Dissociation between changes in glutamate receptor binding sites and their coupling to phosphatidylinositol metabolism following intrahippocampal colchicine injection

Intrahippocampal colchicine injection produces a rapid death of granule cells and pyramidal neurons in the hippocampus in the rat. Under the appropriate assay conditions, [3H]glutamate labels the N-methyl-D-aspartate type of glutamate receptors while [3H]alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate labels the quisqualate type. Unilateral injection of colchicine (15 micrograms) in the dorsal hippocampus did not produce any change in [3H]glutamate and [3H]alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate binding in membrane fractions from the dentate gyrus or CA1 field contralateral to the injection side, at least up to 12 days after the injection. However, it produced a progressive decrease in the binding of both ligands in dentate gyrus and CA1 of the injected hippocampus. In the dentate gyrus the changes in binding as a function of time after the injection were biphasic with a rapid exponential decrease (t1/2 about 8 days for both [3H]glutamate and [3H]alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate) until 12 days after the injection followed by a much slower decrease afterwards. A similar pattern was observed in CA1 although the changes in binding were smaller and delayed by about three days as compared to the dentate gyrus. Kinetic analyses of the binding at equilibrium were performed seven days after the injection and indicated that the changes in [3H]glutamate binding were due to a change in the maximum number of sites but not in affinity for the ligand.(ABSTRACT TRUNCATED AT 250 WORDS)     

The ontogeny of excitatory amino acid receptors in the rat forebrain--II. Kainic acid receptors

The ontogeny of [3H]kainic acid binding in rat forebrain was studied quantitatively using in vitro receptor autoradiography. Specific binding was detectable in ventral thalamus, hippocampus, striatum and olfactory bulb by postnatal day 1. In regions with high densities of receptors in adulthood, such as CA3, dentate gyrus and striatum, binding increased progressively across development peaking at postnatal day 21. In ventral thalamus and the inner lamina of the neocortex, [3H]kainic acid binding was high in the first three postnatal weeks and relatively low thereafter. Saturation studies performed on adults and 14-day-old animals suggest differences in both the affinity and the maximal binding capacity contributed to the observed developmental changes in binding of [3H]kainic acid.     

Chronic amitriptyline administration increases serotonin transporter binding sites in the hippocampus of aged rats

The effects of ageing and of chronic antidepressant treatment upon 5-HT transporter sites ([3H]paroxetine binding) in the rat hippocampus was examined. [3H]paroxetine binding to transporter sites was decreased with ageing in the hippocampus of control rats (38% decrease in dentate gyrus and CA4). Amitriptyline (10 mg/kg, i.p.) had no significant effect on [3H]paroxetine binding in 10 months old rats, but increased binding sites in 24 months rats in all hippocampal subregions (greatest increase of 109% in CA1 compared to saline controls). These data indicate an age-related decrease in hippocampal serotonin transporter sites and upregulation of these sites following 10 weeks of amitriptyline. The observed increase in transporter sites following amitriptyline may contribute to the general lower effectiveness of tricyclic antidepressants with ageing.     

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.     

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)     

Quantitative autoradiographic analysis of glutamate binding sites in the hippocampal formation in normal and schizophrenic brain post mortem

Using quantitative autoradiography, the anatomical distribution of the binding sites (kainate, N-methyl-D-aspartate and quisqualate) for the excitatory neurotransmitter glutamate has been established in the hippocampal formation from control and schizophrenic brains, post mortem. There is a loss of the kainate subtype particularly in schizophrenic hippocampi mainly from the CA4/CA3 mossy fibre termination zone of the cornu ammonis (CA4 and CA3; control and schizophrenic left hippocampus, respectively, 54.2 and 66.6 pmol/g; 18.3 and 17.9 pmol/g), as well as bilateral losses in the dentate gyrus (left 14.2 pmol/g and right 28.0 pmol/g; left 9.5 pmol/g and right 7.9 pmol/g, control and schizophrenic, respectively) and parahippocampal gyrus (left 50.8 pmol/g and right 41.7 pmol/g, left 27.7 pmol/g and right 25.3 pmol/g, control and schizophrenic, respectively). There is complete preservation of N-methy-D-aspartate sites in schizophrenic hippocampi, and a marginally significant loss of the quisqualate binding site in CA4/CA3 regions (left 249 fmol/g and right 306 fmol/g, left 157 fmol/g and right 148 fmol/g, control and schizophrenic, respectively). These findings reflect the possible importance of glutamate in the pathophysiology of schizophrenia and represent novel targets for therapeutic manipulation in schizophrenia.     

