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Afterdischarge thresholds and kindling rates in dorsal and ventral hippocampus and dentate gyrus.

Electrodes were implanted to dorsal hippocampus (CA1), ventral CA1, DOrsal dentate gyrus or ventral dentate gyrus. Epileptiform afterdischarge (AD) thresholds were lower in dorsal areas than in ventral areas. Dorsal areas, however, required a greater number of stimulations to develop ("kindle") a fully generalized convulsion than did ventral areas. Thresholds and kindling rates in the dentate gyrus were intermediate between dorsal and ventral CA1, except for the ventral dentate which had higher AD thresholds than ventral CA1. Secondary sites within the hippocampus subsequently kindled within a few stimulations following completion of kindling in the primary site, regardless of which hippocampal area served as the primary site.

Animals

The morphology of the hippocampus and dentate gyrus in normal and reeler mice.

The morphology of the hippocampus and dentate gyrus in normal and reeler mice has been studied in Nissl, myelin, Golgi, Timm's sulfide silver and gold chloride-sublimate preparations. It is evident from both cell-and fiber-stained sections that despite the obvious defect in the positioning of the hippocampal pyramidal and dentate granule cells in the reeler mouse within the radial dimension, the hippocampal formation as a whole shows a normal and consistent progression of cytoarchitectonic fields along its transverse axis, and a normal and consistent progression of changes in the structure of the hippocampus and dentate gyrus along their longitudinal axes. Thus, at least in these structures, the reeler gene seems to exert its effect only in the radial dimension. Cell counts in the area dentata indicate that the number of dentate granule cells in the reeler mouse is reduced compared to that found in normal or heterozygous animals. Although it has been known for some time that the number of granule cells in the reeler cerebellar cortex is markedly reduced, this appears to be the first evidence for a reduction in cell number in a forebrain structure. All the major cell types normally found in the hippocampus and the dentate gyrus are recognizable in Golgi-stained preparations from the brains of reeler mutants. However, in both regions there are a number of abnormalities in the appearance of the cells which seem to be related to the cellular ectopia. Thus, whereas most of the pyramidal and granule cells which attain a normal position in the mutant usually have normal, or near-normal dendritic arbors, the dendrites of nearly all ectopic cells are severely distorted, both in their orientation and general configuration. In preparations stained by the Timm's sulfide silver technique it is evident that the general lamination pattern seen in normal mice is retained in the reeler hippocampus and dentate gyrus despite the gross malpositioning of many of the relevant neurons. However, although the overall laminar arrangement is preserved, there are some fairly consistent abnormalities; for example, the normal trilaminar staining pattern seen in the stratum moleculare of the dentate gyrus is replaced in the reeler by a bilaminar pattern. In gold chloride-sublimate impregnated preparations there is no obvious alignment of the astrocytes in the stratum moleculare of the dentate gyrus in either normal or reeler mice. Moreover, the distribution of the astrocytes within this zone is fairly normal in the reeler mouse, although, in general, these cells appear to be more consistently stellate in form than in normal animals.

Animals

The development of the hippocampus and dentate gyrus in normal and reeler mice.

The histogenesis, the time of origin and the pattern of migration of the cells in the hippocampus and dentate gyrus, have been studied in normal and reeler mice. The earliest indication of a defect in the reeler hippocampus is seen on the fifteenth embryonic day (E15) which is at least 24 hours after the first indication of a defect in the neocortex. It is not until E18, that the dentate gyrus shows signs of its incipient abnormality. It appears then, that in both the hippocampus and the dentate gyrus the gene defect first manifests itself at the stage at which the definitive cellular layers are assembled. Experiments involving the injection of 3H-thymidine (3H-TdR) at different developmental stages have confirmed that the site and rate of cellular proliferation in the reeler hippocampus and dentate gyrus are normal, as is the initial pattern of cell migration. However, in the reeler dentate gyrus, most postnatal cell proliferation occurs ectopically and in the hippocampus the normal "inside-out" sequence of neurogenesis is reversed, the earliest pyramidal cells generated coming to lie superficially within the stratum pyramidale and the later formed cells being added at progressively deeper levels. There is no discernible gradient in the time of origin of the granule cells in the radial dimension of the reeler dentate gyrus, whereas there is an obvious "outside-in" gradient in the normal animal. The characteristic gradients in cell proliferation seen in the transverse and longitudinal dimensions of the normal dentate gyrus are, however, also evident in the reeler mouse. Taken together, these observations suggest that the reeler gene exerts its effect on neuronal position only in the radial dimension, and does so at a stage of development subsequent to the proliferation and initial migration of the relevant neurons. Timm's sulfide silver preparations indicate that the characteristic staining patterns seen in the dentate gyrus and hippocampus appear at the same time, and mature at the same rate in normal and reeler mice.

