Neurogenesis of glutamic acid decarboxylase immunoreactive cells in the hippocampus of the mouse. II: Area dentata

The temporal patterns of neurogenesis of cells showing glutamic acid decarboxylase (GAD) immunoreactivity were determined in the area dentata of the mouse. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E11 through E17 (E0 being the day of mating). The distribution of (3H)thymidine-labeled, GAD-positive neurons in the hilus and in the different strata of the fascia dentata (stratum infragranulosum, stratum granulosum, stratum moleculare) were recorded in adult animals. A radial gradient of neurogenesis of GAD-positive cells in the area dentata was not apparent. In the transverse axis, neurogenesis of GAD-positive cells seemed to follow a faint suprapyramidal to infrapyramidal gradient, which was due to differential timing of neurogenesis of GAD-positive cells destined for the stratum infragranulosum of the suprapyramidal and infrapyramidal blades of the fascia dentata. GABAergic neurons in the fascia dentata comprise a limited number of well-defined cell types. All of the different morphologic types of GAD-positive neurons present in the area dentata were generated prenatally. These diverse forms did not have specific times of neurogenesis. These results support the concept that the adult morphology of GAD-positive cells in the area dentata of the mouse do not bear any relationship to their times of origin.     

Times of generation of glutamic acid decarboxylase immunoreactive neurons in mouse somatosensory cortex

The birth dates of neurons showing glutamic acid decarboxylase (GAD) immunoreactivity have been determined in mouse somatosensory cortex. Pregnant C57Bl mice received pulse injections of (3H)thymidine from E10 through E17 (E0 being the day of mating). The distributions of thymidine-labeled, GAD-positive and nonimmunoreactive (non-GAD) cells as a function of depth under the pial surface were recorded in adult animals. The maximum rate of generation of GAD-positive neurons occurred at E14, whereas the generation of non-GAD neurons reached its maximum rate at E13. Except for those in layer I, GAD-positive neurons followed an inside-out sequence of positioning. GAD-positive neurons born at E12 and E13 were located in layers VI-IV. GAD-positive neurons born at E14 were found throughout the cortical thickness, with a maximum in layer IV. The GAD-positive neurons labeled after pulses at E15 or E16 or E17 were limited to the superficial strata, forming a band that became narrower as it moved toward the pial surface with increase in age of pulse labeling. GAD-positive neurons in layer I were generated at a constant rate during the whole embryonic period analyzed. Non-GAD neurons also followed an inside-out spatiotemporal gradient. Two partially overlapping phases were distinguished in non-GAD neurogenesis. During the first phase (from E12 to E14) neurons populating adult layers VI and V originated, while neurons located in layers IV through I were generated during the second phase (from E13 to E17). Since GAD-immunoreactive neurons form a heterogeneous population, we envisage further studies in order to test whether differences exist in birth dates among the classes.     

The distribution of acetylcholinesterase in the hippocampal formation of the monkey

In order to study the distribution of acetylcholinesterase (AChE) in the primate hippocampal formation, we have stained serial sections through the brains of nine macaque monkeys for AChE by two variants of the Koelle acetylthiocholine method. We have found a distinctive pattern of staining in the hippocampal formation which varies in intensity both from region to region, and along rostrocaudal and radial gradients within each region. In the dentate gyrus, there is intense staining of the inner one-third of the molecular layer with much lighter staining in the rest of the molecular layer except for a moderately stained band at its outer edge. In the caudal half of the dentate gyrus, the inner portion of the molecular layer is less intensely stained though there is a distinctly denser band of staining just above, and partly within, the superficial margin of the granule cell layer. The granule cells are unstained but there are AChE-positive fibers which run through the granule cell layer to the molecular layer. The hilar region of the dentate gyrus has a narrow band of heavy staining (which corresponds to an acellular layer in Nissl-stained sections) just subjacent to the granule cell layer; the remainder of the hilus, where most of the hilar cells reside, is less intensely stained and at caudal levels is almost entirely unstained. In the regio inferior of the hippocampus, there is intense staining of the stratum oriens which extends into the pyramidal cell layer; the stratum radiatum and the stratum lacunosum- moleculare are also stained and here the staining pattern shows some degree of stratification. By contrast, most of the alveus, the pyramidal cell somata, and the layer of mossy fibers (stratum lucidum) are unstained. The border region between regio inferior and regio superior of the hippocampus (field CA2 of Lorente de No, '34) is especially heavily stained. This contrasts markedly with regio superior, which is more lightly stained than regio inferior. Stratum oriens and stratum radiatum of regio superior have a more evenly distributed pattern of staining, though the intensity of staining increases sharply at the border with the subiculum. Stratum lacunosum- moleculare is only lightly stained throughout much of the transverse extent of regio superior but there is also a conspicuous and constant patch of heavier staining at the border with the subiculum.(ABSTRACT TRUNCATED AT 400 WORDS)     

