Unilateral entorhinal denervation leads to long-lasting dendritic alterations of mouse hippocampal granule cells

Following brain injury, neurons efferently connected from the lesion site are denervated and remodel their dendritic tree. Denervation-induced dendritic reorganization of granule cells was investigated in the dentate gyrus of the Thy1-GFP mouse. After mechanical transection of the perforant path, single granule cells were 3D-reconstructed at different time points post-lesion (3 d, 7 d, 10 d, 30 d, 90 d and 180 d) and their soma size, total dendritic length, number of dendritic segments and dendritic branch orders were studied. Changes in spine densities were determined using 3D-analysis of individual dendritic segments. Following entorhinal denervation the granule cell arbor progressively atrophied until 90 d post-lesion (reduction of total dendritic length to ~ 50% of control). Dendritic alterations occurred selectively in the denervated outer molecular layer, where a loss of distal dendritic segments and a reduction of mean segment length were seen. At 180 d post-lesion total dendritic length partially recovered (~ 70% of control). This recovery appeared to be the result of a re-elongation of surviving dendrites rather than dendritic re-branching, since the number of dendritic segments did not recover. In contrast to the protracted dendritic changes, spine density changes followed a faster time course. In the denervated layer spine densities dropped to ~ 65% of control values and fully recovered by 30 d post-lesion. We conclude that entorhinal denervation in mouse causes protracted and long-term structural alterations of the granule cell dendritic tree. Spontaneously occurring reinnervation processes, such as the sprouting of surviving afferent fibers, are insufficient to maintain the granule cell dendritic arbor.     

Morphological variability is a characteristic feature of granule cells in the primate fascia dentata: A combined Golgi/electron microscope study

This study analyzes the structural variability of granule cells in the monkey fascia dentata. The hippocampi of three adult rhesus monkeys (Macaca mulatta) and two 1-year-old female baboons (Papio anubis) were used for a combined Golgi/electron microscope (EM) study. The results were compared with other Golgi/EM studies on dentate granule cells in small laboratory animals. Whereas the granule cells in small rodents form a relatively uniform population of neurons, we observed a much greater variability of granule cell morphology in monkeys. This variability concerned the size of the cell body, the length and thickness of apical dendrites, the spine density, and the occasional occurrence of basal dendrites. The dendritic length of individual granule cells largely depended on their position in the highly convoluted granular layer. These convolutions caused significant variations in the thickness of the molecular layer and consequently in the length of individual granule cell dendrites. Granule cells with thick dendrites densely covered with spines could be differentiated from those exhibiting much thinner dendritic processes and moderate spine numbers. About 10% of granule cells in the monkey fascia dentata exhibited basal dendrites. Occasionally in the hilus ectopic granule cells were observed that gave rise to long apical dendrites traversing the granular layer. The axons of granule cells, the mossy fibers, entered the hilus, where they gave off several collaterals. In contrast to the light microscopic variability, subtypes of granule cells revealed similar fine structural characteristics, i. e., a round nucleus lacking indentations, a thin rim of cytoplasm, and characteristic spine formations. Large complex spines and smaller, “stubby” spines were observed on apical as well as basal dendrites. This suggests that characteristic spine formations were not induced by specific afferent fibers. All synaptic contacts on spines were of the asymmetric type, whereas both symmetric and asymmetric synapses occured on cell bodies and dendritic shafts. Unlike in rodents, we found a large variability of granule cells in the primate fascia dentata. This variability has to be considered in neropathological studies of this cell type.     

Quantitative, three-dimensional analysis of granule cell dendrites in the rat dentate gyrus