Animals

The organization of certain afferents to the hippocampus and dentate gyrus in normal and reeler mice.

The organization of certain of the major afferents to the hippocampus and dentate gyrus has been studied in normal and reeler mutant mice using the autoradiographic and the anterograde degeneration methods. The distribution of the hippocampal and dentate afferents which arise in the medial and lateral parts of the entorhinal cortex and the hippocampus of both sides, has been found to be generally similar to that previously described in the rat, but there are a few minor differences that are discussed in the text. Despite the marked ectopia of many of the neurons in the hippocampal formation in the reeler mouse, the principal afferents to the hippocampus and the dentate gyrus maintain many of the features seen in normal mice. In particular, they maintain a normal radial sequence and a characteristic laminated and complementary arrangement. However, there are a number of significant differences in their distribution; for example, in the reeler mouse, the entorhinal afferents occupy the entire radial extent of the stratum moleculare of the dentate gyrus, whereas in normal mice they are restricted to the outer four-fifths of this layer. Furthermore, in the mutant the commissural and associational afferents to the dentate gyrus do not occupy the inner one-fifth of the molecular layer (as they do in normal animals) but rather are spread throughout the zone containing granule cells, which includes both the poorly-defined stratum granulosum and most of the hilar region of the dentate gyrus. Some of the developmental and functional implications of these and other abnormalities in the organization of the afferents to the hippocampus and dentate gyrus are discussed.

Animals

A neurophysiological analysis of commissural projections to dentate gyrus of the rat.

The electrophysiological properties of the commissural projections to the dentate gyrus of the rat were investigated using extracellular field-potential and unit-recording techniques. The following conclusions with respect to those investigations were obtained: 1) The CA3c/CA4 region of the contralateral hippocampus proved to be the most effective site for eliciting the commissural field potentials in the dentate gyrus dorsal and ventral leaves. 2) The location of the short-latency negative field potential in the molecular layer of the dentate gyrus was restricted to a region 50-100 mum distal to the granule cell layers corresponding to the inner one-third of the granule cell dendrites. 3) The negative field potential proved to satisfy a number of criteria for the extracellular representation of the summed EPSPs of synchronously activated granule cells. 4) The excitatory nature of the commissural projections to the dentate was confirmed by the short-latency driving of units recorded from the granule cell layers. 5) A comparison of both commissural and entorhinal cortical stimulation procedures showed the field potentials elicited by the different convergent anatomical systems to be localized within different regions of the dentate molecular layer. 6) The distribution of commissural potentials along the septotemporal axis of the dentate gyrus indicated that stimulation sites homotopic to the recording electrode in the contralateral CA3c/CA4 region were the most effective in eliciting these potentials. 7) These findings were discussed with reference to the mode of activation of the dentate granule cells by the commissural system with specific comparison to the larger and apparently more powerful projections from the entorhinal cortex.

Animals

The postnatal development of rat dentate gyrus and the effect of early thyroid hormone treatment.

The postnatal development of the dentate gyrus and the effect of 5 microgram/day triiodothyronine treatment was examined by means of tritiated thymidine autoradiography. The polymorph layer of the dentate gyrus is a secondary germinal layer and forms cells for the granular layer. The cell formation in the dentate gyrus was found to be nearly completed on the 21st day, but development ended only in adulthood. Thyroid-hormone treatment did not affect significantly the postnatal cell formation of the dentate gyrus, but resulted in the appearance of a great number of pycnotic cells in the granular layer.