Distribution of acetylcholinesterase in the hippocampal region of the mouse: II. Subiculum and hippocampus.

The distribution of acetylcholinesterase (AChE) was examined in the subiculum and hippocampus of the adult mouse (Mus musculus domesticus). A distinctly stratified AChE pattern was observed in both areas and was compared in detail with cytoarchitectural fields and layers. In the subiculum, the lateral plexiform layer was lightly stained superficially and moderately stained at depth, where it abutted the lateral, moderately stained cell layer. Medially, a moderately stained deep plexiform layer separated the darkly stained superficial plexiform layer from the equally AChE-intense cell layer. At depth, the subicular cell layer was delimited by a band of very high AChE activity. In regio superior of the hippocampus, AChE-intense bands delimited the moderately stained strata moleculare, radiatum, and oriens toward the subjacent layers. In the stratum pyramidale, precipitate insinuated between the cell bodies gave a dark appearance to the deep part of the layer. The homologous strata of regio inferior appeared darker, but the relative staining intensities corresponded largely to those in regio superior. AChE activity in the layer of mossy fibers was almost absent septally but increased gradually to very high levels temporally. The AChE staining pattern, in conjunction with cytochemical and morphological evidence, strongly suggests a division of the pyramidal cell layer of the mouse and rat into superficial and deep substrata and discourages the definition of a prosubiculum in rodents. A comparative analysis of the AChE pattern reveals that: 1) in the subiculum, differences between species are observed within a generalized pattern of medial darkly staining and lateral lightly staining portions; 2) in the hippocampus, a conservation of the AChE pattern is seen in strata associated with intrinsic hippocampal connection; while 3) numerous interspecific differences are found in the stratum moleculare.     

Commissural afferents to the rat hippocampus terminate on vasoactive intestinal polypeptide-like immunoreactive non-pyramidal neurons. An EM immunocytochemical degeneration study

In the rat hippocampus, bipolar non-pyramidal neurons in stratum radiatum and stratum oriens and multipolar neurons in stratum lacunosum-moleculare react for vasoactive intestinal polypeptide (VIP) immunostaining, but pyramidal cells do not. Such bipolar VIP-like immunoreactive neurons in strata radiatum and oriens of regio superior were studied by electron microscopy for synaptic contacts with commissural afferents. The commissural fibers were identified by their anterograde degeneration induced by contralateral fimbria transections 2 days before sacrifice. Electron-dense degenerated boutons of commissural origin were found in synaptic contact with the cell bodies and dendrites of the VIP-like immunoreactive non-pyramidal cells.     

Immunocytochemical localization of glutaminase-like and aspartate aminotransferase-like immunoreactivities in the rat and guinea pig hippocampus

There is considerable evidence that pathways of the hippocampus use an excitatory amino acids as transmitter. We have attempted to immunocytochemically identify excitatory amino acid neurons in the hippocampus of the rat and guinea pig using antiserum to glutaminase and antiserum to aspartate aminotransferase, which have been proposed as markers for aspartergic/glutamergic neurons. Glutaminase-like immunoreactivity was seen in granule cells in the dentate gyrus and fibers and puncta associated with the mossy fiber pathway in the hilus and stratum lucidum of the hippocampus. At the ultrastructural level, glutaminase-like immunoreactivity was observed in mossy fiber terminals in the stratum lucidum. Glutaminase-like immunoreactivity was also seen in pyramidal cells in regio inferior and regio superior and in cells in layer two of the entorhinal cortex. Schaffer collateral terminals, commissural fiber terminals and perforant pathway terminals were not seen at the light microscopic level. Glutaminase-like immunoreactivity is thus found in the cell bodies of proposed excitatory amino acid neurons of hippocampal pathways, but does not appear to label all terminals. Aspartate aminotransferase-like immunoreactivity was not seen in any cells, fibers or terminals in the rat or guinea pig hippocampus.     

