Septotemporal distribution of [3H]MK-801, [3H]AMPA and [3H]Kainate binding sites in the rat hippocampus

 The distribution of glutamate receptors in transverse hippocampal sections has been well investigated. However, in spite of the known septotemporal gradients of hippocampal connectivity no systematic studies exist about the distribution of glutamate receptors along the septotemporal (longitudinal) hippocampal axis. Therefore, in the present study this issue was investigated using receptor autoradiography for the [³H]MK-801, [³H]AMPA and [³H]Kainate binding sites. Hippocampi from 30-day-old rats were sectioned perpendicularly to their longitudinal axis, yielding a total of 25–30 equidistantly spaced autoradiographs for each hippocampus. For each section layer-specific concentrations of binding sites were calculated by the aid of a computerized image analysing system. The dependency of concentrations of binding sites on the septotemporal position was evaluated by regression analysis. Gradients of binding were confined to distinct hippocampal layers. Significant septotemporal gradients of [³H]MK-801 binding were observed in selected layers of CA1 and the dentate gyrus, a septal to temporal decrease of binding in the oriens and radiatum layers of CA1 being most prominent. For [³H]AMPA, significant septotemporal gradients of binding were restricted to layers of CA3, CA4 and the dentate gyrus, with values generally increasing from septal to temporal levels. The observed septotemporal gradients possibly reflect functional segregations along the longitudinal hippocampal axis and could be important for the comparability of ligand binding studies using transverse hippocampal sections or hippocampal slice cultures. Accepted: 2 April 1998     

Distribution of [3H]MK-801, [3H]AMPA and [3H]Kainate binding sites in rat hippocampal long-term slice cultures isolated from external afferents

The distribution of ionotropic glutamate receptors in transverse hippocampal sections and along the septotemporal hippocampal axis can be correlated to hippocampal connectivity, in particular to area- and layer-specific termination zones of afferents. However, in isolated organotypic hippocampal slice cultures developing without extrinsic afferent input no systematic studies exist about the distribution of glutamate receptors. In the present study we used receptor autoradiography to examine [³H]MK-801, [³H]AMPA and [³H]Kainate binding sites in hippocampal slice cultures prepared from 6-day-old rats. After 24 days in vitro layer-specific concentrations of [³H]MK-801, [³H]AMPA and [³H]Kainate binding sites were compared to age-matched hippocampi in situ (P30 rats). An obvious difference between hippocampal slice cultures and hippocampi in situ was a changed distribution of binding sites among the hippocampal areas showing a relative increase of [³H]MK-801 and [³H]AMPA binding sites in CA3 as compared to CA1 and to the dentate gyrus in the cultures. In CA1, however, the relative layer-specific distributions of [³H]MK-801 and [³H]AMPA binding sites were identical in hippocampal slice cultures and in hippocampi in situ. Interestingly, layer-specific binding of [³H]Kainate in the cultures exceeded that in the hippocampi in situ 3–5 times. Moreover, in the cultures the binding of the three ligands varied systematically showing gradients along the ”superficiomembranal’’ axis. Cultures taken from different positions along the hippocampal axis differed with respect to concentrations of [³H]MK-801 and [³H]Kainate binding sites, but not of [³H]AMPA binding sites. The results suggest a massive sprouting and reorganisation of intrinsic projections in long-term hippocampal slice cultures.     

Distribution of [3H]MK-801, [3H]AMPA and [3H]Kainate binding sites in rat hippocampal long-term slice cultures isolated from external afferents

The distribution of ionotropic glutamate receptors in transverse hippocampal sections and along the septotemporal hippocampal axis can be correlated to hippocampal connectivity, in particular to area- and layer-specific termination zones of afferents. However, in isolated organotypic hippocampal slice cultures developing without extrinsic afferent input no systematic studies exist about the distribution of glutamate receptors. In the present study we used receptor autoradiography to examine [³H]MK-801, [³H]AMPA and [³H]Kainate binding sites in hippocampal slice cultures prepared from 6-day-old rats. After 24 days in vitro layer-specific concentrations of [³H]MK-801, [³H]AMPA and [³H]Kainate binding sites were compared to age-matched hippocampi in situ (P30 rats). An obvious difference between hippocampal slice cultures and hippocampi in situ was a changed distribution of binding sites among the hippocampal areas showing a relative increase of [³H]MK-801 and [³H]AMPA binding sites in CA3 as compared to CA1 and to the dentate gyrus in the cultures. In CA1, however, the relative layer-specific distributions of [³H]MK-801 and [³H]AMPA binding sites were identical in hippocampal slice cultures and in hippocampi in situ. Interestingly, layer-specific binding of [³H]Kainate in the cultures exceeded that in the hippocampi in situ 3–5 times. Moreover, in the cultures the binding of the three ligands varied systematically showing gradients along the ”superficiomembranal’’ axis. Cultures taken from different positions along the hippocampal axis differed with respect to concentrations of [³H]MK-801 and [³H]Kainate binding sites, but not of [³H]AMPA binding sites. The results suggest a massive sprouting and reorganisation of intrinsic projections in long-term hippocampal slice cultures. Accepted: 6 February 2001     

