The Endogenous Stress Hormone CRH Modulates Excitatory Transmission and Network Physiology in Hippocampus.

Memory is strongly influenced by stress but underlying mechanisms are unknown. Here, we used electrophysiology, neuroanatomy, and network simulations to probe the role of the endogenous, stress-related neuropeptide corticotropin-releasing hormone (CRH) in modulating hippocampal function. We focused on neuronal excitability and the incidence of sharp waves (SPWs), a form of intrinsic network activity associated with memory consolidation. Specifically, we blocked endogenous CRH using 2 chemically distinct antagonists of the principal hippocampal CRH receptor, CRHR1. The antagonists caused a modest reduction of spontaneous excitatory transmission onto CA3 pyramidal cells, mediated, in part by effects on IAHP. This was accompanied by a decrease in the incidence but not amplitude of SPWs, indicating that the synaptic actions of CRH are sufficient to alter the output of a complex hippocampal network. A biophysical model of CA3 described how local actions of CRH produce macroscopic consequences including the observed changes in SPWs. Collectively, the results provide a first demonstration of the manner in which subtle synaptic effects of an endogenously released neuropeptide influence hippocampal network level operations and, in the case of CRH, may contribute to the effects of acute stress on memory.

Region-specific alteration of GABAergic markers in the brain of heterozygous reeler mice.

Heterozygous reeler mice (HRM), haploinsufficient for reelin, have been proposed to be a genetic mouse model of schizophrenia. Beside behavioural similarities, HRM also demonstrate several neuroanatomical traits similar to patients suffering from schizophrenia. In the present study using immunocytochemical procedures, we investigated HRM and wild-type mice (WT) for differences in the numbers and densities of glutamic acid decarboxylase (GAD)67 and parvalbumin (PARV)-immunoreactive (IR) neurons in the hippocampus, tyrosine hydroxylase (TH)-IR neurons in the ventral tegmental area (VTA) and substantia nigra (SN), and serotonin transporter (5-HT-T)-IR neurons of the raphe nuclei. We found that HRM, compared with WT, show a significant decrease of GAD67-IR neurons in hippocampal subregion CA1 [stratum pyramidale (SP)], CA2 [stratum oriens (SO), stratum pyramidale (SP) and stratum radiatum (SR)] and dentate gyrus [granule cell layer (GL)], and also a significant decrease of PARV-containing neurons in CA1 (SO, SP) and CA2 (SP). No morphological differences were found in the SN/VTA or raphe nuclei. In conclusion, these results support a hippocampal γ-aminobutyric acid (GABA)ergic dysfunction in HRM as previously described by other authors, and may be based on a downregulation of GAD67 and PARV expressions. In summary, the reelin haploinsufficient mouse may provide a useful model for studying the interaction between reelin and hippocampal GABAergic system, its effect on dendritic spine maturation and plasticity related to schizophrenia.

Targeting gene-modified hematopoietic cells to the central nervous system: use of green fluorescent protein uncovers microglial engraftment.

Gene therapy in the central nervous system (CNS) is hindered by the presence of the blood-brain barrier, which restricts access of serum constituents and peripheral cells to the brain parenchyma. Expression of exogenously administered genes in the CNS has been achieved in vivo using highly invasive routes, or ex vivo relying on the direct implantation of genetically modified cells into the brain. Here we provide evidence for a novel, noninvasive approach for targeting potential therapeutic factors to the CNS. Genetically-modified hematopoietic cells enter the CNS and differentiate into microglia after bone-marrow transplantation. Up to a quarter of the regional microglial population is donor-derived by four months after transplantation. Microglial engraftment is enhanced by neuropathology, and gene-modified myeloid cells are specifically attracted to the sites of neuronal damage. Thus, microglia may serve as vehicles for gene delivery to the nervous system.

Regulation of synaptic efficacy by coincidence of postsynaptic APs and EPSPs.

Activity-driven modifications in synaptic connections between neurons in the neocortex may occur during development and learning. In dual whole-cell voltage recordings from pyramidal neurons, the coincidence of postsynaptic action potentials (APs) and unitary excitatory postsynaptic potentials (EPSPs) was found to induce changes in EPSPs. Their average amplitudes were differentially up- or down-regulated, depending on the precise timing of postsynaptic APs relative to EPSPs. These observations suggest that APs propagating back into dendrites serve to modify single active synaptic connections, depending on the pattern of electrical activity in the pre- and postsynaptic neurons.