Cholecystokinin-, enkephalin-, and substance P-like immunoreactivity in the dentate area,hippocampus, and subiculum of the domestic pig

The distribution of cholecystokinin-like, enkephalin-like, and substance P-like immunoreactivities is described in the dentate area, hippocampus, and subiculum of the domestic pig (Sus scrofa domesticus) as a baseline for future experimental studies. The distributions in the pig are compared with previous observations in other species. Cholecystokinin-like immunoreactive nerve cell bodies were intensely stained and present in large numbers in all subfields studied. Cholecystokinin-like immunoreactive terminals appeared as stained puncta, whereas fibers were only rarely encounterd. The puncta were mainly seen in the dentate molecular layer and dentate granule cell layer, the pyramidal cell layer of the hippocampal regio inferior, stratum moleculare of the hippocampal regio superior, and in the subiculum. Enkephalin-like immunoreactive nerve cell bodies were faintly stained and generally present in very small numbers, except for some pyramidal cells in the subicular cell layer. Enkephalin-like immunoreactive fibers were few in number, whereas stained puncta appeared with variable densities. Puncta of particularly high densities were found in the dentate molecular layer, whereas they appeared of moderate density in the dentate hilus, stratum moleculare of the hippocampal regio superior, and in the subiculum. Substance P-like immunoreactive nerve cell bodies were few and very faintly stined. They primarily occurred in the dentate hilus, stratum oriens of the hippocampus, and in the subicular cell layer. Stained fibers were few in number, whereas stained puncta were present in abundant numbers corresponding to the mossy fiber projection in the dentate hilus and the layer of mossy fibers of the hippocampal regio inferior, and in moderate numbers in stratum moleculare of the hippocampal regio superior and in the subiculum. For all three neuropeptides there were consistent and very characteristic variations in the distribution of immunoreactivity along the septotemporal axis of the hippocampus. When viewed in a comparative perspective the distribution of enkephalin-like and substance P-like terminals in the domestic pig displayed striking differences from the basic pattern observed in other species. This contrasted with the distribution of cholecystokinin-like neurons and terminals, which resembled more closely these species. © 1993 Wiley-Liss, Inc.     

Pattern of neuronal vulnerability in the cat hippocampus after one hour of global cerebral ischemia

Summary The dorsal hippocampus of cat was investigated by light microscopy and immunohistochemistry following 1 h global cerebral ischemia and various recirculation times from 1 day to 1 year. Complete ischemia was produced by combining hypotension with intrathoracic occlusion of major arteries. Postischemic resuscitation was carried out using an intensive care regimen with continuous neurophysiological monitoring. Brains of controls (n=4) and postischemic animals (n=12) were fixed in formaldehyde and prepared for histology and immunohistochemistry of glial fibrillary acidic protein (GFAP). In all post-ischemic animals the hilus and the regio superior of dorsal hippocampus which encompasses the CA1 subfield were severely damaged. Neurons in these regions exhibited the typical sequela of neuronal death. GFAP staining revealed vivid astroglial proliferation in stratum lacunosum-moleculare and stratum oriens. Changes in the regio inferior of dorsal hippocampus, i.e., CA3 subfield, and in dentate gyrus granular layer, were variable. Although most animals exhibited moderate to severe neuronal and glial alterations, groups of surviving cells were observed in the stratum oriens and in the granular layer of dentate gyrus. In one animal the majority of CA3 pyramidal cells and granule cells was preserved. These findings demonstrate that after 1 h of complete cerebral ischemia dorsal hippocampus exhibits two different types of injury: a consistent pattern of selective vulnerability in the hilus and the regio superior, and a variable pattern of non-selective injury in the regio inferior and dentate gyrus. The two patterns can be best explained by intrinsic (pathoclitic) and extrinsic (hemodynamic/edema) factors, respectively and are likely to represent basically different mechanisms of ischemic injury.     