Excitatory amino acid receptors in the human cerebral cortex: a quantitative autoradiographic study comparing the distributions of [3H]TCP, [3H]glycine, L-[3H]glutamate, [3H]AMPA and [3H]kainic acid binding sites

The excitatory amino acids are probably the major neurotransmitters in the cerebral cortex, and they act through at least three receptors: the N-methyl-D-aspartate, the quisqualate and the kainic acid receptors. Under the appropriate conditions, [3H]1-(1-(2-thienyl)-cyclohexyl)piperidine [( 3H]TCP), [3H]glycine and L-[3H]glutamate label different sites on the N-methyl-D-aspartate receptor, [3H]-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid [( 3H]AMPA) labels the quisqualate receptor and [3H]kainic acid the kainic acid receptor. The anatomical localizations of these binding sites were studied in sections of blocks removed from the cerebral cortices of eight post-mortem human brains. The results showed that, in the human cerebral cortex, [3H]TCP, [3H]glycine and L-[3H]glutamate binding sites had congruent distributions, with [3H]AMPA binding sites showing a similar distribution. In the hippocampus, these four ligands had high binding site densities in the CA1 region and the dentate gyrus molecular layer. With the exception of the striate cortex, in the neocortex, a tri-laminar pattern was seen consisting of a high density across laminae I-III, a layer of low density corresponding to the region of lamina IV, and a band of moderate density across laminae V and VI, except for [3H]AMPA where the middle zone of low density was usually wider. [3H]Kainic acid showed a binding pattern which was generally complementary to that of the other four ligands. There were low levels of [3H]kainic acid binding sites in the CA1 region of the hippocampus with higher levels in the CA3 region, the hilus, and the inner third of the dentate gyrus molecular layer. In the neocortex there was a band of high density corresponding to laminae V and VI, with a thin band of moderate binding corresponding to lamina I and the outer region of lamina II. An exception was the motor cortex where the highest level of [3H]kainic acid binding was in laminae I and II. The high degree of congruence between the binding patterns of [3H]TCP, [3H]glycine and L-[3H]glutamate (using conditions appropriate for the N-methyl-D-aspartate receptor) supports data indicating that these ligands bind to different regions of the same receptor complex. The similar distribution of [3H]AMPA binding sites, with the exception of the striate cortex, supports observations made in rodents that N-methyl-D-aspartate receptors and quisqualate receptors have similar distributions and perform different but related functions in excitatory transmission.(ABSTRACT TRUNCATED AT 400 WORDS)     

近交系小鼠海马结构兴奋性氨基酸受体密度的变化(英文)

应用定量放射自显影技术，以氚标配体MK－801、AMPA和KA分别标记NMDA、AMPA和KA受体，定量观察了七种近交系小鼠海马结构内兴奋性氨基酸受体的区域分布类型和受体密度变化．结果表明，海马CA1区含有高密度NMDA和AMPA受体，CA3和齿状回含高密度KA受体。三种受体在海马各区的分布类型无明显系间差异，而其受体密度存在着明显的系间差别。NMDA和AMPA受体在BALB.c鼠海马显示最高密度，而CPB-K鼠以上两种受作密度最低。KA受体在NMRI鼠海马显示最高密度，BA鼠其受体密度最低．这种受体密度的系间差异可能与近交系小鼠合成兴奋性氨基酸受体的基因调控不同有关。     

Differential expression of NMDA and AMPA receptor subunits in rat dorsal and ventral hippocampus

