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Alzheimer's disease (AD) is the most common neurodegenerative disease, and is clinically characterized by cognitive disturbances and the accumulation of the amyloid β (Aβ) peptides in plaques in the brain. Recent studies have shown the links between AD and the immediate-early gene Arc (activity-regulated cytoskeleton-associated protein), involved in synaptic plasticity and memory consolidation. For example, AD mouse models show a decreased expression of Arc mRNA in the brain. In additional, acute Aβ application to brain slices leads to a widespread ARC protein diffusion, unlike the normal defined localization to synapses. In this study, we investigated genetic variation in human ARC and the risk of developing AD. To this end, we genotyped 713 subjects diagnosed with AD and 841 controls without dementia. ARC was sequenced in a group of healthy individuals, and seven previously known SNPs and three novel SNPs were identified. Two of the newly found SNPs were intronic and one, +2852(G/A), was located in the 3'UTR. Three tag SNPs were selected, including the novel SNP +2852(G/A), to relate to risk of AD, Mini Mental State Examination (MMSE) scores and cerebrospinal fluid (CSF) biomarker levels of total tau (T-tau), hyperphosphorylated tau181 (P-tau(181)) and Aβ(1-42). The AA genotype of the newly found 3'-UTR SNP +2852(A/G), was associated with a decreased risk of AD (p (c) = 0.005; OR = 0.74; 95 % CI: 0.61-0.89). No associations of single SNPs or haplotypes with MMSE score or CSF biomarkers were found. Here we report a novel ARC SNP associated with a reduced risk of developing AD. To our knowledge, this is the first study associating a gene variant of ARC with any disease. The location of the SNP within the 3'UTR indicates that dendritic targeting of ARC mRNA could be involved in the molecular mechanisms underlying this protective function. However, further investigation of the importance of this SNP for ARC function, ARC processing and the pathology of AD is needed.

Today, cognitive impairment can be successfully treated with acetylcholine esterase inhibitors (AChE-I) in many, but not all, patients with Alzheimer's disease (AD). To investigate the relation between tacrine treatment, inheritance of ApoE epsilon4 alleles, and rate of progression, the differences in MMSE and CIBIC scores (efficacy parameters) after 6 and 12 months of tacrine (an AChE-I) treatment were investigated in 145 AD patients. Of these, 84 were ApoE epsilon4-positive (ApoE4) and 61 were ApoE epsilon4-negative (ApoE2-3). No differences were found after 6 months of treatment, but after 12 months the CIBIC scores revealed that the ApoE4 patients had declined more than the ApoE2-3 patients (p < 0.05). No differences were found for the last 6 months of treatment. The results primarily suggest a faster rate of decline in the ApoE4 AD compared to the ApoE2-3, but may also reflect that ApoE epsilon4 genotype inheritance is a negative predictor of treatment effect of tacrine in AD patients.

There is a latency period of several weeks before the onset of clinical effect of antidepressant drugs. The detailed mechanisms underlying drug-induced adaptive neuronal changes are not known. To elucidate the involvement of changes in gene expression of candidate transcription factors, we treated rats for 21 days with buspirone, fluoxetine, 8-OH-DPAT and moclobemide. In situ hybridization was used to study mRNAs encoding NGFI-A, NGFI-B and the glucocorticoid receptors, MR and GR. NGFI-A mRNA expression increased profoundly in the hippocampal formation and the cerebral cortex after all drug treatments, especially after moclobemide treatment (77-122% increase); with the exception of buspirone. MR mRNA expression was induced in hippocampal CA1/CA2 subregions (27-37%) by all antidepressants, while moclobemide and 8-OH-DPAT significantly increased GR gene expression mainly in the CA1 region (31-44%). NGFI-B mRNA was significantly decreased in the hippocampal CA3 subfield (23%) and restrosplenial granular cortex (38%) by moclobemide treatment. There are selective effects of antidepressant drugs on specific transcription factors. These may be important for adaptive neuronal and neuroendocrine changes after antidepressant treatment including HPA axis negative feedback regulation.