Focal stimulation of specific pathways in the rat hippocampus causes a reduction in radioligand binding to the haloperidol-sensitive sigma receptor

Summary Focal electrical stimulation of selected excitatory pathways in the hippocampal slice caused a decrease in the binding of [³H]-1,3-di(2-tolyl)guanidine (DTG) or [³H]-(+)-3-[hydroxyphenyl]-N-(1-propyl)piperidine ((³H)-(+)3-PPP) to haloperidol-sensitive sigma binding sites in the slice. Activation of the mossy fibers or perforant path by high frequency electrical stimulation caused the reduction in [³H]-DTG binding; whereas activation of fibers in the strata radiatum, lacunosummoleculare, alveus, or oriens did not affect [³H]-DTG binding. The decrease in binding observed was calcium-dependent and tetrodotoxin sensitive and varied with the frequency, intensity, and duration of stimulation. Although haloperidol-sensitive [³H]-DTG binding sites are distributed throughout the hippocampus, stimulation of the perforant path or mossy fibers resulted in a significant reduction in binding only in the dentate region of the slice. The decrease in binding following perforant path stimulation was blocked by the glutamate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX); whereas the decrease in binding caused by mossy fiber stimulation was not affected by CNQX or DL-APV. The results obtained support the hypothesis that activation of the granule cells in the hippocampal slice caused the release of an endogenous ligand which acts at the haloperidol-sensitive sigma binding site in the dentate gyrus.     

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.     

Post-ischemic synaptic plasticity in the rat hippocampus after long-term survival: histochemical and autoradiographic study.

The hippocampus provides a suitable area in the brain for the analysis of neuronal plasticity after application of a selective lesioning technique. Using histochemistry and autoradiography, we studied synaptic reorganization in the rat hippocampus with selective CA1 pyramidal cell lesioning caused by transient forebrain ischemia after long-term survival. An autoradiographic study was performed on second messenger systems ([3H]inositol 1,4,5-trisphosphate, [3H]forskolin and [3H]phorbol 12,13-dibutyrate binding). One-hundred days after ischemia, depletion of CA1 pyramidal cells and marked shrinkage of the CA1 subfield was noted in spite of unaltered thickness of the CA3 band and of the dentate molecular layers. Although neuronal density in the CA3 region of animals killed seven days after ischemia was not different from the normal group, 78% of animals showed neuronal loss of 30-50% in the stratum pyramidale of the CA3b 100 days after recirculation. Sixty-seven per cent of animals exhibited supragranular mossy fiber sprouting in the dentate gyrus. However, CA3 neuronal loss did not correlate with mossy fiber sprouting. Succinic dehydrogenase was depleted in the CA1 100 days after ischemia, and animals with CA3 damage showed a reduction of succinic dehydrogenase activity in the CA3. In contrast to the unaltered acetylcholinesterase in the animals killed seven days after ischemia, high density bands of acetylcholinesterase activity in the stratum pyramidale of the CA1 were found to be broadened 100 days after ischemia. In the CA1 subfield, subnormal activity of [3H]phorbol 12,13-dibutyrate and [3H]forskolin binding were observed in spite of the depleted [3H]inositol 1,4,5-triphosphate binding. [3H]Forskolin binding in the hilus had increased by 62% 100 days after ischemia, although binding in the stratum lucidum of the CA3 and in the stratum moleculare of the dentate gyrus was unaltered. However, no visible supragranular increase in [3H]forskolin binding was observed. These results indicate that long-term survival after CA1 pyramidal cell depletion caused by transient forebrain ischemia induced the modulation of neuronal activity and synaptic rearrangements in the whole hippocampal formation.     