Several studies have demonstrated anatomical and functional segregation along the dorsoventral axis of the hippocampus. This study examined the possible differences in the AMPA and NMDA receptor subunit composition and receptor binding parameters between dorsal and ventral hippocampus, since several evidence suggest diversification of NMDA receptor-dependent processes between the two hippocampal poles. Three sets of rat dorsal and ventral hippocampus slices were prepared: 1) transverse slices for examining a) the expression of the AMPA (GluRA, GluRB, GluRC) and NMDA (NR1, NR2A, NR2B) subunits mRNA using in situ hybridization, b) the protein expression of NR2A and NR2B subunits using Western blotting, and c) by using quantitative autoradiography, c(1)) the specific binding of the AMPA receptor agonist [(3)H]AMPA and c(2)) the specific binding of the NMDA receptor antagonist [(3)H]MK-801, 2) longitudinal slices containing only the cornus ammonis 1 (CA1) region for performing [(3)H]MK-801 saturation experiments and 3) transverse slices for electrophysiological measures of NMDA receptor-mediated excitatory postsynaptic potentials. Ventral compared with dorsal hippocampus showed for NMDA receptors: 1) lower levels of mRNA and protein expression for NR2A and NR2B subunits in CA1 with the ratio of NR2A /NR2B differing between the two poles and 2) lower levels of [(3)H]MK-801 binding in the ventral hippocampus, with the lowest value observed in CA1, apparently resulting from a decreased receptor density since the B(max) value was lower in ventral hippocampus. For the AMPA receptors CA1 our results showed in ventral hippocampus compared with dorsal hippocampus: 1) lower levels of mRNA expression for GluRA, GluRB and GluRC subunits, which were more pronounced in CA1 and in dentate gyrus region and 2) lower levels of [(3)H]AMPA binding. Intracellular recordings obtained from pyramidal neurons in CA1 showed longer NMDA receptor-mediated excitatory postsynaptic potentials in ventral hippocampus compared with dorsal hippocampus. In conclusion, the differences in the subunit mRNA and protein expression of NMDA and AMPA receptors as well as the lower density of their binding sites observed in ventral hippocampus compared with dorsal hippocampus suggest that the glutamatergic function differs between the two hippocampal poles. Consistently, the lower value of the ratio NR2A/NR2B seen in the ventral part would imply that the ventral hippocampus NMDA receptor subtype is functionally different than the dorsal hippocampus subtype, as supported by our intracellular recordings. This could be related to the lower ability of ventral hippocampus for long-term synaptic plasticity and to the higher involvement of the NMDA receptors in the epileptiform discharges, observed in ventral hippocampus compared with dorsal hippocampus.     

Regional distribution and properties of [3H]MK-801 binding sites determined by quantitative autoradiography in rat brain

[3H]MK-801 binding in rat brain was characterized using a quantitative autoradiographic binding assay. [3H]MK-801 binding (5 nM) reached equilibrium by 120 min at 23 degrees C. [3H]MK-801 appeared to label a single high affinity site with an affinity constant of approximately 11 nM. [3H]MK-801 binding was heterogeneously distributed throughout the brain with the following order of binding densities: hippocampal formation greater than cortical areas greater than striatum greater than thalamus. Competitive N-methyl-D-aspartate antagonists, DL-2-amino-5-phosphonopentanoic acid, DL-2-amino-7-phosphonoheptanoic acid, 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid, and cis-4-phosphonomethyl-2-piperidine carboxylic acid, inhibited [3H]MK-801 binding. Glycine antagonists, 7-chlorokynurenic acid and kynurenic acid, also inhibited [3H]MK-801 binding. Furthermore, the inhibition of [3H]MK-801 binding by the quinoxalinedione compounds 6-cyano-7-nitroquinoxaline-2,3-dione and 6,7-dinitroquinoxaline-2,3-dione was reversed by glycine. [3H]MK-801 binding was also inhibited by zinc ions [3H]MK-801 binding was enhanced by glycine or N-methyl-D-aspartate. These results demonstrate that [3H]MK-801 can be used in a quantitative autoradiographic assay as a functional probe for the N-methyl-D-aspartate receptor complex.     