The three-dimensional organization of dentate granule cell dendritic trees has been quantitatively analyzed with the aid of a computerized microscope system. The dendrites were visualized by iontophoretic injection of horseradish peroxidase into individual granule cells in the in vitro hippocampal slice preparation. Selection criteria insured that the analyzed cells were completely stained and that only neurons with two or fewer cut dendrites in the distal portion of the molecular layer were analyzed. Twenty-nine of the 48 sampled granule cells had no cut dendrites. The granule cells had between one and four primary dendrites. Granule cell dendritic branches were covered with spines and most extended to the hippocampal fissure or pial surface. The mean total dendritic length was 3,221 microns with a range from 2,324 microns to 4,582 microns. The dendrites formed an elliptical plexus with the transverse spread averaging 325 microns and the spread in the septotemporal axis averaging 176 microns. On individual neurons, the maximum branch order ranged from four to eight and the number of dendritic segments ranged from 22 to 40. Approximately 63% of the dendritic branch points occurred in a zone that included the granule cell layer and the inner one-third of the molecular layer. The dendritic tree was organized so that, on average, 30% of the length was in the granule cell layer and proximal third of the molecular layer, 30% was in the middle third, and 40% was in the distal third. Comparisons were made between the dendrites of granule cells in the suprapyramidal and infrapyramidal blades of the dentate gyrus. Suprapyramidal cells had a significantly greater total dendritic length than infrapyramidal cells, their transverse spread was higher, and they had a greater number of dendritic segments. When neurons in the suprapyramidal blade were further subdivided on the basis of somal position within the depth of the cell body layer, superficial neurons were found to have a greater number of primary dendrites, more elliptical trees, and larger transverse spreads of their dendrites. There were no significant differences in dendritic segment number or total dendritic length between superficial and deep cells.     

Axon arbors and synaptic connections of hippocampal mossy cells in the rat in vivo

The axon collateralization patterns and synaptic connections of intracellularly labeled and electrophysiologically identified mossy cells were studied in rat hippocampus. Light microscopic analysis of 11 biocytin-filled cells showed that mossy cell axon arbors extended through an average of 57% of the total septotemporal length of the hippocampus (summated two-dimensional length, not adjusted for tissue shrinkage). Axon collaterals were densest in distant lamellae rather than in lamellae near the soma. Most of the axon was concentrated in the inner one-third of the molecular layer, with the hilus containing an average of only 26% of total axon length and the granule cell layer containing an average of only 7%. Ultrastructural analysis was carried out on three additional intracellularly stained mossy cells, in which axon collaterals and synaptic targets were examined in serial sections of chosen axon segments. In the central and subgranular regions of the hilus, mossy cell axons established a low density of synaptic contacts onto dendritic shafts, neuronal somata, and occasional dendritic spines. Most hilar synapses were made relatively close to the mossy cell somata. At greater distances from the labeled mossy cell (1–2 mm along the septotemporal axis), the axon collaterals ramified predominantly within the inner molecular layer and made a high density of asymmetric synaptic contacts almost exclusively onto dendritic spines. Quantitative measurements indicated that more than 90% of mossy cell synaptic contacts in the ipsilateral hippocampus are onto spines of proximal dendrites of presumed granule cells. These results are consistent with a primary mossy cell role in an excitatory associational network with granule cells of the dentate gyrus. © 1996 Wiley-Liss, Inc.     

Hippocampal mossy fiber sprouting and synapse formation after status epilepticus in rats: Visualization after retrograde transport of biocytin

In complex partial epilepsy and in animal models of epilepsy, hippocampal mossy fibers appear to develop recurrent collaterals, that invade the dentate molecular layer. Mossy fiber collaterals have been proposed to subserve recurrent excitation by forming granule cell-granule cell synapses. This hypothesis was tested by visualizing dentate granule cells and their mossy fibers after terminal uptake and retrograde transport of biocytin. Labeling studies were performed with transverse slices of the caudal rat hippocampal formation prepared 2.6–l70.0 weeks after pilocarpine-induced or kainic acid-induced status epilepticus. Light microscopy demonstrated the progressive growth of recurrent mossy fibers into the molecular layer; the densest innervation was observed in slices from pilocarpine-treated rats that had survived 10 weeks or longer after status epilepticus. Thin mossy fiber collaterals originated predominantly from deep within the hilar region, crossed the granule cell body layer, and formed an axonal plexus oriented parallel to the cell body layer within the inner one-third of the molecular layer. When sprouting was most robust, some recurrent mossy fibers at the apex of the dentate gyrus reached the outer two-thirds of the molecular layer. The distribution and density of mossy fiber-like Timm staining correlated with the biocytin labeling. When viewed with the electron microscope, the inner one-third of the dentate molecular layer contained numerous mossy fiber boutons. In some instances, biocytin-labeled mossy fiber boutons were engaged in synaptic contact with biocytin-labeled granule cell dendrites. Granule cell dendrites did not develop large complex spines (“thorny excrescences”) at the site of synapse formation, and they did not appear to have been permanently damaged by seizure activity. These results establish the validity of Timm staining as a marker for mossy fiber sprouting and support the view that status epilepticus provokes the formation of a novel recurrent excitatory circuit in the dentate gyrus. Retrograde labeling with biocytin showed that the recurrent mossy fiber projection often occupies a considerably greater fraction of the dendritic region than previous studies had suggested. © 1995 Wiley-Liss, Inc.     

