Species differences in the projections from the entorhinal cortex to the hippocampus

Both differences and similarities exist between mammalian species in the projections from entorhinal cortex to the hippocampal formation. In most species, layer II cells of the entorhinal cortex project to the dentate gyrus, and they terminate in the outer two-thirds of the molecular layer of the dentate gyrus. The axons from layer III cells project bilaterally to areas CA(1) and CA(3) of the hippocampus, terminating in the stratum lacunosum moleculare. We have analyzed these projections in mice, and in general, the entorhinal cortex-to-hippocampus projections are similar to those in rats. Axons from layer II neurons terminate in the outer and middle thirds of the molecular layer of the dentate gyrus, and axons from layer III neurons terminate bilaterally in the stratum lacunosum moleculare of areas CA(1) and CA(3), and in the molecular layer of the subiculum. However, in contrast to rat, mouse entorhinal cortex neurons do not appreciably project to the contralateral dentate gyrus. Most species, including mice, show a similar topographical organization of the entorhinal-hippocampal projections, with neurons in the lateral part of both the lateral and medial entorhinal cortex projecting to the dorsal part or septal pole of the hippocampus, whereas the projection to the ventral hippocampus originates primarily from neurons in medial parts of the entorhinal cortex.     

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.     

Ascending projections of the locus coeruleus in the rat. II. Autoradiographic study.

The ascending projections of the locus coeruleus were studied using an autoradiographic method. The major projection of locus coeruleus neurons ascends in a dorsal pathway traversing the midbrain tegmentum in a position ventrolateral to the periaqueductal gray. At the caudal diencephalon the locus coeruleus axons descend to enter the medial forebrain bundle at a caudal tuberal hypothalamic level. They are jointed in the medial forebrain bundle by a much smaller locus coeruleus projection which takes a ventral course through the midbrain tegmentum and enters the medial forebrain bundle via the mammillary peduncle and ventral tegmental area. Terminal projections are evident in the midbrain to the periaqueductal gray, tegmentum and raphe nuclei. There are widespread projections to the dorsal thalamus. The heaviest of these are to the intralaminar nuclei, the anteroventral and anteromedial nuclei, the dorsal lateral geniculate and the paraventricular nucleus. In the hypothalamus the largest projections are to the lateral hypothalamic area, periventricular nucleus, supraoptic nucleus and paraventricular nucleus. As the locus coeruleus projection ascends in the medial forebrain bundle, fibers leave it to traverse the lateral hypothalamus and zona incerta and enter the internal capsule, the ventral amygdaloid bundle and ansa peduncularis. These appear to terminate in the amygdaloid complex and, via the external capsule, in the lateral and dorsal neocortex. At the level of the septum 4 projections are evident. One group of fibers enters the stria medullaris to terminate in the paraventricular nucleus and habenular nuclei. A second group joins the stria terminalis to terminate in the anygdaloid complex. The third group turns into the diagonal band and medial septum; some fibers terminate in the septal nuclei and others continue into the fornix to termimate in hippocampus. A large component continues around the corpus callosum into the cingulum to terminate in the cingulate and adjacent neocortex, the subiculum and hippocampus. The remaining fibers continue rostrally in the medial forebrain bundle to terminate in olfactory forebrain and frontal neocortex. Commissural projections arise at 4 locations. The first decussation occurs in the dorsal tegmentum just below the central gray rostral to the locus coeruleus. The crossing fibers enter the contralateral dorsal bundle. A second group of fibers leaves the ipsilateral dorsal pathway, crosses in the posterior commissure and enters the contralateral dorsal pathway at the level. The third commissural projection arises more rostrally and crosses in the dorsal supraoptic commissure to enter the contralateral medial forebrain bundle. The fourth commissural projection is through the anterior commissure. The termination of the contralateral projection appears similar to that of the ipsilateral projection.     