Aging

Potentiation of excitatory synaptic transmission in the normal and in the reinnervated dentate gyrus of the rat.

Following destruction of the ipsilateral temporo-ammonic tract, which originates in the entorhinal cortex, and terminates on the granule cells of the dentate gyrus, fibers from the surviving contralateral entorhinal area proliferate forming extensive new connections with the denervated dentate granule cells. Utlizing extracellular recording techniques, we have compared the characteristics of synaptic transmission in the lesion induced afferents with the characteristics of the normal ipsilateral afferents by analyzing the responses of dentate granule cells to paired pulse activation of temporo-dentate circuitry. In the dentate gyrus of the normal rat, and extracellularly recorded EPSP evoked by stimulation of the ipsilateral entorhinal cortex is enhanced by as much as 100% by a "conditioning" pulse to the same afferent system. This is called paired pulse potentiation. In the reinnervated dentate gyrus, the extracellular EPSP evoked by a test stimulus delivered to the contralateral entorhinal cortex is also potentiated by a conditioning pulse. Thepaired pulse potentiation in the reinnervated dentate gyrus has a time course which is comparable to that of the normal ipsilateral afferent system, but the magnitude of the potentiation is somewhat less, averaging approximately 140% of control...

Afferent Pathways

Polysynaptic activation of the dentate gyrus of the hippocampal formation: an olfactory input via the lateral entorhinal cortex.

The possibility that olfactory input is transmitted to specific subregions of the hippocampal formation via the entorhinal cortex was investigated electrophysiologically by analyzing the laminar profiles of potentials evoked in the hippocampal formation by stimulation of the lateral olfactory tract (LOT). LOT stimulation resulted in long latency (14--20 ms) evoked responses in the dentate gyrus of the hippocampal formation ipsilateral to the stimulation. The variable long latency of these responses and their inability to follow stimulus rates of 40/s suggested that these potentials reflected polysynaptic activation. Analysis of the laminar profiles of the evoked potentials indicated that the responses originated from a synaptic field localized in the outer portion of the stratum moleculare of the dentate gyrus, a terminal distribution which overlaps that of the lateral entorhinal cortical (LEC) projection to the dentate gyrus. Lesions of the LEC eliminated the long latency responses in the dentate gyrus evoked by LOT stimulation. In addition, a conditioning pulse delivered either to the LOT or to the LEC produced paired pulse potentiation of the response elicited by subsequent stimulation of the other structure. No evidence was found to indicate that responses were generated in regio superior of the hippocampus proper following LOT stimulation. Taken together, these results suggest that stimulation of the LOT activates the dentate gyrus of the hippocampal formation by multisynaptic pathways which relay through the lateral portion of the entorhinal area. This finding is discussed with regard to entorhinal cortical organization and the known olfactory projections to the LEC.

Animals

A note on the distribution of glial cells in the molecular layer of the dentate gyrus.

The distribution of the perikarya of astrocytes and other glial cells in the molecular layer of the dentate gyrus has been studied in gold chloride-sublimate preparations of rats and of normal and reeler mice, and in plastic embedded material from young adult rats. Contrary to previous reports (Rose et al. [7]), we have found no evidence for a distinct "line" or "band" of astrocyte cell bodies along the interface between the zones of termination of the entorhinal and hippocampal afferents to the dentate gyrus. Indeed, apart from a conspicuous accumulation of astrocytes immediately beneath the pial surface and hippocampal fissure, the distribution of glial cells across the extent of the molecular layer appears to be more or less random. In view of this it is difficult to ascribe a critical role to the astrocytes in either determining the normal distribution of afferent fibers to the dentate gyrus or in promoting their re-organization following its partial deafferentation.

Afferent Pathways

Loss of axosomatic synapses in the dentate gyrus of aged rats.