Importance of AMPA receptors for hippocampal synaptic plasticity but not for spatial learning.

Gene-targeted mice lacking the L-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor subunit GluR-A exhibited normal development, life expectancy, and fine structure of neuronal dendrites and synapses. In hippocampal CA1 pyramidal neurons, GluR-A-/- mice showed a reduction in functional AMPA receptors, with the remaining receptors preferentially targeted to synapses. Thus, the CA1 soma-patch currents were strongly reduced, but glutamatergic synaptic currents were unaltered; and evoked dendritic and spinous Ca2+ transients, Ca2+-dependent gene activation, and hippocampal field potentials were as in the wild type. In adult GluR-A-/- mice, associative long-term potentiation (LTP) was absent in CA3 to CA1 synapses, but spatial learning in the water maze was not impaired. The results suggest that CA1 hippocampal LTP is controlled by the number or subunit composition of AMPA receptors and show a dichotomy between LTP in CA1 and acquisition of spatial memory.

Submillisecond AMPA receptor-mediated signaling at a principal neuron-interneuron synapse.

Glutamatergic transmission at a principal neuron-interneuron synapse was investigated by dual whole-cell patch-clamp recording in rat hippocampal slices combined with morphological analysis. Evoked EPSPs with rapid time course (half duration = 4 ms; 34 degrees C) were generated at multiple synaptic contacts established on the interneuron dendrites close to the soma. The underlying postsynaptic conductance change showed a submillisecond rise and decay, due to the precise timing of glutamate release and the rapid deactivation of the postsynaptic AMPA receptors. Simulations based on a compartmental model of the interneuron indicated that the rapid postsynaptic conductance change determines the shape and the somatodendritic integration of EPSPs, thus enabling interneurons to detect synchronous principal neuron activity.

Sprouting in the hippocampus is layer-specific.

Partial removal of layer-specific afferents of the hippocampus is said to induce sprouting of intact fibers from neighboring layers that invade the zone of the degenerating axons. However, recent in vivo and in vitro studies using sensitive anterograde tracers have failed to demonstrate sprouting across laminar boundaries. Sprouting does occur; but, it mainly involves unlesioned fiber systems terminating within the layer of fiber degeneration in addition to the degenerating afferents. These findings point to rigid laminar cues attracting certain fiber systems while repelling others in normal development and after partial deafferentation.

Cajal-Retzius cells, Reelin, and the formation of layers.

Early-generated Cajal-Retzius cells in the marginal zone of the cortex synthesize and secrete the glycoprotein Reelin. The reelin gene is deleted in reeler mice, which show characteristic alterations in cortical lamination. Recent studies have shed some light on the role of Cajal-Retzius cells and Reelin in the formation of cell and fiber layers in the neocortex and hippocampus.

Lesion-induced plasticity of central neurons: sprouting of single fibres in the rat hippocampus after unilateral entorhinal cortex lesion.

In response to a central nervous system trauma surviving neurons reorganize their connections and form new synapses that replace those lost by the lesion. A well established in vivo system for the analysis of this lesion-induced plasticity is the reorganization of the fascia dentata following unilateral entorhinal cortex lesions in rats. After general considerations of neuronal reorganization following a central nervous system trauma, this review focuses on the sprouting of single fibres in the rat hippocampus after entorhinal lesion and the molecular factors which may regulate this process. First, the connectivity of the fascia dentata in control animals is reviewed and previously unknown commissural fibers to the outer molecular layer and entorhinal fibres to the inner molecular layer are characterized. Second, sprouting of commissural and crossed entorhinal fibres after entorhinal cortex lesion is described. Single fibres sprout by forming additional collaterals, axonal extensions, boutons, and tangle-like axon formations. It is pointed out that the sprouting after entorhinal lesion mainly involves unlesioned fibre systems terminating within the layer of fibre degeneration and is therefore layer-specific. Third, molecular changes associated with axonal growth and synapse formation are considered. In this context, the role of adhesion molecules, glial cells, and neurotrophic factors for the sprouting process are discussed. Finally, an involvement of sprouting processes in the formation of neuritic plaques in Alzheimer's disease is reviewed and discussed with regard to the axonal tangle-like formations observed after entorhinal cortex lesion.