Dopamine receptors in human brain: autoradiographic distribution of D2 sites

We have studied the detailed anatomical distribution of D2 receptors in human post mortem brain tissue using quantitative autoradiographic techniques. D2 receptors were labeled using the specific D2 agonist [3H]CV 205-502 and the antagonist [3H]spiroperidol. The pattern of D2 receptor distribution observed with the two ligands was very similar. The highest densities were found in the nucleus caudatus, putamen, nucleus accumbens and olfactory tubercle followed by the substantia nigra, where D2 receptors were mainly concentrated in the pars compacta. Lower but still significant densities were associated with the lateral part of the globus pallidus and CA1 and CA3 fields of the hippocampus. The medial part of the globus pallidus, the dentate gyrus and the amygdala showed low to very low densities of D2 receptors. Almost negligible amounts of binding were observed in the olfactory bulb, diencephalon, brainstem, cerebellum and most parts of the neocortex. Our results are comparable with previously reported localizations of D2 receptors in the human and rat brain. We also report the lack of the so-called spirodecanone binding sites in the human brain. The localization of D2 receptors is compared with the distribution of D1 receptors.     

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.     

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.     

Cholinergic innervation and topographical organization of muscarinic binding sites in rat brain: a comparative autoradiographic study

Employing [3H]hemicholinium-3 ([3H]HC), [3H]pirenzepine([3H]PZ) and [3H]quinuclidinyl benzilate ([3H]QNB), autoradiographic binding studies were performed to identify and quantitate the localization of high-affinity choline carriers, M1-subtype of muscarinic binding sites and a mixed population of M1- and M2-subtypes of muscarinic binding sites, respectively, in 38 anatomically defined areas of rat brain. Labelling of adjacent brain sections with [3H]HC, [3H]PZ and [3H]QNB revealed different topographical binding patterns. [3H]HC binding, which is supposed to reflect cholinergic innervation, was dense in the nucleus accumbens, olfactory tubercle, caudate putamen, basolateral amygdaloid nucleus and the interpeduncular nucleus. Moderate but heterogeneous binding was found in thalamic, hypothalamic, hippocampal and cortical areas. Maximal [3H]PZ binding was observed in the nucleus accumbens, olfactory tubercle and in discrete substructures of the hippocampus, e.g. CA1 and dentate gyrus. Binding to other hippocampal and cortical areas was intermediate, whilst minor binding was found in thalamic, hypothalamic and brain stem areas. The binding of [3H]QNB was more evenly distributed over the brain as compared to that of [3H]PZ. [3H]QNB clearly exceeded the binding of [3H]PZ in the thalamus, hypothalamus and brain stem. A relationship was found between the topography patterns of the [3H]PZ and [3H]QNB binding sites. However, some brain areas showed preference for one of the two ligands, pointing to a distinct localization of M1- and M2-subtypes of muscarinic binding sites. Although M1 sites appeared to predominate in the basal ganglia, hippocampus and cortex, some heterogeneity was observed indicative of the minor occurrence of M2 sites within these structures. There was no relationship between the density of the presumed cholinergic innervation and the binding capacity of either of the muscarinic sites in the various brain areas. However, a relationship was found between M2-selectivity and [3H]HC binding, pointing to a possible presynaptic localization of the M2-sites. In addition, it is suggested that distinct cholinergic cell groups might project their fibres to brain areas containing particular subsets of postsynaptic muscarinic binding sites.     

Enhanced but fragile inhibition in the dentate gyrus in vivo in the kainic acid model of temporal lobe epilepsy: a study using current source density analysis

Temporal lobe epilepsy is related to many structural and physiological changes in the brain. We used kainic acid in rats as an animal model of temporal lobe epilepsy, and studied the neural interactions of the dentate gyrus in urethane-anesthetized rats in vivo. Our initial hypothesis was that sprouting of mossy fibers, the axons of the granule cells, increases proximal dendritic excitatory currents in the inner molecular layer of the dentate gyrus. Extracellular currents were detected in vivo using current source density analysis. Backfiring the mossy fibers in CA3 or orthodromic excitation of the granule cells through the medial perforant path induced a current sink at the inner molecular layer. However, the sink or inferred excitation at the inner molecular layer was not increased in kainic acid-treated rats and the sink actually correlated negatively with the degree of mossy fiber sprouting. It is inferred that the latter sink was mediated mainly by association fibers and not by recurrent mossy fibers. After kainic acid treatment, paired-pulse inhibition of the population spikes in the dentate gyrus was increased. In contrast, reverberant activity that involved looping around an entorhinal-hippocampal circuit was increased in kainic acid-treated rats, compared to control rats. The increase of inhibition in kainic acid-treated rats was readily blocked by a small dose of GABA(A) receptor antagonist bicuculline. The latter dose of bicuculline induced paroxsymal spike bursts in kainic acid-treated but not control rats, demonstrating that the increased inhibition in dentate gyrus was fragile.In conclusion, after kainic acid induced seizures, the dentate gyrus in vivo showed an increase in inhibition that appeared to be fragile. The hypothesized increase in proximal dendritic excitation due to mossy fiber sprouting was not detected. However, the fragile inhibition could explain the seizure susceptibility in patients with temporal lobe epilepsy.     