Axosomatic synapses involving granule cells of the dentate gyrus were studied by means of quantitative electron microscopic analysis in young adult (3-month-old) and aged (25-month-old) rats. The number of axosomatic synapses per unit length of neuronal soma membrane was found to be significantly lower (by 15%) in aged animals than in young adults. This decrease in synaptic numbers is not associated with age-related changes in the size of neuronal soma profiles or in the length of their plasma membranes. The ratio between the total length of synaptic appositions and the membrane length of a neuronal soma profile was diminished by 22% in aged rats, whereas the mean length of synaptic apposition was 10% less in these animals than in young adults. These data, taken together, suggest that an absolute loss of axosomatic synapses occurs with advanced age. It appears, therefore, than not only the loss of axodendritic synapses, described previously, but also the loss of axosomatic synapses, found here, contributes to the process of age-related partial deafferentation of neurons in the rat dentate gyrus.

Afferent Pathways

Evidence for an input to the molecular layer and the stratum granulosum of the dentate gyrus from the supramammillary region of the hypothalamus.

Injections of a mixture of tritiated amino acids were made into the posterior hypothalamus in a series of rats and cats. In every case in which the injection involved a significant proportion of the cells in the supramammillary region, labeled fibers could be followed to the dentate gyrus, the anterior hippocampal rudiment and the induseum griseum of both sides. In the dentate gyrus the hypothalamic afferents terminate in a narrow band in the outer half of the stratum gramulosum and the inner 20 micron or so, of the stratum moleculare, immediately deep to the zone of termination of the associational and commissural afferents. As judged by silver grain counts across the width of the zone of labeled terminals, the projection to the ipsilateral side is several times as heavy as that to the contralateral side, and although it involves the entire septo-temporal (=rostro-caudal) extent of the gyrus on both sides, the projection to the suprapyramidal (inner) blade of the dentate gyrus is approximately twice as heavy as that to the infrapyramidal (outer) blade.

Amino Acids

Amino acids and the synaptic pharmacology of granule cells in the dentate gyrus of the rat.

Granule cells in the dentate gyrus in the hippocampi of anaesthetized rats were excited by stimulation of the contralateral hippocampus (the commissural input) and the ipsilateral entorhinal cortex (the perforant path). The cells were also activated by the electrophoretic administration of various amino acids. A selective antagonism of glutamate and perforant path excitations was obtained with glutamic acid diethylester, and of aspartate and other amino acid induced and commissural excitations with D- or DL-alpha-aminoadipate. An excitatory effect of alpha-aminoadipate which was sometimes observed was prevented by the gamma-aminobutyric acid antagonist bicuculline, and may be a disinhibitory phenomenon. The results lend support to the proposition that the transmitter of the perforant path is glutamate while that of the commissural fibres is aspartate.

2-Aminoadipic Acid

Glutamate secretion and NAD(P)H levels during calcium-dependent depolarization of slices of the dentate gyrus.

Evidence from studies involving release, postsynaptic responses, inactivation, storage and synthesis etc. support the contention that glutamate may be the transmitter of the perforant input to the granule cells in the dentate gyrus of the hippocampus. In the present report the release of endogenous glutamate and the levels of reduced pyridine nucleotides (NAD(P)H) has been measured in parallel experiments on slices from the dentate gyrus of the hippocampus. A Ca-dependent release of glutamate is evoked by tissue depolarization caused either with electrical field stimulation or with elevated KC1. Electrical stimulation induced a transient increase in tissue NAD(P)H levels, the increase being inhibited by approximately 50% during Ca-free conditions. KC1 stimulation, on the other hand, produced a long-lasting decrease in NAD(P)H, the decrease being halved in the absense of Ca. A metabolic relation between stimulus secretion and energy utilization is discussed.

Animals

Development of potentiation in the dentate gyrus of rat: physiology and anatomy.