Selective vulnerability of hippocampal cornu ammonis 1 pyramidal cells to excitotoxic insult is associated with the expression of polyamine-sensitive N-methyl-d-asparate-type glutamate receptors

Excess glutamate release and stimulation of post-synaptic glutamatergic receptors have been implicated in the pathophysiology of many neurological diseases. The hippocampus, and the pyramidal cell layer of the cornu ammonus 1 (CA1) region in particular, has been noted for its selective sensitivity to excitotoxic insults. The current studies examined the role of N-methyl-d-aspartate (NMDA) receptor subunit composition and sensitivity to stimulatory effects of the polyamine spermidine, an allosteric modulator of NMDA NR2 subunit activity, in hippocampal CA1 region sensitivity to excitotoxic insult. Organotypic hippocampal slice cultures of 8 day-old neonatal rat were obtained and maintained in vitro for 5 days. At this time, immunohistochemical analysis of mature neuron density (NeuN); microtubule associated protein-2(a,b) density (MAP-2); and NMDA receptor NR1 and NR2B subunit density in the primary cell layers of the dentate gyrus (DG), CA3, and CA1 regions, was conducted. Further, autoradiographic analysis of NMDA receptor distribution and density (i.e. [¹²⁵I]MK-801 binding) and spermidine (100 μM)-potentiated [¹²⁵I]MK-801 binding in the primary cell layers of these regions was examined. A final series of studies examined effects of prolonged exposure to NMDA (0.1–10 μM) on neurodegeneration in the primary cell layers of the DG, CA3, and CA1 regions, in the absence and presence of spermidine (100 μM) or ifenprodil (100 μM), an allosteric inhibitor of NR2B polypeptide subunit activity. The pyramidal cell layer of the CA1 region demonstrated significantly greater density of mature neurons, MAP-2, NR1 and NR2B subunits, and [¹²⁵I]MK-801 binding than the CA3 region or DG. Twenty-four hour NMDA (10 μM) exposure produced marked neurodegeneration (∼350% of control cultures) in the CA1 pyramidal cell region that was significantly reduced by co-exposure to ifenprodil or dl-2-Amino-5-phosphonopentanoic acid (APV). The addition of spermidine significantly potentiated [¹²⁵I]MK-801 binding and neurodegeneration induced by exposure to a non-toxic concentration of NMDA, exclusively in the CA1 region. This neurodegeneration was markedly reduced with co-exposure to ifenprodil. These data suggest that selective sensitivity of the CA1 region to excitotoxic stimuli may be attributable to the density of mature neurons expressing polyamine-sensitive NR2B polypeptide subunits.     

The degeneration of the excitatory climbing fibers enhances [3H]MK-801 and [3H]CGP 39653 binding sites in the rat cerebellar cortex.

The effect of a single injection of 3-acetylpyridine (3-AP), which led to a degeneration of the excitatory cerebellar climbing fibers, was studied on the binding of [3H]MK-801, a non-competitive NMDA antagonist, in the rat cerebellar cortex. The same treatment increased also the binding of [3H]CGP 39653, a new NMDA competitive antagonist. Saturation isotherms showed a significant increase of the maximal number of binding sites (Bmax) for [3H]CGP 39653 and [3H]MK-801 (+48 and 36% respectively) with no change in the affinity 4-9 days after the administration of 3-AP. Our data demonstrate that in the cerebellar cortex both NMDA recognition site labelled by [3H]CGP 39653 and its modulatory site labelled by [3H]MK-801 may undergo plastic changes when the glutamatergic receptors and transmission are denervated.     

Hippocampal excitatory amino acid receptors in elderly, normal individuals and those with Alzheimer's disease: non-N-methyl-D-aspartate receptors.

Quantitative receptor autoradiography was used to examine the density and distribution of [3H]kainic acid and [3H]alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) binding sites in the hippocampal formation and parahippocampal gyrus obtained at autopsy from 10 Alzheimer's disease and eight normal control individuals. In control and Alzheimer's disease individuals, [3H]kainic acid saturation binding analysis in the outer molecular layer of the dentate gyrus fitted a single-site model. Added calcium ions did not alter the density of [3H]kainic acid binding in the human tissues. These results suggest that calcium-sensitive high-affinity kainic acid binding sites are not present in the human brain in contrast to kainic acid receptors in the rat brain. [3H]AMPA binding was also slightly different in the human brain as compared to the rat, being greatest in the inner third as compared to the outer two-thirds of the dentate gyrus molecular layer. In both control and Alzheimer's disease individuals, [3H]kainic acid and [3H]AMPA binding densities were similar at anterior and posterior levels of the hippocampal formation. In Alzheimer's disease patients, there was a significant increase in [3H]AMPA binding in the infragranular layer. In some, but not all Alzheimer's disease patients, there was an increase in [3H]kainic acid binding densities in the outer half of the dentate gyrus molecular layer. The same individuals which exhibited an increase in [3H]kainic acid binding in the outer molecular layer also displayed increased [3H]AMPA binding in the hilar region. Similar alterations in [3H]kainic acid binding have been observed in rats which had received fimbria-fornix lesions, a model of chronic epilepsy and in individuals with temporal lobe epilepsy. Advanced Alzheimer's disease patients are at risk of developing seizures. The results suggest that several factors including cortical and subcortical pathology and seizure activity may contribute to the alterations in [3H]kainic acid and [3H]AMPA binding observed in the hippocampal formation in Alzheimer's disease.     