A polymorphism consisting of a deletion near the 5' splice site of exon 18 on the alpha2-macroglobulin (A2M) gene (A2M-2) has been suggested to be associated with Alzheimer's disease (AD) in family-based studies. We studied the A2M-2 allele together with the ApoE alleles in a large series on patients with AD (n = 449) and age-matched controls (n = 349). Neuropathologically confirmed diagnoses were available in 199 cases (94 AD and 107 control cases). We found no increase in A2M-2 genotype or allele frequencies in AD (27.5% and 14.6%) versus controls (26.4% and 14.9%). In contrast, a marked increase (p < 0.0001) in ApoE epsilon4 genotype or allele frequencies was found in AD (66.6% and 41.2%) as compared with controls (29.8% and 16.5%), suggesting sufficient statistical power in our sample. No relation was found between the A2M-2 and the ApoE epsilon4 allele. No change in A2M exon 17-18 mRNA size or sequence or A2M protein size was found in cases carrying the A2M-2 deletion, suggesting that there is no biological consequences of the A2M intronic deletion. No change in A2M protein level in cerebrospinal fluid was found in AD, suggesting that the A2M-2 allele does not effect the A2M protein expression in the brain. The lack of an association between the A2M-2 allele and AD in the present study, and the lack of abnormalities in the A2M mRNA or protein suggest that the A2M-2 allele is not associated with AD.

The presynaptically located gamma-aminobutyric acid (GABA) transporter (GAT-1) was studied in a group of patients with Alzheimer's disease (AD) and in a control group using the GAT-1 selective radioligand [3H]tiagabine. Post mortem brain tissue from frontal cortex, temporal cortex, and caudate nucleus from 18 AD patients and 23 age-matched controls were studied. The binding was saturable (Kd 26 nM) and region specific. There were no significant differences between the groups with respect to the binding capacity (Bmax) and binding affinity (Kd). The unaltered [3H]tiagabine binding to GAT-1 protein indicates that intrinsic GABA neurons are spared in Alzheimer's disease.

In this study, drug inhibition and saturation experiments on the binding of the highly selective 5-HT4 antagonist [3H]GR 113808 were performed in human brain membranes so as to better characterize this binding site. Drug competition studies were carried out by incubating 0.2 nM [3H]GR 113808 in the presence of increasing concentrations of six different drugs, i.e. 5-HT, 5-CT, ondansetron, tropisetron, BIMU 1 and BIMU 8 (mixed 5-HT3 and 5-HT4 agonists). The binding displaced by 5-HT showed a drug inhibition constant (Ki) value of 197nM. The use of 5-CT or ondansetron also showed the existence of single-site models albeit with Ki values in the micromolar range (11,5 microM). Tropisetron, BIMU 1 and BIMU 8 displaced bound [3H]GR 113808 according to a two-site binding model, with the high affinity component in the nanomolar range and the low affinity site in the micro or milimolar range. Saturation experiments revealed high binding densities in basal ganglia (187 fmol/mg in putamen, and 149 fmol/mg in caudate nucleus), while lower densities were observed in cortical regions (49 fmol/mg in temporal cortex, 45 fmol/mg in parietal cortex and 71 fmol/mg in cingulate cortex). The apparent affinity (Kd) was similar in the brain regions studied, ranging from 0.13 to 0.34 nmol/l. Despite the enrichment of 5-HT receptors in human brain, their functional correlate in brain diseases remains to be clarified.

The effect of age on the monoamines 5-hydroxytryptamine (5-HT), noradrenaline (NA) and dopamine (DA), their metabolites 5-hydroxyindoleacetic acid (5-HIAA), homovanillic acid (HVA), 3,4-dihydroxphenylacetic acid (DOPAC), and the 5-HT precursor 5-hydroxy-L-tryptophan (5-HTP), together with the peptides neuropeptide Y (NPY), somatostatin (SOM), and corticotropin-releasing factor (CRF), was studied in frontal cortex, gyrus cinguli and hypothalamus from 23 healthy control subjects, aged 16-75 years. After correcting for postmortem interval, significant decreases in gyrus cinguli NA, NPY and CRF, and hypothalamic DA, HVA, and 5-HIAA concentrations were obtained with advancing age. The involvement of the monoaminergic system in several functional abnormalities appearing in senescence is suggested. Furthermore, evidence is given of the participation of the peptidergic systems in the aging process.