Axon arbors and synaptic connections of a vulnerable population of interneurons in the dentate gyrus in vivo

The predominant gamma-aminobutyric acid (GABA)ergic neuron class in the hilus of the dentate gyrus consists of spiny somatostatinergic interneurons. We examined the axon projections and synaptic connections made by spiny hilar interneurons labeled with biocytin in gerbils in vivo. Axon length was 152-497 mm/neuron. Sixty to 85% of the axon concentrated in the outer two thirds of the molecular layer of the dentate gyrus. The septotemporal span of the axon arbor extended over 48-82% of the total hippocampal length, which far exceeds the septotemporal span of axons of granule cells whose complete axon arbors extended over 15-29%. A three-dimensionally reconstructed 216-microm-long spiny hilar interneuron axon segment in the outer third of the molecular layer formed an average of 1 synapse every 5.1 microm. Of the 42 symmetric (inhibitory) synapses formed by the reconstructed segment, 88% were with spiny dendrites of presumed granule cells, and 67% were with dendritic spines that also receive an asymmetric (excitatory) contact from an unlabeled axon terminal. Postembedding GABA-immunocytochemistry revealed that 55% of the GABAergic synapses in the outer third of the molecular layer were with spines. Therefore, in the outer molecular layer, spiny hilar interneurons form synaptic contacts that appear to be positioned to exert inhibitory control near sites of excitatory synaptic input from the entorhinal cortex to granule cell dendritic spines. These findings demonstrate far-reaching, yet highly specific, connectivity of individual interneurons and suggest that the loss of spiny hilar interneurons, as occurs in temporal lobe epilepsy, may contribute to hyperexcitability in the hippocampus.     

Postnatal development of dendritic spines on olfactory bulb granule cells in rats

Postnatal morphological changes in granule cell dendritic spines and filopodia (collectively referred to as "spines/filopodia") were examined in the rat main olfactory bulb to characterize the development of the neural circuitry for olfaction. Granule cells were labeled with a membrane dye and confocal laser scanning microscope images of labeled spines/filopodia were acquired in the following three dendritic domains: apical dendrites in the external plexiform layer, those in the granule cell layer, and basal dendrites. In all three domains the proportion of typical spines slightly increased during development, with a concomitant decrease in the proportion of "stubby" spines lacking a neck; the proportion of filopodia remained unchanged, accounting for 20-40% of all protrusions. The mean diameter and length of the spine/filopodium population were nearly constant throughout development. On the other hand, the developmental pattern of the spine/filopodium density varied markedly, depending on the domain of the dendrites. In the external plexiform layer, the density did not change remarkably during development. The density in apical dendrites in the granule cell layer increased during the initial 2 postnatal weeks, then gradually decreased. The spine/filopodium density in basal dendrites, however, continued to increase until 4 weeks of age, and then began to decrease. These results suggest that a substantial amount of input-specific synaptic remodeling occurs in granule cells during development, which proceeds from superficial dendritic domains to deeper ones, occurring most prominently in the basal dendrites.     

Dendritic patterns of granule cells in organotypic explants of fetal and neonatal mouse hippocampal formation

Granule and granule-like neurons were labeled by Golgi and HRP techniques in the dentate area of fetal and neonatal organotypic hippocampal explants after 1 day-8 weeks in vitro. These cells resembled granule cells labeled in situ with similar techniques, although the dendritic pattern and spine development were not as elaborate as observed on granule cells from adult rodents. Many of these neurons retained basilar or multiple dendrites after 8 weeks in culture, a characteristic often associated with immature granule cells, granule cells in the reeler mutant mouse and tissues removed from human epileptic foci.     