Distribution of nicotinic cholinergic receptors in the rat tel- and diencephalon: a quantitative receptor autoradiographical study using [3H]-acetylcholine, [α-125l]bungarotoxin and [3H]nicotine

The topographical distribution of [α:-¹²T]bungarotoxin [¹²⁵I]BTX, [³H]nicotine ([³H]Nic), [³H]acetylcholine ([³H]ACh) (in the presence of atropine) binding in rat teland diencephalon was investigated using a quantitative receptor autoradiographical technique. With the [³H|ACh and [³H]Nic radioligands, a strong labelling was observed in various thalamic nuclei, including the medial habenula, a moderate labelling in different areas of the cortex cerebri, the nucleus caudatus putamen, the nucleus accumbens and tuberculum olfactorium and a uniform weak labelling in the hypothalamus. When the binding data for [³H]Nic were plotted against binding data for [³H]ACh in various brain nuclei, a significant correlation was obtained. Considering [¹²⁵I]BTX, the strongest labelling was observed in the lateral mammillary nucleus and the hilus gyrus dentatus of the hippocampal formation. A weak labelling occurred in areas such as the nucleus causatus putamen, the thalamus and the cerebral cortex. No significant correlation was therefore obtained between the degree of [¹²⁵I]BTX binding in various brain nuclei and the degree of binding observed with [³H]Nic or [³H]ACh. The present results underline the view that the high-affinity |³H]Nic and [³H]ACh binding sites label the same cholinergic nicotinic receptor binding site, while [¹²⁵I]BTX labels another subpopulation of nicotinic cholinergic receptors, predominantly found in discrete areas of the hypothalamus and the limbic cortex.     

Postischemic binding of [3H]phorbol 12,13-dibutyrate and [3H]inositol 1,4,5-trisphosphate in the gerbil brain: an autoradiographic study.

Postischemic alteration of second messenger systems was investigated in the Mongolian gerbil, utilizing [3H]phorbol 12,13-dibutyrate and [3H]inositol 1,4,5-trisphosphate receptor autoradiography. Transient ischemia was induced for 10 min, and animals were allowed to survive for various recirculation periods of up to one month. [3H]Phorbol 12,13-dibutyrate binding in selectively vulnerable areas showed no significant change 1-24 h after ischemia except for a transient decline in a few regions. Thereafter, the binding in most of the selectively vulnerable areas showed significant alteration 48 h or seven days after ischemia. Interestingly, dentate molecular layer which was resistant to ischemia showed a significant elevation in the number of [3H]phorbol 12,13-dibutyrate binding sites. One month after ischemia, [3H]phorbol 12,13-dibutyrate binding showed significant reduction only in the striatum and the hippocampal CA1 sector where severe neuronal damage was seen morphologically. A significant elevation in the number of [3H]phorbol 12,13-dibutyrate binding sites was still seen in the dentate molecular layer one month after ischemia. In contrast, [3H]inositol 1,4,5-trisphosphate binding showed significant reduction in the selectively vulnerable regions 1-24 h after ischemia. Thereafter, [3H]inositol 1,4,5-trisphosphate binding in most of the selectively vulnerable areas markedly decreased up to one month after ischemia. In the dentate molecular layer, [3H]inositol 1,4,5-trisphosphate binding also showed significant reduction during recirculation except for a slight recovery 48 h and seven days after ischemia. One month after ischemia, the binding in all regions showed significant reduction. These results suggest that postischemic alteration of two second messenger (protein kinase C and inositol 1,4,5-trisphosphate) binding sites was produced with different processes in selectively vulnerable areas.(ABSTRACT TRUNCATED AT 250 WORDS)     

