscopic immunocytochemical study

Glutamate decarboxylase (GAD) immunocytochemistry was used to study GABAergic neurons and synapses in intracerebral allografts of the rat hippocampus and fascia dentata. Tissue blocks of regio inferior of Ammon's horn (hippocampal field CA3) or of the fascia dentata were taken from newborn rats and transplanted to the hippocampal region of young adult rats. After 6 1/2 months' survival the recipient brains were fixed by perfusion and serially sectioned on a Vibratome. Sections containing the transplant and/or the host hippocampal region were immunostained for GAD and flat-embedded in Araldite for a correlated light and electron microscopic analysis. Immunostained neurons and terminals in the transplants were compared to immunoreactive elements in the hippocampus and fascia dentata of the hosts and other, normal rats. As in the hippocampal formation in situ, GAD-immunoreactive neurons and terminals in the transplants were observed in all layers. In dentate transplants a preponderance of immunostained cells was found just beneath the granule cell layer. In both hippocampal and dentate transplants, immunoreactive terminals were most abundant in the cell layers where they formed characteristic pericellular baskets around the pyramidal and granule cell bodies. In the electron microscope, the transplant GAD-immunoreactive neurons exhibited numerous cytoplasmic organelles, deeply infolded nuclei, and nuclear rods. Immunoreactive terminals formed symmetric synaptic contacts on the cell bodies, dendritic shafts, and spines of transplant pyramidal cells, granule cells, and hilar neurons. These are normal characteristics of GAD-immunoreactive neurons and terminals as also observed in the hippocampus of the host rats and the normal controls. Our results demonstrate that GABAergic neurons survive transplantation and develop a cell-specific morphology that includes the axonal projections.     

Afferent and efferent synaptic connections of somatostatin-immunoreactive neurons in the rat fascia dentata

The aim of this study was to determine whether somatostatin (SS)-immunoreactive neurons of the rat fascia dentata are involved in specific excitatory circuitries that may result in their selective damage in models of epilepsy. Synaptic connections of SS-immunoreactive neurons were determined at the electron microscopic level by using normal and colchicine pretreated rats. Vibratome sections prepared from both fascia dentata of control animals and from rats that had received an ipsilateral lesion of the entorhinal cortex 30-36 hours before sacrifice were immunostained for SS by using a monoclonal antibody (SS8). Correlated light and electron microscopic analysis demonstrated that many SS-immunoreactive neurons in the hilus send dendritic processes into the outer molecular layer of the fascia dentata, and dendrites of the same neurons occupy broad areas in the dentate hilar area. The majority of SS-immunoreactive axon terminals form symmetric synapses with the granule cell dendrites in the outer molecular layer and also innervate deep hilar neurons. Via their dendrites in the outer molecular layer, the SS-immunoreactive neurons receive synaptic inputs from perforant pathway axons which were identified by their anterograde degeneration following entorhinal lesions. The axons from the entorhinal cortex are the first segment of the main hippocampal excitatory loop. The hilar dendrites of the same SS-immunoreactive cells establish synapses with the mossy axon collaterals which represent the second member in this excitatory neuronal chain. These observations suggest that SS-immunoreactive neurons in the dentate hilar area may be driven directly by their perforant path synapses and via the granule cells which are known to receive a dense innervation from the entorhinal cortex. These observations demonstrate that SS-immunoreactive neurons in the hilar region are integrated in the main excitatory impulse flow of the hippocampal formation.     

Laminar differentiation of the hippocampus, fascia dentata and subiculum in developing rats, observed with the Timm sulphide silver method.

The laminar staining of the rat hippocampal region with the Timm sulphide silver method is from studies on adult rats known to depend on the various fibersystems terminating in these laminae. In order to illustrate the development of these fibersystems the laminar differentiation of the Timm staining of fascia dentata, hippocampus and subiculum is presented for rats between 1 and 31 days old. Corresponding changes in cytoarchitectonics as revealed by thionin staining are briefly demonstrated. Even on the first postnatal day there are indications of the mature, laminar staining pattern, and between three and nine days all the laminae corresponding to the terminal fields of the major afferent and intrinsic systems appear. After 12 days only minor additions to the laminar pattern develop, but there are adjustments of absolute and relative dimensions of layers and fields, and also the staining densities of individual laminae change. These observations are in good correlation with the available information on both hippocampal neurogenesis and cytodifferentiation, and the few fiber tracing studies performed on the developing hippocampal region. Compared to the latter, which ideally marks only one system or one lamina per animal, the Timm method provides an excellent means for getting an overview of the general developmental situation at the different ages. Thus developmental gradients along septotemporal, medio-lateral and basal-apical axes are found; which should be heeded in future studies on hippocampal synaptogenesis. The observations are discussed in relation to current models for neuronal growth and formation of nervous connections.     