Aberrant dentate gyrus cytoarchitecture and fiber lamination in Lis1 mutant mice

Mutant mice with a heterozygous deletion of LIS1, show varying degrees of hippocampal abnormality and enhanced excitability. To examine how LIS1 affects cytoarchitecture and fiber lamination in dentate gyrus (DG), we performed a series of immunohistochemistry studies. By using different neuronal- and glial-specific antibodies, we found that the majority of hippocampal cell populations were affected by heterozygous mutation of LIS1; some reelin-positive Cajal-Retzius cells were left undisturbed. Granule cell dispersion was significant in hippocampal sections from Lis1-deficient mice. However, the fiber termination of commissural/associational fibers and mossy fibers appeared relatively compact despite obvious granule cell dispersion and CA1-CA3 pyramidal cell disorganization. vGlut1-immunoreactive axon terminals were found aberrantly traversing the dispersed granule cell layer. Consistent with previous observations, we also found that immature granule cells in Lis1 mutants, here stained with antibodies to doublecortin (DCX) and Mash-1, are aberrantly located and bear an abnormal cellular morphology. Our findings suggest that LIS1 mutants exhibit abnormal cell positioning and aberrant hippocampal neurogenesis, but maintain relatively normal fiber termination patterns. The functional consequences of hippocampal granule cell dispersion could offer critical insight to the epileptic and cognitive disorder associated with LIS1 haploinsufficiency.     

Fine structure of identified neurons in the primate hippocampus: a combined Golgi/EM study in the baboon

The hippocampi of two 1-year-old female baboons (Papio anubis) were used for a combined Golgi/electron microscope (EM) study of characteristic cell types in the hippocampus proper and fascia dentata. Results were compared with previous Golgi/EM studies of hippocampal neurons in small laboratory animals. Cell bodies of pyramidal neurons in CA1 were more loosely distributed than known from studies on the rat or guinea pig. Numerous basal and horizontal dendrites originating from the perikaryon filled in the space between neighboring cell bodies. Apical stem dendrites were varying in length, depending on the position of the parent cell body in outer or inner portions of the pyramidal layer. Dendrites were densely covered with spines which in the EM showed very complex synaptic contacts. In contrast to our observations in rats and guinea pigs, CA3 pyramidal cells in the monkey hippocampus exhibited numerous large spines or excrescences not only on apical dendrites but also on basal dendrites running through stratum oriens. These excrescences appeared to be more complex than in small rodents. They often branched, protruding deeply into presynaptic mossy fiber boutons, and formed multiple asymmetric synaptic contacts. Granule cells of the monkey fascia dentata, in contrast to those of the rodent, occasionally had basal dendrites extending into the hilar region. In the EM, granule cells either with or without basal dendrites exhibited fine structural characteristics that were very similar to those described in Golgi/EM studies of granule cells in the rat fascia dentata. Of the various types of nonpyramidal neurons the horizontal cells in stratum oriens with dendrites parallel to the alveus were analyzed. As seen in rats, these cells exhibited large amounts of rough endoplasmic reticulum, indentations of the nuclear membrane, and nuclear inclusions. Numerous terminals formed synaptic contacts on dendritic shafts. In contrast to rodents, numerous spines arose from dendrites and cell bodies of these neurons. In the EM, often single spines were found to establish synaptic contacts with several presynaptic boutons. In summary, our correlated light and EM study of four characteristic cell types, which are present in both nonprimates and primates, demonstrates a much more complex dendritic pattern and synaptic organization of these neurons in primates than in commonly studied small laboratory animals.     

A Golgi-electron microscopic study of fusiform neurons in the hilar region of the dentate gyrus