An EM autoradiographic study of the hypothalamo-hippocampal projection

Previous studies have shown that in many different mammals there is a small but distinct projection from the supramammillary region in the caudal hypothalamus to the junctional region between the regio superior and regio inferior of the hippocampus. We have analyzed the mode of termination of this hypothalamo-hippocampal projection in the rat by electron microscopic (EM) autoradiography following injections of [3H]proline into the caudal hypothalamus. The projection is confined to the regio inferior where it is centered over the subicular end of field CA3, but also spans the adjoining region, field CA2. In our material the highest densities of labeling have been seen over the deeper part of the pyramidal cell layer and in the adjacent stratum oriens but, in addition, above background levels of labeling have been found superficial to the pyramidal cell layer in the stratum lucidum and the deeper part of the stratum radiatum. Most of the labeled synapses appear to be on the perikarya and primary dendrites of the hippocampal pyramidal cells, but some axo-spinous contacts have also been seen. All the labeled boutons contained clear, spheroidal synaptic vesicles and made asymmetric, Type I, contacts with their targets.     

Prolonged sojourn of developing pyramidal cells in the intermediate zone of the hippocampus and their settling in the stratum pyramidale

In radiograms of rat embryos that received a single dose of [3H]thymidine between days E16 and E20 and were killed 24 hours after the injection, the heavily labeled cells (those that ceased to multiply soon after the injection) form a horizontal layer in the intermediate zone of the hippocampus, called the inferior band. The fate of these heavily labeled cells was traced in radiograms of the dorsal hippocampus in embryos that received [3H]thymidine on day E18 and were killed at different intervals thereafter. Two hours after injection the labeled proliferative cells are located in the Ammonic neuroepithelium. The heavily labeled cells that leave the neuroepithelium and aggregate in the inferior band 1 day after the injection become progressively displaced toward the stratum pyramidale 2-3 days later, and penetrate the stratum pyramidale of the CA1 region on the 4th day. In the stratum pyramidale of the CA3 region, farther removed from the Ammonic neuroepithelium, the heavily labeled cells are still sojourning in the intermediate zone 4 days after labeling. Observations in methacrylate sections suggest that two morphogenetic features of the developing hippocampus may contribute to the long sojourn of young pyramidal cells in the intermediate zone: the way in which the stratum pyramidale forms and the way in which the alveolar channels develop. The stratum pyramidale of the CA1 region forms before that of the CA3 region, which is the reverse of the neurogenetic gradient in the production of pyramidal cells. We hypothesize that this is so because the pyramidal cells destined to settle in the CA3 region, which will be contacted by granule cells axons (the mossy fibers), have to await the formation of the granular layer on days E21-E22. Concordant with this is the observation that the hippocampal intermediate zone, which contains the sojourning young pyramidal cells, greatly enlarges between days E16 and E20, then suddenly diminishes and disappears by day E22. The other factor that may contribute to the prolonged sojourn of pyramidal cells, specifically those destined to settle in the CA1 region, is the pattern of alveolar channel development. This transient extracellular matrix begins to form several days after the onset of pyramidal cell neurogenesis, grows in a direction opposite to the settling of pyramidal cells in the stratum pyramidale, and does not reach the subicular end of Ammon's horn until day E21.(ABSTRACT TRUNCATED AT 400 WORDS)     

Some intrinsic connections of the hippocampus in the rat: an experimental analysis