Cholinergic neurons within the rat hippocampus: response to fimbria-fornix transection

The distribution and morphologic characteristics of choline acetyltransferase (ChAT)-containing neurons were studied throughout the rostrocaudal extent of the rat hippocampus and in a midline area just dorsal to the dorsal hippocampus. Peroxidase reaction product was observed with the aid of immunohistochemical methods and a high-titer polyclonal antibody against ChAT, the acetylcholine biosynthetic enzyme. ChAT-positive cells in the hippocampus were characterized by small, round or oval perikarya with two or more proximal processes. They were located within the caudal and temporal hippocampal formation, predominantly within the subiculum, in the stratum lacunosum moleculare, at the border of the stratum lacunosum moleculare and the stratum radiatum, and in the molecular layer of the dentate gyrus. The cells resembled in morphology the small, bipolar and multipolar neocortical ChAT-immunoreactive cells. In addition to the hippocampus, ChAT-positive neurons were observed caudally in a region just above the dorsal hippocampal commissure and rostrally in the columns of the fornix. These cells were large with an oval perikarya and darkly labeled compared to neurons in the hippocampus. They more closely resembled the ChAT-positive neurons in the midline raphe of the medial septal nucleus. Examination of the rat hippocampus 2 and 8 weeks following unilateral lesioning of the fimbria-fornix and supracallosal striae revealed a sparse innervation of ChAT-positive fibers in the hippocampus ipsilateral to the lesion. ChAT-labeled neurons in the hippocampus did not appear to sprout in response to the lesion. In contrast, ChAT-positive cells in the midline did appear to sprout into the medial dorsal subiculum and dorsal medial hippocampus. We conclude that these two populations of cells are distinct with respect to their response to hippocampal denervation and, furthermore, that this distinction may be attributed to a differential response to nerve growth factor.     

The efferent connections of the nucleus raphe centralis superior in the rat as revealed by radioautography.

By chronically implanting a glass micropipette filled with tritiated leucine in the raphe centralis superior of the rat, the projection of this nucleus was traced by radioautography. The majority of the ascending projections were located within the ventral tegmental area and, further rostrally, the median forebrain bundle. Along the course of this bundle numerous fibers branched successively into the mammillary peduncle, the fasciculus retroflexus, the stria medullaris, the fornix and the cingulum. The most significant projections included the ones to the interpeduncular nucleus, the mammillary bodies, the habenular nuclei and the hippocampus. No projections were detected in the striatum, the cortex piriformis or the amygdala. Descending projections diffused to the pontine reticular formation and central gray through the medial and the dorsal longitudinal bundles. In addition widespread projections were also seen in nuclei located near the raphe centralis superior: raphe nuclei, dorsal and ventral tegmental nuclei.     

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.     

Projections of the ventral subiculum to the amygdala, septum, and hypothalamus: a PHAL anterograde tract-tracing study in the rat.

The projections of the ventral subiculum are organized differentially along the dorsoventral (or septotemporal) axis of this cortical field, with more ventral regions playing a particularly important role in hippocampal communication with the amygdala, bed nuclei of the stria terminalis (BST), and rostral hypothalamus. In the present study we re-examined the projection of the ventral subiculum to these regions with the Phaseolus vulgaris leucoagglutinin (PHAL) method in the rat. The results confirm and extend earlier conclusions based primarily on the autoradiographic method. Projections from the ventral subiculum course either obliquely through the angular bundle to innervate the amygdala and adjacent parts of the temporal lobe, or follow the alveus and fimbria to the precommissural fornix and medial corticohypothalamic tract. The major amygdalar terminal field is centered in the posterior basomedial nucleus, while other structures that appear to be innervated include the piriformamygdaloid area, the posterior basolateral, posterior cortical, posterior, central, medial, and intercalated nuclei, and the nucleus of the lateral olfactory tract. Projections from the ventral subiculum reach the BST mainly by way of the precommissural fornix, and provide rather dense inputs to the anterodorsal area as well as the transverse and interfascicular nuclei. The medial corticohypothalamic tract is the main route taken by fibers from the ventral subiculum to the hypothalamus, where they innervate the medial preoptic area, "shell" of the ventromedial nucleus, dorsomedial nucleus, ventral premammillary nucleus, and cell-poor zone around the medial mammillary nucleus. We also observed a rather dense terminal field just dorsal to the suprachiasmatic nucleus that extends dorsally and caudally to fill the subparaventricular zone along the medial border of the anterior hypothalamic nucleus and ventrolateral border of the paraventricular nucleus. The general pattern of outputs to the hypothalamus and septum is strikingly similar for the ventral subiculum and suprachiasmatic nucleus, the endogenous circadian rhythm generator.     