Novel and transient populations of corticotropin-releasing hormone-expressing neurons in developing hippocampus suggest unique functional roles: a quantitative spatiotemporal analysis.

Robust physiological actions of the neuropeptide corticotropin-releasing hormone (CRH) on hippocampal pyramidal neurons have been demonstrated, which may contribute to synaptic efficacy and to learning and memory processes. These excitatory actions of the peptide, as well as the expression of the CRH receptor type that mediates them, are particularly prominent during early postnatal life, suggesting that endogenous CRH may contribute to processes involved in maturation of hippocampal circuitry. To further elucidate the function(s) of endogenous CRH in developing hippocampus, we used neurochemical and quantitative stereological methods to characterize in detail CRH-expressing neuronal populations during postnatal hippocampal differentiation. These experiments revealed progressively increasing numbers of CRH-expressing neurons in developing hippocampus that peaked on postnatal day 11-18 and then declined drastically to adult levels. These cells belonged to several discrete populations, distinguished by GAD67 mRNA expression, morphology, and distinct spatiotemporal distribution profiles. Importantly, a novel population of Cajal-Retzius-like CRH-expressing neurons was characterized that exists only transiently in early postnatal hippocampus and is positioned to contribute to the establishment of hippocampal connectivity. These findings suggest novel, age-specific roles for CRH in regulating early developmental events in the hippocampal formation.

Rapid signaling at inhibitory synapses in a dentate gyrus interneuron network.

Mutual synaptic interactions between GABAergic interneurons are thought to be of critical importance for the generation of network oscillations and for temporal encoding of information in the hippocampus. However, the functional properties of synaptic transmission between hippocampal interneurons are largely unknown. We have made paired recordings from basket cells (BCs) in the dentate gyrus of rat hippocampal slices, followed by correlated light and electron microscopical analysis. Unitary GABA(A) receptor-mediated IPSCs at BC-BC synapses recorded at the soma showed a fast rise and decay, with a mean decay time constant of 2.5 +/- 0.2 msec (32 degrees C). Synaptic transmission at BC-BC synapses showed paired-pulse depression (PPD) (32 +/- 5% for 10 msec interpulse intervals) and multiple-pulse depression during repetitive stimulation. Detailed passive cable model simulations based on somatodendritic morphology and localization of synaptic contacts further indicated that the conductance change at the postsynaptic site was even faster, decaying with a mean time constant of 1.8 +/- 0.6 msec. Sequential triple recordings revealed that the decay time course of IPSCs at BC-BC synapses was approximately twofold faster than that at BC-granule cell synapses, whereas the extent of PPD was comparable. To examine the consequences of the fast postsynaptic conductance change for the generation of oscillatory activity, we developed a computational model of an interneuron network. The model showed robust oscillations at frequencies >60 Hz if the excitatory drive was sufficiently large. Thus the fast conductance change at interneuron-interneuron synapses may promote the generation of high-frequency oscillations observed in the dentate gyrus in vivo.

Cerebral amyloid induces aberrant axonal sprouting and ectopic terminal formation in amyloid precursor protein transgenic mice.

A characteristic feature of Alzheimer's disease (AD) is the formation of amyloid plaques in the brain. Although this hallmark pathology has been well described, the biological effects of plaques are poorly understood. To study the effect of amyloid plaques on axons and neuronal connectivity, we have examined the axonal projections from the entorhinal cortex in aged amyloid precursor protein (APP) transgenic mice that exhibit cerebral amyloid deposition in plaques and vessels (APP23 mice). Here we report that entorhinal axons form dystrophic boutons around amyloid plaques in the entorhinal termination zone of the hippocampus. More importantly, entorhinal boutons were found associated with amyloid in ectopic locations within the hippocampus, the thalamus, white matter tracts, as well as surrounding vascular amyloid. Many of these ectopic entorhinal boutons were immunopositive for the growth-associated protein GAP-43 and showed light and electron microscopic characteristics of axonal terminals. Our findings suggest that (1) cerebral amyloid deposition has neurotropic effects and is the main cause of aberrant sprouting in AD brain; (2) the magnitude and significance of sprouting in AD have been underestimated; and (3) cerebral amyloid leads to the disruption of neuronal connectivity which, in turn, may significantly contribute to AD dementia.