Lesion-induced plasticity of high affinity choline uptake in the developing rat fascia dentata

After removal of the perforant path input to the rat fascia dentata at the age of 11 days, cholinergic septohippocampal fibers invade the denervated area. We have examined the effect of this lesion on hemicholinium-sensitive, high affinity choline uptake and its coupling to acetylcholine synthesis, specific properties of the septohippocampal input. Removal of the ipsilateral perforant path fibers increased the velocity of high affinity choline uptake by dentate particulate preparations, usually within 1 day. Studies conducted 5–104 days after operation showed a consistent 50–65% elevation in the molecular (denervated) layer. In contrast, the choline uptake rate in the granular layer eventually decreased slightly. Calculation of choline uptake rates independently of protein (per whole region) revealed that fasciae dentatae from operated and control sides accumulated choline at approximately equal rates, but on the operated side a greater percentage was transported by structures from the molecular layer and a lesser percentage by those from the granular layer. The rate of acetylcholine synthesis from exogenous choline increased to the same extent as high affinity choline uptake from 3 days after operation onward. The changes in high affinity choline uptake and acetylcholine synthesis coincided spatially and temporally with the reactive growth of septohippocampal fibers. Our results support the view that a perforant path lesion during development permanently alters the distribution of functional septohippocampal boutons in the fascia dentata. Acetylcholine synthesis is regulated to the same extent by high affinity choline uptake in the anomalous boutons as in normally located boutons.     

Ultrastructural localization of activated NF-kappaB in granule cells of the rat fascia dentata

The distribution of activated NF-kappaB (p65) was studied in granule cells of the rat fascia dentata using confocal laser scanning microscopy and preembedding immunogold electron microscopy. Activated NF-kappaB, detected with a monoclonal antibody specific for the nuclear localization signal-epitope, was found in proximal dendrites, somata, and nuclei but not in axon terminals of granule cells. Within the nuclei of granule cells, clusters of NF-kappaB immunoreactivity were detected. These data are in line with the proposed function of NF-kappaB as a retrograde signal transducer which transports information from synaptic sites to the nucleus to initiate gene expression.     

Lateralized fascia dentata lesion and blockade of one hippocampus: Effect on spatial memory in rats

Unilateral blockade of the dorsal hippocampus by tetrodotoxin makes it possible to form lateralized spatial memories, which rapidly transfer to the naive hippocampus when training continues with intact brain. Unilateral X-ray irradiation of newborn rats causes irreversible destruction of granule cells in the ipsilateral fascia dentata (FD). Possible compensation of poor learning in the lesioned hemisphere by commissural transfer of memories from the intact hippocampus was examined in seven rats with unilateral FD lesion, which were first trained in the Morris water maze to asymptotic performance (mean escape latency 6 ± 1 s). Subsequent testing during functional ablation either of the intact or of the lesioned hippocampus by tetrodotoxin revealed escape latencies 35 ± 8 s or 8 ± 1 s, respectively. Probe trial tests during inactivation of the intact and lesioned hippocampus showed target quadrant preference of 32 ± 2% or 54 ±3%, respectively. The results indicate: (a) that one intact hippocampus alone can support the water maze task, (b) that no, or only a very weak, memory trace is available in the lesioned hippocampus. It is concluded that the above results are due to the inability of the FD lesioned hippocampus to process the information received from the ipsilateral entorhinal cortex. Hippocampus 1998;8:647–650. © 1998 Wiley-Liss, Inc.     

Heterotypic neuronal differentiation of adult subependymal zone neuronal progenitor cells transplanted to the adult hippocampus

Neuronal progenitor cells (NPCs) residing in the adult subependymal zone (SEZ) are a potential source of expandable cells for autologous transplantation to replace neurons lost in multiple types of brain injury. To characterize the capacity of these cells for neuronal differentiation in a mature ectopic environment, NPCs expanded from the SEZ of adult rats were transplanted to the adult dentate gyrus. Cultures comprised a heterogeneous population of proliferating cells, which expressed nestin (47%) and GFAP (37%), with many cells expressing both progenitor cell markers (31%). In grafts of undifferentiated cells, as well as in grafts of cells that were induced to differentiate in vitro with retinoic acid, 35% of the transplanted SEZ-derived cells exhibited immunohistochemical and morphological features characteristic of hippocampal granule cell neurons. These novel results indicate that in vitro expanded adult SEZ NPCs are capable of heterotypic neuronal differentiation in a neurogenic region of the adult brain.     

Comparison of commissural sprouting in the mouse and rat fascia dentata after entorhinal cortex lesion

Reactive axonal sprouting occurs in the fascia dentata after entorhinal cortex lesion. This sprouting process has been described extensively in the rat, and plasticity-associated molecules have been identified that might be involved in its regulation. To demonstrate causal relationships between these candidate molecules and the axonal reorganization process, it is reasonable to analyze knockout and transgenic animals after entorhinal cortex lesion, and because gene knockouts are primarily generated in mice, it is necessary to characterize the sprouting response after entorhinal cortex lesion in this species. In the present study, Phaseolus vulgaris-leucoagglutinin (PHAL) tracing was used to analyze the commissural projection to the inner molecular layer in mice with longstanding entorhinal lesions. Because the commissural projection to the fascia dentata is neurochemically heterogeneous, PHAL tracing was combined with immunocytochemistry for calretinin, a marker for commissural/associational mossy cell axons. Using both techniques singly as well as in combination (double-immunofluorescence) at the light or electron microscopic level, it could be shown that in response to entorhinal lesion mossy cell axons leave the main commissural fiber plexus, invade the denervated middle molecular layer, and form asymmetric synapses within the denervated zone. Thus, the commissural sprouting response in mice has a considerable translaminar component. This is in contrast to the layer-specific commissural sprouting observed in rats, in which the overwhelming majority of mossy cell axons remain within their home territory. These data demonstrate an important species difference in the commissural/associational sprouting response between rats and mice that needs to be taken into account in future studies.