The fusiform cells of the dentate gyrus are located in a portion of the hilus within 100 micron of the granule cell layer. They have ovoid somata and bipolar dendrites that generally run parallel to the granule cell layer. The dendrites of these cells are either spiny or sparsely spiny. The spiny fusiform cell has numerous spines along its dendrites, which are contacted by terminals with the features of granule cell axon collaterals. This cell type also displays somal spines that are contacted by similar terminals. In contrast, the sparsely spiny fusiform cell displays only a few spines, which are contacted by multiple small axon terminals that synapse with both the stalk and end bulb of the spine. Most synaptic input for this cell type is made with the smooth surfaces of the soma and dendrites. A variety of terminals form synapses with the sparsely spiny fusiform cell, including terminals that resemble the fine axon collaterals of mossy fibers. The somata of these two cell types also display differences in the amount of Nissl bodies and the degree of nuclear infolding. The results indicate that spiny fusiform cells are similar to mossy cells, another hilar cell type that receives its major synaptic input from axon collaterals of mossy fibers from granule cells. The distribution of the dendrites of spiny fusiform cells and the pattern of granule cell axon collaterals suggest a high degree of convergence from granule cells. In contrast, the variety of axodendritic synapses for sparsely spiny fusiform cells suggests that more diverse inputs affect this cell's activity. Therefore, the structure and circuitry of these two hilar cell types are probably different. This study adds further evidence to indicate that the hilus contains a large variety of cell types with different neuronal connections.     

Extranuclear estrogen receptor beta immunoreactivity is on doublecortin-containing cells in the adult and neonatal rat dentate gyrus

In adult female rats, estrogen receptor (ER) activation, particularly of ERbeta, promotes hippocampal neurogenesis. We previously reported that extranuclear ERbeta immunoreactivity (ir) in adult rats is on cellular profiles in or near the granule cell layer, which is the location of newly generated cells. During development, cells in or near the granule cell layer transiently express high levels of estrogen binding and nuclear ERs. Thus, we sought to determine if extranuclear ERbeta is in newly generated cells in adult and neonatal rat dentate gyrus. Sections from the dentate gyrus of adult proestrus or postnatal day 7 and 14 female rats were dual-labeled for ERbeta and the new-cell marker doublecortin (DCX) and examined by electron microscopy. DCX-containing neurons were found in the subgranular hilus in adult rats and were more widespread throughout the granule cell layer and hilus of neonatal rats. In both adults and neonatal rats, ERbeta immunoreactivity was found in a subset of DCX-labeled neurons. Electron microscopic examination of the adult dentate gyrus revealed that most perikarya with DCX-ir had the morphological characteristics of granule cells, although a few resembled interneurons. Dendrites with DCX-ir also were observed. In both adults and neonates, DCX-labeled neuronal perikarya and dendrites contained ERbeta-ir; ERbeta-ir usually was aggregated near the plasma membrane, mitochondria or endoplasmic reticula. ERbeta-ir was in glial profiles that apposed DCX-labeled perikarya and dendrites. These findings are consistent with data showing that estrogens can exert non-genomic effects directly and indirectly on newly generated cells in neonatal and adult rat dentate gyrus.     

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.     

Postnatal development of mossy cells in the human dentate gyrus: a light microscopic Golgi study.

Mossy cells in the human dentate gyrus of adults and children of different ages were impregnated using the rapid-Golgi method. In every case the cause of death was verified by autopsy and the brains were used when neither the history of the patient nor autopsy revealed brain-related disease. Mossy cells in the human share common light microscopic features with the same cell type in rats and monkeys. Their most characteristic feature is the extremely large and complex excrescences on their proximal dendrites. Distal dendrites display pedunculate spines. Mossy cells have a few somal spines. The axon of mossy cells originates from the cell body and gives rise to several collaterals in the hilar region. The axons could be followed for several hundred microns, but in only one case did an axon collateral enter the granule cell layer of the adult dentate gyrus. In the newborn child, mossy cells display immature somal and dendritic features. The soma frequently bear spines. The dendrites are varicose and terminate in presumed growth cones. Both proximal and distal portions of the dendrites bear a few pedunculate spines and long-irregular filopodia. A few small excrescences are present on the proximal dendrites. The first large, complex excrescences on the proximal dendrites of mossy cells appeared in the 7-month-old child. Both somata and dendrites display adult-like characteristics in mossy cells from a 5-year-old child. However, not all mossy cells are alike and some dendrites still display long filopodia. The axons of immature mossy cells were similar to adults. The present results indicate that connections between granule cells and hilar mossy cells of the human dentate gyrus develop through an extended postnatal period of time that may last until the fifth year.     