Limbic seizures cause pronounced changes in the expression of neurokinin B in the hippocampus of the rat.

Immunohistological and in situ hybridization techniques were used to study the influence of kainic acid-induced seizures and of pentylenetetrazol kindling on neurokinin B immunoreactivity and neurokinin B mRNA in the rat hippocampus. Pronounced increases in neurokinin B immunoreactivity were observed in the terminal field of mossy fibres 10-60 days after intraperitoneal injection of kainic acid. These slow but persistent increases in immunoreactivity were accompanied by markedly enhanced expression of neurokinin B mRNA in the granule cells and in hilar interneurons adjacent to the granule cell layer. These changes were preceded by transient increases in neurokinin B mRNA and immunoreactivity in CA1 pyramidal cell layer two and 10 days after kainic acid, which, however, subsided later on. Pentylenetetrazol kindling caused similar increases in neurokinin B mRNA expression in granule cells and in CA1 pyramidal cells, but not in hilar interneurons. In CA1, increased neurokinin B message was present two days after termination of the kindling procedure but not after 10 days. Sixty days after kainic acid injection, neurokinin B immunoreactivity extended to the inner-third of the molecular layer of the dentate gyrus. After pentylenetetrazol kindling, a neurokinin B-immunoreactive band was observed in the infrapyramidal region of CA3. Lesions of the dentate granule cells by local injection of colchicine in kainic acid-treated rats abolished the supragranular neurokinin B-positive staining, whereas it was almost unchanged after transection of the ventral hippocampal commissure. These observations suggest that neurokinin B immunoreactivity may be located in ipsilateral mossy fibres undergoing collateral sprouting to the inner molecular layer or to the infrapyramidal region in CA3, respectively. Preprotachykinin A mRNA, which encodes for neurokinin A and substance P, and substance P immunoreactivity were not changed in the hippocampus of epileptic rats compared with untreated animals. The observed changes in neurokinin B immunoreactivity and mRNA indicate that specific functional and morphological changes may be induced in hippocampal neurons by recurrent limbic seizures.     

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     

Kainic acid-mediated increase of preprotachykinin-A messenger RNA expression in the rat hippocampus and a region-selective attenuation by dexamethasone.

The hippocampus contains the highest number of glucocorticoid-sensitive neurons in the rat brain and excessive exposure to glucocorticoids can cause damage to hippocampal neurons and impair the capacity of the hippocampus to survive neuronal insults. In this study in situ hybridization combined with quantitative image analysis was used to study preprotachykinin-A mRNA levels after administration of a toxic dose of kainic acid in animals pretreated with glucocorticoids. Kainic acid was injected into dorsal hippocampus CA3 region in animals pretreated with the synthetic glucocorticoid receptor agonist dexamethasone and in control animals. Preprotachykinin-A mRNA was not detected in the hippocampus of untreated animals or in animals analysed 30 min after a kainic acid injection. However, 4 h after injection of kainic acid, the level of preprotachykinin-A mRNA increased to 20-times above the detection limit both in the dentate gyrus and the CA3 region of the hippocampus. Treatment of kainic acid-injected animals with dexamethasone 30 min before and 2 h after the injection attenuated the increase in the granule cells of the dentate gyrus by 50%. In contrast, dexamethasone pretreatment had no significant effect on the kainic acid-induced increase of preprotachykinin-A mRNA in pyramidal cells in regions CA3 or CA1. These results show that an excitatory stimulus within the hippocampus causes a substantial increase in the level of preprotachykinin-A mRNA in hippocampal granule and pyramidal cells and suggest that in granule cells of the dentate gyrus this increase can be modulated by glucocorticoids.     

Alterations in the ryanodine receptor calcium release channel correlate with Alzheimer's disease neurofibrillary and beta-amyloid pathologies.