Neuronal basic helix-loop-helix proteins (NEX and BETA2/Neuro D) regulate terminal granule cell differentiation in the hippocampus.

The transcription factors neuronal helix-loop-helix protein (NEX)/mammalian atonal homolog 2 (Math-2), BETA2/neuronal determination factor (NeuroD), and NeuroD-related factor (NDRF)/NeuroD2 comprise a family of Drosophila atonal-related basic helix-loop-helix (bHLH) proteins with highly overlapping expression in the developing forebrain. The ability of BETA2/NeuroD and NDRF to convert ectodermal cells into neurons after mRNA injection into Xenopus oocytes suggested a role in specifying neuronal cell fate. However, neuronal bHLH genes are largely transcribed in CNS neurons, which are fully committed. Here we analyze a defect in mice lacking BETA2/NeuroD, and in NEX*BETA2/NeuroD double mutants, demonstrating that bHLH proteins are required in vivo for terminal neuronal differentiation. Most strikingly, presumptive granule cells of the dentate gyrus are generated but fail to mature, lack normal sodium currents, and show little dendritic arborization. Long-term hippocampal slice cultures demonstrate secondary alterations of entorhinal and commissural/associational projections. The primary developmental arrest appears to be restricted to granule cells in which an autoregulatory system involving all three neuronal bHLH genes has failed.

Neurosteroid synthesis in the hippocampus: role in synaptic plasticity.

Neurosteroids are still found in the brain after steroidogenic glands were removed, indicating that they are synthesized either de novo or from endogenous precursors by enzymes present in the CNS. In fact, steroidogenic acute regulatory protein, and aromatase, two molecules essential for estrogen synthesis, are expressed in the hippocampus. We recently showed, for the first time, that estrogens are synthesized de novo in hippocampal neurons and that these hippocampus-derived estrogens are essential for synaptic plasticity. Both estrogen receptor isoforms, estrogen receptor alpha and estrogen receptor beta, are expressed in the hippocampus, and estradiol treatment of the cultures leads to an upregulation of estrogen receptor alpha. This finding confirmed the presence of functional estrogen receptors in hippocampal neurons and showed the responsiveness of the cultured hippocampal neurons to estradiol. By using letrozole, an inhibitor of aromatase, estradiol levels in hippocampal dispersion cultures as well as in hippocampal slice cultures were significantly suppressed which in turn led to a downregulation of estrogen receptor alpha. Letrozole treatment was followed by a significant decrease in the density of spines and spine synapses and in the number of presynaptic boutons. Quantitative immunohistochemistry revealed a dose-dependent downregulation of spinophilin, a spine marker, and of synaptophysin, a presynaptic marker, and of growth-associated protein 43 after letrozole treatment. Our data provide strong evidence for estrogens being potent modulators of structural synaptic plasticity and point to a paracrine rather than endocrine mechanism of estrogen action in the hippocampus.

Src controls neuronal migration by regulating the activity of FAK and cofilin.

Migration of postmitotic neurons in the developing cortex along radial glial fiber is essential for the formation of cortical layers. Several neurological diseases are caused by defects in neuronal migration, underlining the importance of this process for brain function. Multiple molecules are involved in this process. However, the precise mechanisms are largely unknown. In the present study, we examined the expression of Src in the developing cortex and investigated the role of Src in neuronal migration and its cellular and molecular mechanisms. Our results showed that Src was strongly expressed in the cerebral cortex during corticogenesis and mainly targeted to the leading processes of migrating neurons. Overexpression of wildtype Src (Src-WT) and its mutants, constitutively active Src (Src-CA) and dominant negative Src (Src-DN) in the mouse brain by in utero electroporation perturbed neuronal migration through affecting the adhesion properties and cytoskeletal dynamics of migrating neurons. Overexpression of Src-WT and Src-CA induced aggregation and branching of migrating neurons, whereas overexpression of Src-DN led to abnormal elongation of the leading processes of migrating neurons. Furthermore, we showed that Src activates the focal adhesion kinase (FAK) and cofilin by regulating their phosphorylation levels. We conclude that Src controls neuronal migration by regulating adhesion properties and F-actin dynamics of migrating neurons.

Heterogeneity in the Membrane Current Pattern of Identified Glial Cells in the Hippocampal Slice.