Some internal pathways of the hippocampus were mapped in adult rats using Fink-Heimer silver impregnation methods for the demonstration of anterograde axon degeneration. Cases with lesions of regio superior of the Ammon's horn showed that regio superior projects onto the subiculum by way of fibers in the alveus. The termination is most dense in the portion of the subiculum near the presubiculum. The fibers exhibit an orderly arrangement, in that the dorso-ventral level of the subicular degeneration depends on the level of the regio-superior lesion. No evidence of connections from regio superior to regio inferior and area dentata appeared in this study. The terminal fields of ipsilateral fibers from regio inferior to regio inferior and superior are pervaded by commissural fibers. This precludes selective destruction of the ipsilateral fibers in normal rats. In order to investigate these latter ipsilateral fibers, special animals were used in which the commissures had been transsected at the age of eight days. Secondary lesions were made in the hippocampus of the adults when no stainable commissural degeneration was demonstrable. The results obtained from these animals were the following. Lesions of regio inferior cause degeneration both in this subfield and in regio superior. The fibers terminate in the stratum oriens and radiatum, and not in the stratum lacunosum-moleculare or on pyramidal cell bodies. The total spread of fibers along the longitudinal axis of the hippocampus is 4–5 mm, being slightly less in the regio inferior than in the regio superior. When the lesion involves only the dentate area and the adjacent part of the regio inferior, the degeneration in the regio superior is most pronounced superficially in the stratum radiatum and is absent from the stratum oriens. The fibers remaining within the regio inferior have a special mode of termination: dorsal to the lesion, degeneration is present in the deeper half of the stratum radiatum and in the stratum oriens; ventral to the lesion, degeneration is found predominantly in the superficial one half of the stratum radiatum.     

Commissural and intrinsic connections of the rat hippocampus.

The commissural and intrinsic connections of the hippocampus were studied using the Fink-Heimer method and the horseradish peroxidase (HRP) uptake technique. A conspicuous septo-termporal gradient was found of the density of the commissural projection that passes through the psalterium ventrale to the Ammon's horn. The degeneration resulting from transection of the psalterium ventrale was most dense in the septal tip and decreased towards the temporal tip. The commissural and ipsilateral connections from the hilus fasciae dentatae (CA4) and regio inferior (CA3/CA2) were found to terminate in different parts of the hippocampus. The hilus fasciae dentatae gave rise to ipsilateral and commissural projections to the dentate area only. The regio inferior has ipsilateral and commissural projections to the Ammon's horn. A specific termination pattern was found of the projection from regio inferior to stratum radiatum of both the ipsilateral and contralateral regio superior (CA1) and regio inferior (CA2/CA3). At levels temporal to the lesion, the projection is primarily to the superficial part of stratum radiatum, while at levels septal to the lesion the terminal zone occupies the deep part of the layer. This pattern was not related to the position of the cells of origin, along the septo-temporal or subiculo-dentate axes. In general, the commissural projections showed the same degree of septo-temporal divergence as the ipsilateral projections. The only major difference in the terminal fields of the two sets projections to the Ammon's horn was that the terminal zone of the commissural projection to stratum oriens was always more dense than that of the ipsilateral projection to this layer, while an inverse gradient was seen in stratum radiatum. The projections from the septal and middle dorso-ventral parts of regio inferior differed. The temporal spread of the projections from the septal part was large while that from the projections arising at middle dorso-ventral levels was more restricted. Moreover, a longitudinal association path interconnecting different parts of the regio inferior along the septo-temporal axis was seen to arise only from the cells in the septal parts of the regio inferior. Each part of the regio inferior projected to all parts of stratum radiatum and oriens of the contralateral Ammon's horn. However, the projection to the contralateral regio inferior was most dense at the site homotopic to that lesioned. The ventricular part of regio inferior projected primarily to the contralateral stratum oriens of the Ammon's horn, while the part adjacent to the dentate area mostly supplied stratum radiatum.     

Topographic organization of the projections from the entorhinal area to the hippocampal formation of the rat.