Investigation of the integrity of the ryanodine receptor in Alzheimer's disease is important because it plays a critical role in the regulation of calcium release from the endoplasmic reticulum in brain, impairment of which is believed to contribute to the pathogenesis of Alzheimer's disease. The present study compared ryanodine receptor levels and their functional modulation in particulate fractions from control and Alzheimer's disease temporal cortex, occipital cortex and putamen. Relationships between ryanodine receptor changes and the progression of Alzheimer's disease pathology were determined by examining autoradiographic [3H]ryanodine binding in entorhinal cortex/anterior hippocampus sections from 22 cases that had been staged for neurofibrillary changes and beta-amyloid deposition. A significant (P < 0.02) 40% decrease in the Bmax for [3H]ryanodine binding and significantly higher IC50 values for both magnesium and Ruthenium Red inhibition of [3H]ryanodine binding were detected in Alzheimer's disease temporal cortex particulate fractions compared to controls. Immunoblot analyses showed Type 2 ryanodine receptor holoprotein levels to be decreased by 20% (P < 0.05) in these Alzheimer's disease cases compared to controls. No significant differences were detected in [3H]ryanodine binding comparing control and Alzheimer's disease occipital cortex or putamen samples. The autoradiography study detected increased [3H]ryanodine binding in the subiculum, CA2 and CA1 regions in cases with early (stage I-II) neurofibrillary pathology when compared to Stage 0 cases. Analysis of variance of data with respect to the different stages of neurofibrillary pathology revealed significant stage-related declines of [3H]ryanodine binding in the subiculum (P < 0.02) with trends towards significant decreases in CA1, CA2 and CA4. Post-hoc testing with Fisher's PLSD showed significant reductions (74-94%) of [3H]ryanodine binding in the subiculum, and CA1-CA4 regions of the late isocortical stage (V-VI) cases compared to the early entorhinal stage I-II cases. [3H]Ryanodine binding also showed significant declines with staging for beta-amyloid deposition in the entorhinal cortex (P < 0.01) and CA4 (P < 0.05) with trends towards a significant decrease in the dentate gyrus. We conclude that alterations in ryanodine receptor binding and function are very early events in the pathogenesis of Alzheimer's disease, and may be fundamental to the progression of both neurofibrillary and beta-amyloid pathologies.     

Neuropeptide Y biosynthesis is markedly induced in mossy fibers during temporal lobe epilepsy of the rat

Neuropeptide Y (NPY) immunoreactivity and gene expression was investigated in the hippocampus after kainic acid-induced seizures and pentylenetetrazol kindling in the rat. Pronounced increases of NPY immunoreactivity were found in the terminal field of mossy fibers in both animal models. In kainic acid-treated rats the peptide progressively accumulated in the hilus and the stratum lucidum of CA3, 5-60 days after injection of the toxin and, at the later intervals, extended to the supragranular molecular layer of the dentate gyrus indicating sprouting of these neurons. Unilateral injection of colchicine into the hilus abolished NPY staining of the mossy fibers. Using in situ hybridization, in both animal models markedly enhanced expression of prepro-NPY mRNA was observed in the granular layer, containing the perikarya of the mossy fibers. It is suggested that sustained expression of the neuromodulatory neuropeptide NPY, in addition to the observed plastic changes, may contribute to altered excitability of hippocampal mossy fibers in epilepsy. Neither somatostatin immunoreactivity nor gene expression were enhanced in granule cells/mossy fibers.     

癫痫发作敏感大鼠齿状回苔状纤维侧枝发芽

目的　探讨海马齿状回苔状纤维侧枝发芽与癫痫发作敏感性形成之间的关系。方法　在颈部皮下注射惊厥剂量的海人酸 (KA ,10mg/kg)诱发大鼠出现癫痫发作后 ,采用Timm’s染色法 ,分别在注射KA后3d、7d和 1个月 3个时间点观察致痫大鼠海马齿状回内苔状纤维发芽的情况。结果　Timm’s染色发现 ,注射KA后 7d ,海马齿状回分子层内带和颗粒细胞上层出现苔状纤维的异常发芽 ,注射KA后 1个月海马齿状回内Timm’s染色颗粒颜色加深 ,范围增大。提示海马苔状纤维发芽形成的时间过程与癫痫发作敏感性形成的时间过程一致。结论　海马齿状回分子层内带和颗粒细胞上层出现异常的苔状纤维发芽可能与癫痫发作敏感性形成有关。