J. Ułas

Reduced density of adenosine A1 receptors and preserved coupling of adenosine A1 receptors to G proteins in alzheimer hippocampus: A quantitative autoradiographic study

Binding to adenosine A1 receptors and the status of their coupling to G proteins were studied in the hippocampus and parahippocampal gyrus of Alzheimer individuals and age-matched controls. The binding to A1 receptors was compared with binding to the N-methyl-D-aspartate receptor complex channel-associated sites (labeled with (+)-[3H]5-methyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-imine maleate). In vitro quantitative autoradiography demonstrated a similar anatomical distribution of A1 receptors labeled either with an agonist ((-)-[3H]phenylisopropyladenosine) or antagonist ([3H]8-cyclopentyl-1,3-dipropylxanthine) in the brains of elderly controls. In Alzheimer patients, significant decreases in the density of both agonist and antagonist binding sites were found in the molecular layer of the dentate gyrus. Decreased A1 agonist binding was also observed in the CA1 stratum oriens and outer layers of the parahippocampal gyrus, while reduced antagonist binding was found in the subiculum and CA3 region. Reduced density of the N-methyl-D-aspartate receptor channel sites was found in the CA1 region and parahippocampal gyrus. The reductions in binding to adenosine A1 and N-methyl-D-aspartate receptors were due to a decrease in the density of binding sites (Bmax), and not changes in receptor affinity (KD). In both elderly control and Alzheimer subjects, GTP substantially reduced the density of A1 agonist binding sites with a concomitant increase in the KD values, whereas antagonist binding was unaffected by GTP. The results suggest that adenosine A1 receptor agonists and antagonists recognize overlapping populations of binding sites. Reduced density of A1 receptors in the molecular layer of the dentate gyrus most probably reflects damage of the perforant path input in Alzheimers disease, while altered binding in the CA1 and CA3 regions is probably due to loss of intrinsic neurons. Similar effects of GTP on binding to A1 receptors in control and Alzheimer individuals suggest lack of alterations in coupling of A1 receptors to G proteins in Alzheimers disease, thus supporting the notion of normal receptor coupling to their effector systems in Alzheimers disease.

N-methyl-d-aspartate receptor complex in the hippocampus of elderly, normal individuals and those with Alzheimer's disease

The various ligand binding sites of the N-methyl-D-aspartate receptor complex in the hippocampal formation and parahippocampal gyrus of Alzheimers disease patients and age-matched normal individuals were examined using quantitative autoradiography. The hippocampus and parahippocampal gyrus of the normal elderly brain exhibited virtually identical distributions of L-[3H]glutamate, [3H]5-methyl-10,11-dihydro-5H- dibenzo[a,d]cyclohepten-5,10-iminemaleate ([3H]MK-801), [3H][(+/-)2-carboxypiperazine-4-yl]propyl-1-phosphonic acid ([3H]CPP) and strychnine-insensitive [3H]glycine binding sites (r greater than 0.87) suggesting that binding occurred to different domains of the same receptor macromolecule. The binding of [3H]MK-801 to channel-associated phencyclidine sites appeared to be most severely impaired in Alzheimers disease, especially at the anterior hippocampal level. When the data were averaged and the means for Alzheimers disease and control group compared, a 34% decrease (P less than 0.01) in [3H]MK-801 binding was identified in the CA1 stratum pyramidale and a smaller decrease was found in the dentate gyrus molecular layer, parahippocampal gyrus and subiculum. The CA1 region exhibited a similar 35% reduction (P less than 0.05) in L-[3H]glutamate binding to N-methyl-D-aspartate-sensitive sites. This decrease most probably reflected a decline in receptor density. Binding of [3H]CPP to antagonist-preferring sites and [3H]glycine to glycine modulatory sites did not change significantly. However, a marked intersubject variability in N-methyl-D-aspartate receptor binding was observed in control and Alzheimers disease groups. This variability was not related to age, sex or post mortem delay. Some Alzheimers disease patients showed markedly reduced receptor binding levels, while others showed no changes or even increased binding. The loss of N-methyl-D-aspartate-sensitive sites did not correlate with a loss of neurons in the CA1 region (r = 0.286). Similarly, no correlation between the level of binding to N-methyl-D-aspartate-sensitive sites and the density of neuritic plaques and neurofibrillary tangles was found. Intersubject variability in N-methyl-D-aspartate receptor responses in the Alzheimers disease group may partially explain conflicting reports in the literature on the N-methyl-D-aspartate receptor changes in Alzheimers disease, and imply that caution should be exercised before making any generalizations about receptor changes in Alzheimers disease based on mean values only. The analysis of the individual Alzheimers disease cases may also be valuable in determining the mechanism(s) underlying the disease.

Elevated NMDA receptors in parkinsonian striatum.

Dopamine-glutamate interactions contribute to normal striatal function and have been implicated in neurotoxicity at nigrostriatal dopamine (DA) terminals. The present study examined the striata of idiopathic Parkinsons disease (PD) patients and age-matched controls for regional differences in the DA transporter and binding to N-methyl-D-aspartate (NMDA) receptors. [3H]Mazindol labeling of the DA transporter was reduced by 70-80% in the caudate and putamen of PD patients, with reductions being more extensive dorsally than ventrally. In contrast, L-[3H]glutamate binding to NDMA-sensitive receptors was 20-40% higher in PD cases than in controls. These findings raise the possibility that modifications occur within corticostriatal glutamate synapses of PD patients, possibly as a consequence of reduced nigrostriatal DA activity.