The present study re-examines, with autoradiographic methods, the pattern of termination of fibers originating from various medio-lateral divisions of the entorhinal cortex on dentate granule cells and on hippocampal pyramidal cells of the rat. Entorhinal fibers were found to distribute in a proximo-distal gradient along the dendrites of dentate granule cells, with afferents from the medial entorhinal area terminating in the innermost portion of the entorhinal synaptic field, afferents from the lateral entorhinal area terminating in the most superficial portions of the entorhinal synaptic field, and intermediate medio-lateral locations in the entorhinal area terminating in intermediate locations in the entorhinal synaptic zone. A similar graded pattern of termination of medial and lateral entorhinal fibers was apparent in the very slight crossed projection of the entorhinal area to the contralateral dentate gyrus. In addition, a comparable gradient in the pattern of termination of entorhinal fibers was evident in the entorhinal projection field in the distal regions of the pyramidal cells of regio inferior of the hippocampus proper. Entorhinal projections to regio superior were, however, organized in quite a different fashion. In this zone, there was no evidence of a proximo-distal gradient in the patterns of termination of medial and lateral entorhinal areas along the dendrites of regio superior pyramidal cells. Rather, the medio-lateral organization was in a longitudinal dimension, with medial entorhinal afferents terminating in the portions of regio superior near the CA1-CA2 transition, and lateral entorhinal afferents terminating furthest from the CA1-CA2 transition, immediately adjacent to the CA1-subicular transition, and in the molecular layer of the subiculum proper. A comparable longitudinal organization of entorhinal projections to regio superior was also evident in the zones of termination of the crossed temporo-ammonic tract, contralateral to the injection. These results demonstrate a heretofore unrecognized complexity in the patterns of projection of the entorhinal area to the hippocampal formation, and illustrate that the entorhinal cortex cannot be divided into only two discrete divisions on the basis of the pattern of projection.     

The organization and development of the hippocampal mossy fiber system

The anatomical organization and development of the hippocampal mossy fiber system has been reviewed with special reference to its organization in the common laboratory rat. The mossy fibers originate from the granule cells of the dentate granular layer and the few granule cells found scattered in the dentate molecular layer and hilus. Via a complex system of collaterals the mossy fibers terminate on several types of neurons in the hilus, e.g. the basket cells and the mossy cells. Upon leaving the hilus to pass into Ammon's horn, the mossy fibers converge to form a distinct band of fibers that terminates on the proximal part of the apical and basal dendrites of the pyramidal and basket cells of the regio inferior. In some mammalian species the mossy fibers may continue into the adjacent part of the regio superior. Despite differences in the number of granule cells and pyramidal cells at different septotemporal levels this organization is relatively uniform along the septotemporal extent of the hippocampus. During development the mossy fibers grow out in a sequential manner that matches the pattern of neurogenesis and the aggregation of the cells of origin. From the level at which they originate, the fibers diverge along the septotemporal axis in such a way that the oldest granule cells have the most extensive projections. The adult topographic organization, which is already apparent at the earliest developmental stages, is thus formed in a stepwise fashion. It is concluded that the organization of the hippocampal mossy fibers indicates that neuronal specificity should not be explained by cellular recognition alone, but rather as the cumulated product of the preceding sequence of developmental events that include neurogenesis, migration, aggregation and directed axonal outgrowth.     

The organization and development of the hippocampal mossy fiber system

The anatomical organization and development of the hippocampal mossy fiber system has been reviewed with special reference to its organization in the common laboratory rat.The mossy fibers originate from the granule cells of the dentate granular layer and the few granule cells found scattered in the dentate molecular layer and hilus. Via a complex system of collaterals the mossy fibers terminate on several types of neurons in the hilus, e.g. the basket cells and the mossy cells. Upon leaving the hilus to pass into Ammon's horn, the mossy fibers converge to form a distinct band of fibers that terminates on the proximal part of the apical and basal dendrites of the pyramidal and basket cells of the regio inferior. In some mammalian species the mossy fibers may continue into the adjacent part of the regio superior. Despite differences in the number of granule cells and pyramidal cells at different septotemporal levels this organization is relatively uniform along the septotemporal extent of the hippocampus.During development the mossy fibers grow out in a sequential manner that matches the pattern of neurogenesis and the aggregation of the cells of origin. From the level at which they originate, the fibers diverge along the septotemporal axis in such a way that the oldest granule cells have the most extensive projections. The adult topographic organization, which is already apparent at the earliest developmental stages, is thus formed in a stepwise fashion. It is concluded that the organization of the hippocampal mossy fibers indicates that neuronal specificity should not be explained by cellular recognition alone, but rather as the cumulated product of the preceding sequence of developmental events that include neurogenesis, migration, aggregation and directed axonal outgrowth.     