Localization of the ras-like rab3A protein in the adult rat brain.

Rab3A is a small GTP-binding synaptic vesicle protein, shown to dissociate from synaptic vesicle membranes upon depolarization-induced exocytosis. Using an antiserum raised against rab3A, we found that the antigen was localized to the neuropil of specific brain regions, but was not present in major fiber tracts or most cell bodies. For example, the neuropil of several thalamic nuclei (i.e, dorsal lateral geniculate nucleus, lateral posterior nucleus, ventroposterior nucleus), cerebral cortex, upper layers of the superior colliculus and matrix zones of the neostriatum, were strongly immunoreactive, while the anterior commissure, corpus callosum, optic tract and internal capsule were devoid of staining. The hippocampus, dark staining was observed in the stratum oriens, stratum radiatum and molecular layer of the dentate gyrus, while the pyramidal, stratum lacunosum moleculare and dentale granule layers were not stained. In cerebellum the molecular layer and to a lesser extent, the underlying granule cell layer showed enhanced immunoreactivity. Seven days after excitotoxic lesions of the cerebral cortex, rab3A immunoreactivity was diminished in the mirror locus in the contralateral coritcal hemisphere and in certain thalamic nuclei ipsilateral to the injection site. These results show that rab3A is localized to a number of specific regions. Its absence from other areas suggests that this synaptic vesicle protein is not universal to all neuronal terminals and pathways. In addition, our lesion studies indicate that for some brain regions, much of the antigen originates in cortical neurons and is distributed within specific axonal projections.     

Localization of the ras-like rab3A protein in the adult rat brain

Rab3A is a small GTP-binding synaptic vesicle protein, shown to dissociate from synaptic vesicle membranes upon depolarization-induced exocytosis. Using an antiserum raised against rab3A, we found that the antigen was localized to the neuropil of specific brain regions, but was not present in major fiber tracts or most cell bodies. For example, the neuropil of several thalamic nuclei (i.e, dorsal lateral geniculate nucleus, lateral posterior nucleus, ventroposterior nucleus), cerebral cortex, upper layers of the superior colliculus and matrix zones of the neostriatum, were strongly immunoreactive, while the anterior commissure, corpus callosum, optic tract and internal capsule were devoid of staining. The hippocampus, dark staining was observed in the stratum oriens, stratum radiatum and molecular layer of the dentate gyrus, while the pyramidal, stratum lacunosum moleculare and dentale granule layers were not stained. In cerebellum the molecular layer and to a lesser extent, the underlying granule cell layer showed enhanced immunoreactivity. Seven days after excitotoxic lesions of the cerebral cortex, rab3A immunoreactivity was diminished in the mirror locus in the contralateral coritcal hemisphere and in certain thalamic nuclei ipsilateral to the injection site. These results show that rab3A is localized to a number of specific regions. Its absence from other areas suggests that this synaptic vesicle protein is not universal to all neuronal terminals and pathways. In addition, our lesion studies indicate that for some brain regions, much of the antigen originates in cortical neurons and is distributed within specific axonal projections.     

Alterations in muscarinic cholinergic receptor densities induced by thiamine deficiency: autoradiographic detection of changes in high- and low-affinity agonist binding

Animals fed a diet deficient in thiamine or treated with a drug preventing the utilization of thiamine (thiamine antagonist) exhibited alterations in ligand binding to muscarinic receptors in several brain regions. Using quantitative techniques of receptor autoradiography, an increase in muscarinic receptor binding was demonstrated in such regions as the corpus callosum, lamina VI of the parietal cortex, caudate-putamen, ventral nucleus of the thalamus, stratum lacunosum moleculare and stratum oriens of the hippocampus, and the hilus of the area dentata. As a result of thiamine deficiency, this increase in muscarinic receptor populations was primarily due to an increase in the binding of the low-affinity agonist site. In the same experiment, a decrease in muscarinic receptor binding was found in the ventromedial region of the hypothalamus. Thiamine deficiency thus causes an up-regulation of muscarinic receptor binding in several regions of rat brain while causing a down-regulation of these same receptors in other brain areas.     