Glial cells, acutely isolated or in tissue culture, have previously been shown to express a variety of voltage-gated channels. To resolve the question whether such channels are also expressed by glial cells in their normal cellular environment, we have applied the patch-clamp technique to study glial cells in hippocampal slices of 10 - 12-day-old mice. Based on the membrane current pattern, we distinguished four glial cell types. One was characterized by passive, symmetrical K+ currents activated in depolarizing and hyperpolarizing directions. A second population showed a similar current pattern, but with a marked decay of the current during the 50-ms voltage jumps. In a third population, the decaying passive currents were superimposed with a delayed rectifier outward current and, in some cases, with a slow inward current activated by depolarization. The fourth population expressed delayed rectifying outward currents, an inward rectifier K+ current and fast inward currents activated by depolarization. To unequivocally identify the glial cells we combined electrophysiological and ultrastructural characterizations. Therefore, cells were filled with the fluorescent dye lucifer yellow during characterization of their membrane currents, the fluorescence of the dye was used to convert diaminobenzidine to an electron-dense material, and subsequently slices were inspected in the electron microscope. Recordings were obtained from cells in the stratum radiatum and were identified as glial by their size, the characteristic chromatin distribution, and the lack of synaptic membrane specializations.

GABAergic innervation of the rat fascia dentata: a novel type of interneuron in the granule cell layer with extensive axonal arborization in the molecular layer.

By using the combined Golgi/electron microscopy (EM) technique and postembedding immunocytochemistry for gamma-aminobutyric acid (GABA), we describe a novel type of local circuit neuron in the rat fascia dentata that gives rise to an axon profusely ramifying in the dentate molecular layer. The relatively small ovoid cell body (long axis 12-15 microns) is located directly underneath the granular layer. From both poles of the cell body dendritic processes emerge that enter the molecular layer and hilar region, respectively. The apical dendrites traverse the granular layer, invade the molecular layer, and branch in the same way as granule cell dendrites. Some branches reach the hippocampal fissure. Thus, the apical dendrites of these neurons may receive a similar input pattern as the granule cells. The dendrites are smooth, occasionally bearing varicosities. A few spines are regularly observed. The axon originates from the apical dendrite and traverses the molecular layer horizontally for up to 500 microns. It gives off numerous collaterals that are distributed throughout the entire width of the molecular layer and only rarely enter the granule cell layer. Electron microscopy of the cell body of gold-toned neurons revealed the well-known fine-structural characteristics of nonpyramidal neurons, i.e., an indented nucleus with nuclear inclusions and large aggregations of endoplasmic reticulum. Apical as well as basal dendrites are densely covered with presynaptic boutons, mainly forming asymmetric synapses. The axon terminals of these cells form symmetric synapses with dendritic shafts and, to a lesser extent, with spines. These symmetric synapses, together with the results of our GABA postembedding immunocytochemical study, suggest that this cell is a GABAergic inhibitory neuron that almost exclusively innervates the dentate molecular layer. Together with data from the literature on dentate axoaxonic cells (which innervate the axon initial segments of the granule cells) and GABAergic basket cells (which innervate the granule cell somata and proximal dendrites in the granular layer), the present results indicate that there is a lamination of the GABAergic innervation of the fascia dentata corresponding to the well-known segregated termination of entorhinal and commissural afferents to this region.

Spiny nonpyramidal neurons in the CA3 region of the rat hippocampus are glutamate-like immunoreactive and receive convergent mossy fiber input.

There is increasing evidence that the various types of hippocampal nonpyramidal neurons control the principal cells in different ways. In the present study a type of spiny nonpyramidal cell in stratum lucidum of rat hippocampal region CA3 was studied by Golgi impregnation. Three Golgi-impregnated and gold-toned neurons of this type were further analyzed by electron microscopy and postembedding immunocytochemistry. The dendrites of these bipolar neurons seemed to be restricted to stratum lucidum and ran parallel with the mossy fibers that terminate in this layer. A characteristic feature of this neuron is the presence of long, thin spines on both cell body and dendrites. Although these dendrites were exposed to a large number of mossy fibers, no thorny excrescences were formed which are characteristic postsynaptic elements of CA3 pyramidal neurons for synaptic contact with the mossy fibers. Semithin sections of Golgi-impregnated and gold-toned stratum lucidum cells displayed immunoreactivity of the cell body region for glutamate but not for GABA. A fine-structural analysis of gold-toned sections revealed a large cell body with numerous cytoplasmic organelles and an indented nucleus. Numerous asymmetric synapses were found on dendritic shafts as well as on the long, thin somatic and dendritic spines. Usually, several presynaptic boutons contacted a single spine. The majority of these asymmetric spine synapses were probably of mossy fiber origin, although no giant mossy fiber synapses were formed. The long spines were contacted by much smaller en passant synapses of preterminal axons. In contrast, giant mossy fiber boutons were found presynaptic to dendritic shafts and cell bodies of these cells. Our morphological analysis of a glutamate-immunoreactive, GABA-negative type of nonpyramidal neuron that receives convergent mossy fiber input suggests that the impulse flow within the "trisynaptic pathway" is more complex than previously assumed.