Localization of enkephalin-like immunoreactivity to identified axonal and neuronal populations of the rat hippocampus

The distribution of enkephalin-like immunoreactivity in the hippocampal formation of the rat was analyzed. Two specific projection systems are described. The first emerges from the hilus of the dentate gyrus and appears to terminate with notably large boutons on the proximal apical and, to a lesser extent, basal dendrites of hippocampal regio inferior pyramidal cells. This projection corresponds in source, position, and character to the hippocampal mossy fiber system. The second axonal population enters the temporal hippocampal formation from the medial wall of the subicular complex and follows the hippocampal fissure to occupy stratum lacunosum-moleculare of the hippocampus proper and the distal third of the dentate gyrus molecular layer; this pattern corresponds to the distribution of afferent input from the lateral entorhinal cortex and/or perirhinal area. Lesions of the hilus or retrohippocampal area caused a selective depletion of immunoreactivity in the mossy fiber fields and molecular layers of the dentate gyrus, respectively. Enkephalin-like immunoreactivity was found within the somata of three types of hippocampal neurons: (1) granule cells of the dentate gyrus, (2) occasional pyramidal shaped cells of field CA1 stratum pyramidale, and (3) varied scattered interneurons. Of this last group, two types of interneurons were consistently seen. The first occupy the border between stratum radiatum and stratum lacunosum-moleculare and extend processes at right angles to the long axis of the pyramidal cell dendrites, whereas the second lie within stratum radiatum of field CA1 and extend processes in alignment with the long axis of the pyramidal cell dendrites. Cells containing enkephalin-like immunoreacactivity were also observed in the subiculum and retrohippocampal region, most notably including layers II and III of the lateral entorhinal cortex-perirhinal area—the probable source of extrinsic immunoreactive input to the hippocampal formation. Intraventricular colchicine treatment intensified the immunoreactive staining of some hippocampal neurons but did not reveal any cell types not seen to be labeled in untreated rats.     

Histochemical demonstration of zinc in the hippocampal region of the domestic pig: II. Subiculum and hippocampus.

The distribution of zinc has been described in two areas of the hippocampal region of the domestic pig, viz., the subiculum and the hippocampus. Zinc was demonstrated histochemically according to the Neo-Timm method, a modification of the sulphide-silver procedure. In each of the examined areas the staining displayed a distinctly stratified pattern which has been compared in detail to fields and layers defined on the basis of cyto- and fibroarchitecture, resulting in a combined chemo- and cytoarchitectonic map. Most of the staining was confined to the neuropil, but a considerable number of stained nerve cell bodies were seen in both the subiculum and the hippocampus. In the subiculum, the plexiform layer was divided into a superficial, weakly stained subzone and a deep, better stained subzone. The cell layer was generally well stained, but displayed a complex staining pattern with differences in staining intensity of both the cell bodies and neuropil. In regio superior of the hippocampus, the stratum moleculare appeared weakly stained, with the exception of a tapering process of more darkly stained tissue projecting from the plexiform layer of the subiculum into the deepest part of the layer. Stratum radiatum and the superficial subzone of stratum oriens showed a weak staining intensity, contrasting to the relatively darkly stained pyramidal cell layer and the intensely stained deep subzone of stratum oriens. In regio inferior, the stratum moleculare was divided into a moderately stained superficial part and an unstained deep part. Stratum radiatum and stratum oriens both appeared weakly stained. The layer of mossy fibers was very intensely stained and appeared almost homogeneously black in its main suprapyramidal part, whereas the infrapyramidal part was looser in character. The pyramidal cell layer was darker than in regio superior. The distribution of zinc in the pig was compared with that in the guinea pig and rat, described previously. The staining pattern is fundamentally similar in all three species, though notable species-specific traits do exist.     

Somatostatin- and neuropeptide Y-like immunoreactivity in the dentate area, hippocampus, and subiculum of the domestic pig.