Projections from the periamygdaloid cortex to the amygdaloid complex, the hippocampal formation, and the parahippocampal region: a PHA-L study in the rat

The periamygdaloid cortex, an amygdaloid region that processes olfactory information, projects to the hippocampal formation and parahippocampal region. To elucidate the topographic details of these projections, pathways were anterogradely traced using Phaseolus vulgaris leukoagglutinin (PHA-L) in 14 rats. First, we investigated the intradivisional, interdivisional, and intra-amygdaloid connections of various subfields [periamygdaloid subfield (PAC), medial subfield (PACm), sulcal subfield (PACs)] of the periamygdaloid cortex. Thereafter, we focused on projections to the hippocampal formation (dentate gyrus, hippocampus proper, subiculum) and to the parahippocampal region (presubiculum, parasubiculum, entorhinal, and perirhinal and postrhinal cortices). The PACm had the heaviest intradivisional projections and it also originated light interdivisional projections to other periamygdaloid subfields. Projections from the other subfields converged in the PACs. All subfields provided substantial intra-amygdaloid projections to the medial and posterior cortical nuclei. In addition, the PAC subfield projected to the ventrolateral and medial divisions of the lateral nucleus. The heaviest periamygdalohippocampal projections originated in the PACm and PACs, which projected moderately to the temporal end of the stratum lacunosum moleculare of the CA1 subfield and to the molecular layer of the ventral subiculum. The PACm also projected moderately to the temporal CA3 subfield. The heaviest projections to the entorhinal cortex originated in the PACs and terminated in the amygdalo-entorhinal, ventral intermediate, and medial subfields. Area 35 of the perirhinal cortex was lightly innervated by the PAC subfield. Thus, these connections might allow for olfactory information entering the amygdala to become associated with signals from other sensory modalities that enter the amygdala via other nuclei. Further, the periamygdalohippocampal pathways might form one route by which the amygdala modulates memory formation and retrieval in the medial temporal lobe memory system. These pathways can also facilitate the spread of seizure activity from the amygdala to the hippocampal and parahippocampal regions in temporal lobe epilepsy.     

Electrical signs of an oligosynaptic visual projection to the rat hippocampus

In rats with pons transection photic or optic nerve stimulation elicits a response in dorsal hippocampus with approximately with same latency, amplitude and time course as the response in striate cortex. After optic nerve stimulation a positive polyphasic deflection with the last peak at 10 ms is followed by a larger biphasic major deflection with peaks at 15 and 25 ms and later, slower deflections at intervals of 125 ms. The polyphasic deflection is maximal in the inferior part of the hippocampus but does not reverse polarity; the other deflections reverse above and below an isoelectric point in the hilum of the dentate gyrus, a distribution attributable to depolarization in the molecular layers, perhaps also cell bodies, of that structure. The major deflection is more sensitive to changes in optic nerve stimulus strength than the response in striate cortex and is more resistant to reduction in amplitude during repetitive stimulation, following frequencies up to 50/s. The pathway between retina and hippocampus for all parts of the response is interrupted by lesions in the tectum. The major deflection is abolished by lesions in the posterior cingulum. In the posterior cingulum the pathway has fast and slow components in the lower and upper portions, respectively, associated with the first and second parts, respectively, of the major deflection. The pathway is not interrupted by lesions in the fornix, septal nuclei, anterior cingulum, anterior medial thalamus or medial midbrain ventral to the superficial tectum. There are complex interactions, up to several hundred milliseconds in duration, between the response to optic nerve stimulation and those elicited by stimulation in the cingulum, midbrain and thalamus. Tonic influences on the dentate gyrus from cingulum and tectum are described.     