Chronic haloperidol treatment enhances binding to Nmda receptors in rat cortex

Hyperactivity of the dopaminergic system and a hypoglutamatergic state have been hypothesized to underlie schizophrenia. It has also been proposed that neuroleptics may interact not only with the dopaminergic system but also with the glutamatergic system. We found that daily intraperitoneal injections of haloperidol (1 mg kg-1) for 21 days resulted in increased binding (10-20%) to the NMDA type of glutamate receptors in the outer layers of rat parietal cortex. Quantitative receptor autoradiography indicates that the action of haloperidol is regionally specific since no changes in NMDA receptors were found in the hippocampus and thalamus. Our data suggest that haloperidol may exert its antipsychotic effects by enhancement of glutamatergic functions as well as by the blockade of dopamine 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 Alzheimers disease and eight normal control individuals. In control and Alzheimers 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 Alzheimers disease individuals, [3H]kainic acid and [3H]AMPA binding densities were similar at anterior and posterior levels of the hippocampal formation. In Alzheimers disease patients, there was a significant increase in [3H]AMPA binding in the infragranular layer. In some, but not all Alzheimers 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 Alzheimers 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 Alzheimers disease.

Plastic response of hippocampal excitatory amino acid receptors to deafferentation and reinnervation

In vitro autoradiography was used to examine the response of excitatory amino acid receptors in the hippocampus of rat following unilateral lesions of the entorhinal cortex. The density of N-methyl-D-aspartate and quisqualate receptor binding was determined on days 1, 3, 7, 14, 21, 30 and 60 postlesion both ipsilateral and contralateral to the lesion and in unoperated controls. The results are compared to the time-course of deafferentation and reinnervation. The molecular layer of the dentate gyrus contralateral to the lesion is only minimally denervated, but is known to exhibit extensive synapse loss and replacement. N-Methyl-D-aspartate receptor binding density in the contralateral hippocampus increased (10-15% relative to unoperated controls) as early as 3 days postlesion and remained elevated through all postlesion times examined. In contrast, the quisqualate receptors in the contralateral hippocampus were unaffected at all times investigated. In the deafferented molecular layer of the ipsilateral dentate gyrus there was a small transient decrease (15-20%) in the binding density of quisqualate receptors 3 days postlesion. At later postoperative times (30-60 days postlesion) the density of both N-methyl-D-aspartate and quisqualate receptors in the ipsilateral molecular layer was higher (15-50%) than that of unoperated controls. These results indicate that N-methyl-D-aspartate and quisqualate receptors are differentially regulated in response to deafferentation. The rapid decrease in quisqualate (and perhaps also N-methyl-D-aspartate) receptor binding at 3 days postlesion may simply reflect the loss of presynaptic receptors, the turnover of postsynaptic receptors or the down-regulation of postsynaptic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Decreased expression of N-methyl-D-aspartate receptor 1 messenger RNA in select regions of Alzheimer brain.

An antisense oligonucleotide probe was used to examine the expression of gene encoding the obligatory NMDAR1 subunit of the N-methyl-D-aspartate receptor in the hippocampus and adjacent cortical areas (entorhinal and perirhinal cortices) of seven Alzheimer patients and in the same brain regions of seven control individuals. Both groups were matched according to age, sex, cause of death, post mortem delay, and tissue storage time. Densitometric analysis of in situ hybridization autoradiograms revealed a 34% (P<0.05) decrease in NMDAR1 messenger RNA levels in layer III of the entorhinal cortex in Alzheimer brains. Similar deficits. although statistically not significant, were observed in layers II and IV-VI of the entorhinal cortex, and in granule cells of the dentate gyrus. Reduced levels of NMDAR1 messenger RNA were also found in layers II-VI of the perirhinal cortex (41 53% decrease, P<0.02). There were no changes in NMDAR1 messenger RNA expression in the CA1, hilus, or subiculum. Both Alzheimer and control group show substantial intersubject variation in levels of NMDAR1 messenger RNA. The analysis of emulsion-dipped tissue revealed a trend toward a decrease in the number of silver grains overlying individual neurons in the CA1, entorhinal cortex, and granule cell layer of some Alzheimer patients. No significant relationship was detected between the levels of NMDAR1 messenger RNA and post mortem delay, tissue storage, age of the subjects, or mini mental state exam score either in control or Alzheimer individuals. In contrast, a strong inverse correlation between NMDAR1 expression and disease duration was found. These data suggest that reduction in expression of the NMDAR1 gene observed in certain regions of Alzheimer hippocampus and adjacent cortical regions is specific for the disease itself. We postulate that reduced transcript levels may reflect either regional cell loss or anomalies in glutamatergic input to the hippocampus and entorhinal cortex in Alzheimers disease. When followed by changes at the receptor subunit protein level, altered expression of the NMDAR1 gene in Alzheimer brain may contribute, through the formation of N-methyl-D-aspartate receptors with different properties, to the previously reported modified N-methyl-D-aspartate receptor ligand binding, abnormal vulnerability of select neuronal populations to excitotoxic insult, and may also be involved in learning and memory deficits.

Chapter 5 Plasticity of excitatory amino acid receptors: implications for aging and Alzheimer's disease

Publisher Summary In view of the critical role of the N-methyl-D-aspartate (NMDA) and other excitatory amino acid (EAA) receptors in higher brain function, it is essential to understand their properties in the developing, mature, aged, injured, and diseased brain. This chapter develops radioligand binding techniques to examine the various receptor components of the EAA system. NMDA receptors have been found to be important in the process of stabilizing appropriate synaptic contacts during development. In the adult brain, no such role has been established, but it is possible that these receptors at least in part play some role in reactive synaptogenesis. The chapter focuses on the hippocampus, in particular the hippocampal alterations that occur following the loss of the major cortical input to the hippocampus, which originates in the entorhinal cortex. The entorhinal cortex is vulnerable in normal aging and in Alzheimers disease, and lesions of the entorhinal cortex have been widely used as a model system in which to examine synaptic plasticity.