With the principal aim of providing baseline observations for future experimental studies, the distribution of somatostatin-like and neuropeptide Y-like immunoreactivities is described in the dentate area, hippocampus, and subiculum of the domestic pig (Sus scrofa domesticus) and compared with the distribution described in other mammals. Intensely stained somatostatin-like immunoreactive nerve cell bodies were present throughout the region, with highest densities in the dentate hilus, stratum radiatum and stratum oriens of the hippocampal regio inferior, stratum oriens of the hippocampal regio superior, and in the subicular cell layer. Somatostatin-like immunoreactive terminals were represented by both stained fibers and stained puncta. Scattered somatostatin-like immunoreactive nerve fibers were seen in most areas, but regular fiber plexuses were present in the dentate molecular layer and dentate hilus, stratum moleculare of the hippocampus, and in the subicular plexiform layer. Somatostatin-like immunoreactive puncta were seen in the dentate molecular layer, stratum moleculare of the hippocampus, and in the subicular plexiform layer. Neuropeptide Y-like immunoreactive nerve cell bodies were less numerous than somatostatin-like immunoreactive ones. They were mainly seen in the dentate granule cell layer and dentate hilus, stratum radiatum and stratum oriens of the hippocampus, and in the subicular cell layer. Intensely stained neuropeptide Y-like immunoreactive fibers were numerous, and present in all areas examined. They formed fiber plexuses in the dentate molecular layer and dentate hilus, stratum moleculare of the hippocampal regio superior, and in the subicular plexiform layer. Neuropeptide Y-like immunoreactive puncta were present in the dentate molecular layer, stratum moleculare of the hippocampus, and in the subicular plexiform layer. Consistent and very characteristic variation in the distribution of somatostatin-like and neuropeptide Y-like immunoreactivity was found along the septotemporal axis of the hippocampus. The distribution of somatostatin-like and neuropeptide Y-like neurons and terminals in the domestic pig displayed striking similarities with the basic pattern of organization of these neuropeptides in other species, although more subtle species-specific characteristics were also observed in the pig.     

Detailed projection patterns of septal and diagonal band efferents to the hippocampus in the rat with emphasis on innervation of CA1 and dentate gyrus

The detailed patterns of afferentation to the ammon's horn and dentate gyrus of the hippocampus in the rat were investigated employing the anterograde tracer Phaseolus vulgaris leuco-agglutinin (PHA-L) after punctate iontophoretic injections in the medial septum (MS) and vertical limb of the diagonal band of Broca (VDB). The topographically ordered innervation pattern was different in the regio superior (or CA1) vs. the regio inferior (or CA3) and in the dorsal vs. ventral aspects of ammon's horn and dentate gyrus. The CA1 pyramidal and dentate granule cell layers in the dorsal hippocampus received afferent input almost exclusively from the VDB, whereas those cell layers in ventral hippocampus were supplied from both VDB and MS. The PHA-L labeled projecting fibers could be differentiated into two distinct fiber systems. One class of thick and coarse axons (tentatively called type I fibers) carried fewer but larger terminal boutons and were found to infiltrate the entire stratum oriens, dentate hilus, all layers of the regio inferior and the CA1 str. moleculare. A second, delicate thin (type II) fiber system provided with numerous and passant varicosities showed a much more restricted laminar innervation pattern and appeared to originate from areas in MS-VDB which are rich in AChE-positive neurons. The densest type II fiber networks could be observed in the CA1 subpyramidal and dentate supragranular zones, in the CA1 stratum lacunosum-moleculare and in the dentate middle third molecular layer. This laminar type II innervation pattern showed a remarkable coincidence with the reported distribution of cholinergic marker enzymes. The topographic and spatial organization of the projections described above will be discussed in relation to their possible functional significance.     

The development of Ammon's horn and the fascia dentata in the cat: a [3H]thymidine analysis

The present [3H]thymidine autoradiographic analysis of neurogenesis demonstrates that the neurons which populate the adult cat hippocampus are born between embryonic day (E)22 and E42. In contrast, although neuronal production in the fascia dentata begins on the same day, granule cells in this area continue to be produced throughout prenatal life and into early postnatal life, and probably continues at an extremely low rate well into adulthood. Three major sets of spatiotemporal gradients characterize the production of neurons in Ammon's horn and the fascia dentata. The first set involves the radial axis. Within the hippocampus there exists an inside-out gradient. The reverse gradient is present in the fascia dentata, i.e. outside-in. The second set of gradients involves the transverse or rhinodentate axis. In general the CA3 neurons are born earlier than the CA1 neurons. Within both neuronal layers of the fascia dentata, the hidden blade cells tend to be born earlier than those of the exposed blade. Again, the pattern in the fascia is the reverse of that in the hippocampus proper. A temporal to septal gradient is also present, but this is the weakest of the gradients.