Lesion-induced mossy fibers to the molecular layer of the rat fascia dentata: identification of postsynaptic granule cells by the Golgi-EM technique.

The axons of the dentate granule cells, the hippocampal mossy fibers, sprout "backward" into the dentate molecular layer when this is heavily denervated. Using the combined Golgi-electron microscopy (EM) technique we now demonstrate that these aberrant supragranular mossy fibers at least in part terminate on granule cell dendrites. Sprouting of mossy fibers into the dentate molecular layer was induced in adult rats by simultaneous surgical removal of the commissural and entorhinal afferents to the fascia dentata. After at least 7 weeks survival, the presence of mossy fiber terminals in the inner part of the dentate molecular layer was demonstrated by light microscopy. In the electron microscope the mossy fiber terminals were identified by their unique structural characteristics, namely, the unusually large size of the terminals, the dense packing of clear synaptic vesicles with a few dense core vesicles intermingled, the presence of asymmetric synaptic contacts with spines and desmosome-like contacts with dendritic shafts, and the continuity with a thin unmyelinated preterminal axon. Golgi-stained granule cells were first identified in the light microscope, and then, after deimpregnation, the same cells were examined in the electron microscope. In ultrathin, serial sections lesion-induced mossy fiber terminals were found in synaptic contact with spines on proximal dendritic segments of such identified Golgi-impregnated granule cells. From this we conclude that the aberrant, supragranular mossy fibers can innervate dendrites of the parent cell group, the dentate granule cells. The results, moreover, provide an example of reactive synaptogenesis where both the sprouted afferents and its postsynaptic element have been identified.

The cholinergic innervation of the rat fascia dentata: identification of target structures on granule cells by combining choline acetyltransferase immunocytochemistry and Golgi impregnation.

A monoclonal antibody against choline acetyltransferase (ChAT), the acetylcholine-synthesizing enzyme, was used to study cholinergic synapses on identified (Golgi stained) granule cells in the rat fascia dentata. Choline acetyltransferase immunocytochemistry was applied to 40-microns Vibratome sections cut perpendicular to the longitudinal axis of the hippocampus. Light microscopy revealed fine varicose ChAT-immunoreactive axons in all layers of the fascia dentata, i.e., in the stratum moleculare, the stratum granulosum, and the subgranular polymorph zone. Most fibers were observed in the vicinity of granule cell bodies where they ran mainly parallel to the granular layer. Next, the immunostained Vibratome sections were sandwiched between small pieces of Parafilm and piled to form a block that was covered with agar and Golgi stained. After that, the sections were separated by cutting away the agar and removing the Parafilm. Sections containing well-impregnated granule cells were gold-toned (Fairén et al., '77), embedded in Araldite, and subjected to ultrathin sectioning for electron microscopy. A total of 14 gold-toned granule cells were examined in the electron microscope for synaptic contacts with cholinergic afferents. Choline acetyltransferase-immunoreactive axon terminals were observed that established symmetric synaptic contacts with the cell bodies and dendritic shafts of the gold-toned identified granule cells. Two types of contact were observed on spines arising from gold-toned granule cell dendrites. Immunoreactive terminals established asymmetric synaptic contacts with the head of small spines and symmetric contacts with the stalk of large, complex spines. The boutons forming asymmetric synaptic contacts with the cup-shaped spine head of the complex spines were not found to be immunoreactive. Our results demonstrate that cholinergic fibers to the rat fascia dentata establish characteristic types of synaptic contact with different postsynaptic elements of granule cells, suggesting a complex function of this afferent system.