The topographical organization of the hippocampal projection to the septal area: a comparative neuroanatomical analysis in the gerbil, rat, rabbit, and cat

An experiment was performed to determine the origin of the projection from the hippocampus to the septal area in the subrimate mammalian nervous system. Lesions were made by aspiration or by radio frequency in 4 gerbils, 17 rats, 8 rabbits, and 7 cats. Survival times varied from 2–5 days. Tissues were stained principally with the Fink Heimer I method for identification of degenerating axons and their terminals. Following lesions destroying any one or more of the fields of the dorsal hippocampus of the gerbil, rat, rabbit, or cat, terminal degeneration was observed only in the medial septal area, olfactory tubercle, and adjacent portions of the diagonal band. In addition, lesions producing total destruction of all dorsal hippocampal fields also resulted in the presence of terminal degeneration restricted to the medial septal area. In contrast, superficial lesions of field CA1 of the ventral hippocampus produced terminal degeneration in the lateral septal area, nucleus accumbens, olfactory tubercle, and adjacent portions of the diagonal band. Similar findings were also observed following more widespread lesions of the ventral hippocampus which produced damage to other CA fields as well. Superficial lesions of the posterior crus of the hippocampus (i.e., a position midway between dorsal and ventral hippocampus) resulted in terminal degeneration localized to an intermediolateral region of the septum. Combined lesions of the dorsal hippocampus and fimbria produced widespread terminal degeneration in both the lateral and medial septum indicating that the axons contained within the fimbria arise only from the ventral hippocampus. Finally, lesions of the medial and lateral segments of the fornix of the cat produced terminal degeneration in the medial and lateral regions of the septum, respectively. These findings, collectively, indicate that the origin of the topographical projection to the medial and lateral septum are the dorsal and ventral hippocampus, respectively. This projection is unrelated to cytoarchitectonic fields within the hippocampus and is also invariant among the species considered in this study.     

Hypothalamic afferent connections mediating adrenocortical responses that follow hippocampal stimulation

This study identified some neural pathways which mediate the adrenocortical responses that follow hippocampal stimulation. The increase in plasma corticosterone following dorsal hippocampus stimulation, in rats with electrodes chronically implanted under pentobarbital anesthesia, was blocked by dorsal fornix and lateral septal lesions and by small posterior hypothalamic deafferentation. Fimbria transection, lateral septal lesions, and posterior hypothalamic deafferentation, but not midbrain reticular formation lesions, also blocked the adrenocortical responses to ventral hippocampus stimulation. Our present and previous studies indicate that the dorsal and ventral hippocampal effects on the hypothalamus, which increase plasma corticosterone concentrations, are mediated by the dorsal fornix and fimbria, respectively, as well as by the lateral septum. A posterior hypothalamic input, which does not involve the medial forebrain bundle or the midbrain reticular formation is also essential for the activation of this response.     

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.     

Chromogranins as markers of altered hippocampal circuitry in temporal lobe epilepsy

Chromogranins are polypeptides which are widely expressed in the central nervous system. They are stored in dense core vesicles of nerve terminals, from where they are released upon stimulation. Using immunocytochemistry, we investigated the distribution of chromogranin A, chromogranin B, secretoneurin, and, for comparison, dynorphin in hippocampal specimens removed at routine surgery from patients with drug-resistant mesial temporal lobe epilepsy and in autopsy tissues from nonneurologically deceased subjects. In post mortem controls (n = 21), immunoreactivity for all 4 peptides (most prominently for chromogranin B and dynorphin) was observed in the terminal field of mossy fibers. For chromogranins, staining was observed also in sectors CA1 to CA3 and in the subiculum. Chromogranin B immunoreactivity was found in the inner molecular layer of the dentate gyrus, the area of terminating associational-commissural fibers. Secretoneurin and dynorphin immunoreactivity labeled the outer molecular layer and the stratum lacunosum moleculare of sectors CA1 to CA3, where projections from the entorhinal cortex terminate. In specimens with Ammon's horn sclerosis (n = 25), staining for all 3 chromogranins and for dynorphin was reduced in the hilus of the dentate gyrus. Instead, intense staining was observed in the inner molecular layer, presumably delineating terminals of sprouted mossy fibers. Specimens obtained from temporal lobe epilepsy patients without Ammon's horn sclerosis (n = 4) lacked this pronounced rearrangement of mossy fibers. In the stratum lacunosum moleculare of sector CA1, secretoneurin and dynorphin immunoreactivity was reduced in sclerotic, but not in nonsclerotic, specimens, paralleling the partial loss of fibers arising from the entorhinal cortex. Instead, presumably sprouted secretoneurin-immunoreactive fibers were found in the outer dentate molecular layer in sclerotic specimens. These changes in staining patterns for chromogranins and dynorphin mark profound plastic and functional rearrangement of hippocampal circuitry in temporal lobe epilepsy.     