Intracerebral transplants of the rat fascia dentata: A Golgi/electron microscope study of dentate granule cells

In the present study we describe the morphological characteristics of dentate granule cells in intracerebral allografts of the rat fascia dentata. Blocks of hippocampal tissue containing the fascia dentata were taken from late embryonic and newborn rats and transplanted to the hippocampal region of other newborn and young adult rats. After survival periods of several months the recipient brains were fixed by perfusion and serially sectioned on a Vibratome. Some sections were stained with thionin to determine the localization and general histological organization of the transplants, while others were Golgi stained with a modification of the section Golgi technique. Well-impregnated transplant granule cells were gold-toned and deimpregnated thus allowing a correlated, light and electron microscopic analysis of identified neurons to be done. At the light microscopic level the morphology of the dentate granule cells in the transplants was very similar to Golgi-impregnated, gold-toned granule cells in the fascia dentata of normal rats (controls). A few irregular, more obliquely curved dendrites occurred, but basal dendrites passing into the hilar region were never observed. Following an initial spine-free segment granule cell dendrites were densely covered with spines. The axon, the mossy fiber, originated as usual from the basal pole of the cell body. In the electron microscope, both small and larger complex spines (v and w types) were seen to emerge from the gold-toned dendrites of the identified granule cells. The thin unmyelinated granule cell axons gave rise to giant mossy fiber boutons in the dentate hilus, but in addition numerous aberrant mossy fiber terminals were found innermost in the dentate molecular layer just above the granule cell layer. The results demonstrate that dentate granule cells that have gone through the major part of their differentiation-after transplantation develop characteristic dendritic and axonal elements very similar to those of granule cells in the fascia dentata in situ. The minor changes observed correspond to the redistribution of intrinsic connections that results from the absence of major extrinsic afferents.     

Subdivisions in the Multiple GABAergic Innervation of Granule Cells in the Dentate Gyrus of the Rat Hippocampus

The sources of GABAergic innervation to granule cells were studied to establish how the basic cortical circuit is implemented in the dentate gyrus. Five types of neuron having extensive local axons were recorded electrophysiologically in vitro and filled intracellularly with biocytin (Han et al., 1993). They were processed for electron microscopy in order to reveal their synaptic organization and postsynaptic targets, and to test whether their terminals contained GABA. (1) The hilar cell, with axon terminals in the commissural and association pathway termination field (HICAP cell), formed Gray's type 2 (symmetrical) synapses with large proximal dendritic shafts (n= 18), two-thirds of which could be shown to emit spines, and with small dendritic branches (n= 6). Other boutons of the HICAP neuron were found to make either Gray's type 1 (asymmetrical) synapses (n= 4) or type 2 synapses (n= 6) with dendritic spines. Using a highly sensitive silver-intensified immunogold method for the postembedding visualization of GABA immunoreactivity, both the terminals and the dendrites of the HICAP cell were found to be immunopositive, whereas its postsynaptic targets were GABA-immunonegative. The dendritic shafts of the HICAP cell received synapses from both GABA-negative and GABA-positive boutons; the dendritic spines which densely covered the main apical dendrite in the medial one-third of the molecular layer received synapses from GABA-negative boutons. (2) The hilar cell, with axon terminals distributed in conjunction with the perforant path termination field (HIPP cell), established type 2 synapses with distal dendritic shafts (n= 17), most of which could be shown to emit spines, small-calibre dendritic profiles (n= 2) and dendritic spines (n= 6), all showing characteristics of granule cell dendrites. The sparsely spiny dendrites of the HIPP cell were covered with many synaptic boutons on both their shafts and their spines. (3) The cell with soma in the molecular layer had an axon associated with the perforant path termination field (MOPP cell). This GABA-immunoreactive cell made type 2 synapses exclusively on dendritic shafts (n= 20), 60% of which could be shown to emit spines. The smooth dendrites of the MOPP cell were also restricted to the outer two-thirds of the molecular layer, where they received both GABA-negative and GABA-positive synaptic inputs. (4) The extensive axonal arborization of the dentate basket cell terminated mainly on somata (n= 26) and proximal dendrites (n= 9) in the granule cell layer, and some boutons made synapses on somatic spines (n= 6); all boutons established type 2 synapses. (5) The dentate axo-axonic cell established type 2 synapses (n= 14) exclusively on axon initial segments of granule cells in the granule cell layer, and on initial segments of presumed mossy cells in the hilus. The results demonstrate that granule cells receive inputs from the local circuit axons of at least five distinct types of dentate neuron terminating in mutually exclusive domains of the cell's surface in four out of five cases. Four of the cell types (HICAP cell, MOPP cell, basket cell, axo-axonic cell) contain GABA, and the HIPP cell may also be inhibitory. The specific local inhibitory neurons terminating in conjunction with particular excitatory amino acid inputs to the granule cells (types 1 – 3) are in a position to interact selectively with the specific inputs on the same dendritic segment. This arrangement provides a possibility for the independent regulation of the gain and long-term potentiation of separate excitatory inputs, through different sets of GABAergic local circuit neurons. The pairing of excitatory and inhibitory inputs may also provide a mechanism for the downward reseating of excitatory postsynaptic potentials, thereby extending their dynamic range.     