Autoradiographic distribution of [H]nicotine binding in human cortex: Relative abundance in subicular complex

Distinct patterns of [³H]nicotine (3 nm) binding were apparent in various regions of adult human neo- and archicortex. Receptor binding was greatest in the subicular complex—particularly presubiculum—and entorhinal cortex, where it was prominent in the characteristic parvo- and magnocellular islands of these regions and in middle layers of entorhinal cortex. In somatosensory cortex (Brodmann areas 3, 1 and 2) and occipital (area 17) cortex binding was highest in the upper and lower layers, and relatively sparse in the sensory input, layer IV. In primary motor (area 4) and temporal (area 21) cortex, binding in the outer half of the cortical ribbon was denser than that in the inner half and a distinct band was apparent in temporal and cingulate (area 32) in the lower portion of layer III. In prefrontal association cortex the pattern of binding was less distinct although slightly higher in the lower architectonic layers. There was generally little binding in the hippocampus (areas CA1–4) and dentate gyrus with the exception of the stratum lacunosum moleculare in CA2–3 and, to a lesser extent, supra- and subgranule zones of the dentate. These patterns of reactivity, which are distinct from that of the major cortical cholinergic innervation, suggest that the nicotinic receptor, detected using nanomolar concentrations of [³H]nicotine, may primarily be associated with intracortical circuitry in the neocortex. The relatively high density in entorhinal and subicular regions may be related to the extensive phylogenetic development of these regions which has occurred in conjunction with the development of multimodal association circuitry in the human cortex.     

Autoradiographic analysis of [H]ryanodine binding sites in rat brain: Regional distribution and the effects of lesions on sites in the hippocampus

Quantitative and qualitative autoradiographic methods together with lesion approaches were used to determine the distribution of [3H]ryanodine binding sites in rat brain and the neuronal localization of these sites in the hippocampus. In normal animals, levels of [3H]ryanodine binding sites ranged from a low of about 1 fmol/mg tissue in subcortical structures to a high of 12-18 fmol/mg tissue in subregions of the hippocampus and the olfactory bulb. Relatively high densities of sites (5-9 fmol/mg tissue) were also seen in the olfactory tubercle, most areas of the cerebral cortex, accumbens nucleus, striatum, lateral septal nuclei, pontine nucleus, superior colliculus and granule cell layer of the cerebellum. Specific binding was undetectable in white matter. In experimental animals, intracerebral injections of kainic acid caused neuronal degeneration and a near total depletion of [3H]ryanodine binding sites in the dentate gyrus and in fields CA1, CA2 and CA3 of the hippocampus. Injections of kainic acid that left dentate granule cells largely intact while destroying all neurons in field CA3 had no effect on binding sites in the dentate gyrus. However, these lesions substantially reduced the density of binding in field CA3, leaving a narrow band of sites outlining the position of the degenerated CA3 pyramidal cells. Mechanical knife-cut lesions that severed the granule cell mossy fiber input to field CA3 reduced the density of binding sites in the CA3 region. The results indicate that [3H]ryanodine binding sites in brain are heterogeneously distributed and suggest that a proportion of these sites in the hippocampus may be contained in mossy fiber terminals where a presumptive calcium channel/ryanodine receptor complex may be involved in the regulation of calcium mobilization and/or neurotransmitter release.     

Systemic administration of kainic acid induces selective time dependent decrease in [I]insulin-like growth factor I, [I]insulin-like growth factor II and [I]insulin receptor binding sites in adult rat hippocampal formation