The binding of [3H]paroxetine and [3H]citalopram to the human brain serotonin (5-HT) uptake site has been characterized and compared. Our results reveal that the binding exclusively involved with the 5-HT uptake site is identical for both [3H]ligands. The selective 5-HT uptake inhibitor citalopram displays the highest affinity for this uptake site, as compared with the affinities obtained for desipramine and norzimeldine, which is in accordance with their respective blockage of 5-HT uptake. Similar Bmax values were obtained for both radioligands in the brain regions studied, indicating their binding to the same presynaptic membrane protein. Together these findings suggest that both [3H]paroxetine and [3H]citalopram are good markers of the 5-HT transporter as both bind selectively and with high affinity to the serotonin uptake sites. However, the higher affinity of [3H]paroxetine confirms that this compound is the best radioligand for the 5-HT uptake site available today.

Several reports have shown heterogeneity of [3H]imipramine binding to brain membranes. Recently, a high affinity and 5-HT sensitive [3H]imipramine binding site of protein nature, that was suggested to be identical to the substrate recognition site for 5-HT uptake, was demonstrated. Since most studies on the regulation of the [3H]imipramine binding sites by antidepressants have used desipramine displaceable binding, which is heterogenous in nature and contains binding not related to 5-HT uptake sites, the present report studies the possible effects of chronic (3 weeks) administration of imipramine or norzimeldine (10 mg/kg intraperitoneally twice daily) on 5-HT sensitive [3H]imipramine binding sites. For comparison, desipramine sensitive binding was also studied, as well as the physiological correlate 5-HT uptake. There were no changes in either [3H]imipramine binding or 5-HT uptake after the antidepressant treatment.

The binding of (3H)5-hydroxytryptamine ([3H]5-HT, serotonin) to crude homogenates of brains from three different strains of mice has been studied. The strains, C57/BL, DBA and BALB did not show significant differences in the binding characteristics, with Kd values around 6-7 nM and Bmax 270-310 fmoles/mg protein. Various methodological aspects were investigated and found to be important for the binding assay. The presence of ascorbic acid (5.7 mM) thus caused a significant increase in Bmax by 30% without any change in the Kd values. This increase seemed to be due to a decrease in the non-specific binding rather than to an increase in the total binding of serotonin. The presence of a specific MAO-A inhibitor, clorgyline (10 microM), during the assay, resulted in a significant reduction of Bmax by 20% without any change in the affinity for serotonin, when tissue which had been frozen was used. Less than 2% of added serotonin was metabolized during the binding procedure in the absence of clorgyline. Thus, the decrease in binding capacity caused by clorgyline, may be due to a non-competitive blocking effect of the serotonin binding structure. These results indicate that the use of MAO inhibitors in (3H)5-HT binding assays on frozen tissue, rather than being necessary, might negatively affect the result. Neither storage of the brains at -70 degrees nor postmortem storage of the animals for 60 hours at 4 degrees resulted in obvious changes in the (3H)5-HT binding characteristics.

The effect of age upon monoamine oxidase -A and -B (MAO-A and -B) in 23 different, regions of human brain was determined. There was a significant positive correlation with age in 19 out of 23 regions for MAO-B, but no positive correlation with age was found for MAO-A. The increased MAO-B activity was found, in 5 out of 5 regions tested, to be due entirely to an increased enzyme concentration, rather than due to an increased molecular turnover number of the enzyme. The responses of the mitochondrial marker enzymes succinate dehydrogenase (SDH) and malate dehydrogenase (MDH) were studied in 5 brain regions, and no consistent change in activity found with age. The lysosomal enzyme acid phosphatase was found to tend towards an increased activity with age. No difference in either the specific activities or molecular characteristics of MAO were found between men and women. Cross-correlation studies of the data, after compensation for the effects of age, indicated that the activities of the two enzyme forms are under some form of organized control across the whole brain. Such a finding is consistent with a genetic regulation of the enzyme forms.

The binding of [3H]-5-hydroxytryptamine ([3H]-5-HT) to homogenates of human brain has been studied. The specific binding is saturable, with a Kd (frontal cortex) of 12 /+- 2 nM, and is inhibited by non-radioactive 5-HT (IC50=26 nM) and D-Lysergic acid diethylamide (IC50=20 nM). Specific, but not non-specific binding of [3H]-5-TH was inhibited by incubation of the homogenates at 50 degrees C. The binding of [3H]-5-HT across the human brain was not uniform, the highest binding being found in the substantia nigra and hippocampus, and the lowest in the thalamus and pons. The Kd of the binding sites towards 5-HT did, however, appear to be similar for the different brain regions.