Dendritic caliber and the 3/2 power relationship of dentate granule cells

A quantitative examination of granule cell dendritic caliber and knowledge of dendritic lengths allows assessment of the distribution of dendritic membrane and the 3/2 power relationship at branch points. This paper presents caliber data of Golgi-impregnated rat dentate gyrus. We used camera lucida drawings of the dendritic trees of 15 dorsal leaf and 15 ventral leaf granule cells to quantify mean dendritic caliber, dendritic taper, the 3/2 power relationship of parent and sibling dendritic diameters at branch points, and surface area. First-order dendrites vary substantially in diameter. However, the mean caliber of all other dendrites is uniform across the proximal two-thirds of the molecular layer for the dorsal and ventral leaves. The average diameter here is 1 micron. More distally, only mean ventral leaf dendritic caliber declines. Granule cell dendritic taper is due primarily to caliber decreases at branch points and not to a gradual decline in diameter across the length of a dendritic segment. Comparing the parent segment diameter raised to the 3/2 power with the sum of the 3/2 powers of the two sibling segment diameters reveals, for the dendritic tree located within the distal two-thirds of the molecular layer, the desired 3/2 power relationship for the dorsal and ventral leaves. More proximally, where first-, second-, and third-order dendrites branch sequentially across a 60-100-micron extent, a 3/2 power relationship is not obtained. For the average dorsal leaf granule cell, dendritic surface area (without spines) is 11,984 micron2. The ratio of dendritic to somatic surface area is 28:1. Discussion of these data includes their implications for electrotonic modeling of the dentate granule cell.     

Interneurons in the Hippocampal Dentate Gyrus: an In Vivo intracellular Study

Interneurons in the dentate area were characterized physiologically and filled with biocytin in urethane-anaesthetized rats. On the basis of axonal targets the following groups could be distinguished. (i) Large multipolar interneurons with spiny dendrites in the deep hilar region densely innervated the outer molecular layer and contacted both granule cells and parvalbumin-positive neurons (hilar interneuron with perforant pathway-associated axon terminals; HIPP cells). (ii) A pyramidal-shaped neuron with a cell body located in the subgranular layer innervated mostly the inner molecular layer and the granule cell layer (hilar interneuron with commissural-associational pathway-associated axon terminals; HICAP cell). It contacted both granule cells and interneurons. Axon collaterals of HIPP and HICAP neurons covered virtually the entire Septo-temporal extent of the dorsal dentate gyrus. (iii) Calbindin-immunoreactive neurons with horizontal dendrites in stratum oriens of the CA3c region gave rise to a rich axon arbor in strata oriens, pyramidale and radiatum and innervated almost the entire extent of the dorsal hippocampus, with some collaterals entering the subicular area (putative trilaminar cell). (iv) Hilar basket cells innervated mostly the granule cell layer and to some extent the inner molecular layer and the CA3c pyramidal layer. HIPP and trilaminar interneurons could be antidromically activated by stimulation of the fimbria. Only the HICAP cells could be monosynaptically discharged by the perforant path input. All interneurons examined showed phase-locked activity to the extracellularly recorded theta/gamma oscillations or to irregular dentate electroencephalogram spikes. These observations indicate that the interconnected interneuronal system plays a critical role in coordinating population activity of the dentate gyrus and Ammon's horn.