Administration of kainic acid evokes acute seizure in hippocampal pathways that results in a complex sequence of functional and structural alterations resembling human temporal lobe epilepsy. The structural alterations induced by kainic acid include selective loss of neurones in CA1–CA3 subfields and the hilar region of the dentate gyrus followed by sprouting and permanent reorganization of the synaptic connections of the mossy fibre pathways. Although the neuronal degeneration and process of reactive synaptogenesis have been extensively studied, at present little is known about means to prevent pathological conditions leading to kainate-induced cell death. In the present study, to address the role of insulin-like growth factors I and II, and insulin in neuronal survival as well as synaptic reorganization following kainate-induced seizure, the time course alterations of the corresponding receptors were evaluated. Additionally, using histological preparations, the temporal profile of neuronal degeneration and hypertrophy of resident astroglial cells were also studied. [¹²⁵I]Insulin-like growth factor I binding was found to be decreased transiently in almost all regions of the hippocampal formation at 12 h following treatment with kainic acid. The dentate hilar region however, exhibited protracted decreases in [¹²⁵I]insulin-like growth factor I receptor sites throughout (i.e. 30 days) the study. [¹²⁵I]Insulin-like growth factor II receptor binding sites in the hippocampal formation were found to be differentially altered following systemic administration of kainic acid. A significant decrease in [¹²⁵I]insulin-like growth factor II receptor sites was observed in CA1 subfield and the pyramidal cell layer of the Ammon's horn at all time points studied whereas the hilar region and the stratum radiatum did not exhibit alteration at any time. A kainate-induced decrease in [¹²⁵I]insulin receptor binding was noted at all time points in the molecular layer of the dentate gyrus whereas binding in CA1–CA3 subfields and discrete layers of the Ammon's horn was found to be affected only after 12 h of treatment.

These results, when analysed with reference to the observed histological changes and established neurotrophic/protective roles of insulin-like growth factors and insulin, suggest possible involvement of these growth factors in the cascade of neurotrophic events that is associated with the reorganization of the hippocampal formation observed following kainate-induced seizures.     

Binding of [3H]inositoltrisphosphate and [3H]phorbol 12,13-dibutyrate in rat hippocampus following transient global ischemia: a quantitative autoradiographic study

An in vitro quantitative autoradiographic binding study of the phosphatidylinositol system ligands [3H]inositol 1,4,5-trisphosphate (IP3) and [3H]phorbol 12,13-dibutyrate (PDBU) to rat brain slices was performed at 6, 12, 24, 28 and 72 h following a 20 min ischemic injury. PDBU binding showed a transient 20% decrease in the dentate gyrus and the CA3 the first 24 h as well as a 50% decrease in the CA1 at 72 h. A 50% decline in IP3 binding was seen in all regions at 6-12 h except the pyramidal cell layer of the CA1. This downregulation of calcium mobilizing intracellular receptors is probably a defence against ischemic neuronal cell death.     

Autoradiographic distribution of phencyclidine receptors in the rat brain using [3H]1-(1-(2-thienyl)cyclohexyl)piperidine ([3H]TCP)

The distribution of phencyclidine (PCP) receptors in the rat brain was determined by autoradiography using 1-(1-(2-thienyl)cyclohexyl)piperidine ([3H]TCP). [3H]TCP appeared to bind to PCP receptors as only PCP-like drugs and sigma-opioids inhibited the binding of [3H]TCP. The areas of the rat brain with the highest density of radiolabeled binding sites were the superficial layers of cerebral cortex, hippocampus and dentate gyrus. Moderate densities of binding sites were found in the medial geniculate nuclei, caudate nucleus, nucleus accumbens, interpeduncular nucleus, superior colliculus, periaqueductal gray and cerebellum. Low densities of binding sites were observed in spinal cord, most of the brainstem, the substantia nigra and most of the hypothalamus.     

3H]TCP binding sites in Alzheimer's disease

Quantitative autoradiography was used to determine the density and distribution of [3H]1-[1-(2-thienyl)-cyclohexyl]piperidine ([3H]TCP) binding sites in human hippocampal tissue sections from control and Alzheimer's disease patients. Some Alzheimer's cases showed no changes in binding site density while other cases showed substantial declines in the CA1 region. [3H]TCP binding in the CA1 region from Alzheimer's patients was reduced an average of 40% while the other hippocampal regions were unaffected. It is proposed that the loss of [3H]TCP sites in the hippocampal CA1 region of certain Alzheimer's cases is associated with the greater cell loss observed in cases of severe Alzheimer's disease.     

Evidence for the presence of a histaminergic neuron system in the rat brain: an immunohistochemical analysis

The binding of calcium antagonists in the rat hippocampal formation was studied using autoradiography. Hippocampal slices were labeled in vitro with [3H]PN 200-110. High densities of binding sites for calcium antagonists were found in the molecular layer of the dentate gyrus and in the CA3 subfield of the hippocampus. After ablation of the granule cells by local injection of colchicine a marked decrease in the number of [3H]PN 200-110 binding sites density was observed on these areas, while binding to other parts of the hippocampal formation and brain was spared. These results strongly suggest the localization of high densities of calcium channels to the granule cells of the dentate gyrus.