Hippocampal innervation by serotonin neurons of the midbrain raphe in the rat.

The organization of the brainstem serotonin neuron projection to the hippocampal formation was analyzed in the rat. This projection arises in the raphe nuclei of the midbrain. Following destruction of the midbrain raphe nuclei, chiefly nucleus centralis superior, there is a 72% decrease in hippocampal serotonin content. Injection of tritiated amino acid into the midbrain raphe nuclei results in transport of tritiated protein to the hippocampal formation and this transport is blocked in animals pretreated by intraventricular administration of 5,6-dihydroxytryptamine (5,6-DHT). Autoradiographic analysis indicates that the transport reaches the hippocampal formation primarily via two major pathways, the cingulum and the fornix. Cingulum fibers terminate predominantly in the dorsal hippocampus whereas the fornix distributes throughout the entire hippocampal formation. Some fibers reach the ventral hippocampus from the entorhinal area. Within the hippocampus there is dense labeling in a restricted lamina of the CA1 stratum lacunosum-moleculare with moderate labeling in stratum radiatum. Stratum oriens is sparsely labeled in CA1 and moderately so in CA2 and CA3. Stratum radiatum and stratum lacunosum-moleculare are moderately densely labeled in CA2 and Ca3. The area dentata is sparsely to moderately labeled in the molecular layer and heavily labeled in a thin lamina of the hilar zone immediately beneath the granule cell layer. The remaining hilar zone is moderately labeled. All of the discrete labeling of the hippocampus and area dentata described above is absent in animals pretreated with 5,6-DHT. These observations indicate that serotonin neurons of the midbrain raphe provide a highly organized innervation of the hippocampal formation in the rat.     

Release of endogenous opioid peptides displaces [3H]diprenorphine binding in rat hippocampal slices

Pharmacological depolarization by KCl or veratrine reduced [3H]diprenorphine binding to opioid receptors in the hippocampal slice in a transient, calcium-dependent, and peptide-sensitive manner. These results suggest that endogenous opioid peptides were released from synaptic terminals and competitively displaced [3H]diprenorphine binding to opioid receptors. [3H]diprenorphine binding was significantly reduced by calcium-dependent depolarization throughout the hippocampus as determined by subsequent receptor autoradiography and quantitative densitometry. Displacement of binding was evident at sites in the CA1 and CA3 regions, the dentate gyrus, and the subiculum. The most dramatic reduction was evident in stratum lacunosum moleculare of CA3. Correlating the sites of maximal [3H]diprenorphine displacement with the previously described distribution of the opioid peptides suggests that the perforant path fibers release enkephalins in stratum lacunosum moleculare of CA3 and stratum moleculare of the dentate gyrus, and that mossy fibers may release both dynorphins and enkephalins near stratum pyramidale of CA3 and stratum granulosum. The lack of complete overlap between the distribution of opioid terminals and the sites of displacement indicates that these peptides may diffuse a moderate distance to their sites of action. Radioligand displacement defines the sites of endogenous opioid binding, suggests the likely sources of peptide release, and thus predicts the sites of endogenous opioid action within the hippocampus.     

Kindling induced changes in calmodulin kinase II immunoreactivity

The distribution of type II calmodulin kinase (CaM kinase) immunoreactivity was studied in control and septally kindled rat brains. CaM kinase was concentrated in limbic structures, such as the hippocampus, lateral septum and amygdala. Within the hippocampus, the molecular layer of the endal limb of the dentate gyrus, the stratum radiatum, and lacunosum moleculare of CA1 were the most heavily stained regions. The cerebellum was stained only in the molecular and Purkinje cell layers, and very low amounts of immunoreactive protein were present in the brainstem and white matter. Kindling resulted in a significant decrease in CaM kinase immunoreactivity in CA3 and in the dentate of the ventral hippocampus but not in the lateral septum. These data suggest that kindling decreases the number of CaM kinase molecules or alters its antigenic distribution, and provides further evidence that alterations of this enzyme may be important in the kindling phenomenon.