The physiological development of potentiating processes in the rat dentate gyrus were compared to morphological development. Rapid Golgi techniques were coupled with in vitro studies of dentate granule cell frequency potentiation, post-tetanic potentiation and long-term potentiation. Frequency potentiation and long-term potentiation exhibited a developmental progression between 7 and 210 days postnatal. Posttetanic potentiation remained constant across this period. The relation of these findings to synaptogenesis and dendritic spine formation are discussed.

Animals

Development of habituation in the dentate gyrus of rat: physiology and anatomy.

The physiological development of monosynaptic response habituation in the rat dentate gyrus was compared to morphological development. Rapid Golgi techniques were coupled with in vitro studies of dentate granule cell habituation to several frequencies and intensities of monosynaptic excitation. Except for the youngest group, the degree of habituation increased as a function of age, paralleling the morphological development.

Animals

A study of glial cell proliferation in the molecular layer of the dentate gyrus of the rat following interruption of the ventral hippocampal commissure.

The proliferation of glial cells in the molecular layer of the dentate gyrus in response to lesions of the ventral hippocampal commissure, has been studied autoradiographically following intraventricular injections of 3H-thymidine. Within 24 h of commissurotomy there is an appreciable increase in the number of labeled cells throughout the molecular layer which reaches its peak at approximately 36 h. This generalized glial hyperplasia persists for at least 5--6 weeks and there does not appear to be a secondary re-distribution of the newly-generated glial cells as has been reported after entorhinal lesions (Gall et al., 1979). In semi-thin plastic sections most of the proliferating cells more closely resemble the "medium-shade oligodendrocytes" of Ling et al. (1973) than typical microglia; the reactive astrocytes do not appear to participate in the glial proliferation.

Animals

Terminal proliferation in the partially deafferented dentate gyrus: time courses for the appearance and removal of degeneration and the replacement of lost terminals.

The time courses for the appearance and removal of degenerating terminals and the loss and reappearance of intact terminals were investigated in the partially denervated inner molecular layer of the dentate gyrus of the adult rat. Dense degeneration was evident in the neuropil within 26 hours following contralateral hippocampectomy. These profiles increased rapidly in number until the maximal degree was reached at two to three days postlesion, after which the degenerating terminals were quickly removed from the neuropil. A more rapid rate of removal occurred during the 3-to 5-day survival period than from 6 to 50 days postlesion. The intact terminal population dropped 35% within two days of the lesion and remained at this level until six to eight days postlesion when the number began to steadily increase. The time course for this reappearance can be divided into two phases: a period of rapid terminal addition from 6 to 15 days followed by a phase of slower acquisition. This recovery continued until the normal synaptic density was regained by 50 to 65 days postlesion. These results indicate that a substantial proportion of degenerating endings are removed well in advance of the time at which terminal proliferation begins, suggesting that certain changes other than merely the removal of competitive inputs must take place prior to growth of new terminals. Possible explanation suggested by the present results for the delay in the onset of sprouting include: (1) an absence of appropriate postsynaptic targets during the 2-to 5-day postlesion period and (2) inhibition of axonal growth by the glial cells which are phagocytizing the degenerating products. Beyond the sixth postlesion day the rate at which new terminals appear does correlate with the rate at which degeneration is removed. This suggests that once underway the time course for sprouting may be determined by the avaiabliity of postsynaptic sites.

Afferent Pathways

Dipole-like neuronal sources of theta rhythm in dorsal hippocampus, dentate gyrus and cingulate cortex of the urethane-anesthetized rat.

Spatial distribution of theta activity was investigated in the dorsal hippocampal formation and overlying neocortex of the urethane-anesthetized rat. Laminar phase profiles from semi-microelectrode penetartions showed approximately 180 degrees phase shifts combined with small amplitude values in stratum radiatum of CA1, instratum moleculare of the dentate gyrus and in layer V/VI of the cingulate cortex at theta peak frequency. Evidence has been presented that layers of neurons in CA1, in the dorsal granular layer and in the cingulate cortex are the sources of dipole-like theta field potentials. A strong linear relationship between the neuronal theta sources in hippocampal CA1, dentate area and cingulate cortex was found.

Animals