Long-term effects of maternal choline supplementation on CA1 pyramidal neuron gene expression in the Ts65Dn mouse model of Down syndrome and Alzheimer's disease.

Choline is critical for normative function of 3 major pathways in the brain, including acetylcholine biosynthesis, being a key mediator of epigenetic regulation, and serving as the primary substrate for the phosphatidylethanolamine N-methyltransferase pathway. Sufficient intake of dietary choline is critical for proper brain function and neurodevelopment. This is especially important for brain development during the perinatal period. Current dietary recommendations for choline intake were undertaken without critical evaluation of maternal choline levels. As such, recommended levels may be insufficient for both mother and fetus. Herein, we examined the impact of perinatal maternal choline supplementation (MCS) in a mouse model of Down syndrome and Alzheimer's disease, the Ts65Dn mouse relative to normal disomic littermates, to examine the effects on gene expression within adult offspring at ∼6 and 11 mo of age. We found MCS produces significant changes in offspring gene expression levels that supersede age-related and genotypic gene expression changes. Alterations due to MCS impact every gene ontology category queried, including GABAergic neurotransmission, the endosomal-lysosomal pathway and autophagy, and neurotrophins, highlighting the importance of proper choline intake during the perinatal period, especially when the fetus is known to have a neurodevelopmental disorder such as trisomy.-Alldred, M. J., Chao, H. M., Lee, S. H., Beilin, J., Powers, B. E., Petkova, E., Strupp, B. J., Ginsberg, S. D. Long-term effects of maternal choline supplementation on CA1 pyramidal neuron gene expression in the Ts65Dn mouse model of Down syndrome and Alzheimer's disease.

CA1 pyramidal neuron gene expression mosaics in the Ts65Dn murine model of Down syndrome and Alzheimer's disease following maternal choline supplementation.

Although there are changes in gene expression and alterations in neuronal density and afferent inputs in the forebrain of trisomic mouse models of Down syndrome (DS) and Alzheimer's disease (AD), there is a lack of systematic assessments of gene expression and encoded proteins within individual vulnerable cell populations, precluding translational investigations at the molecular and cellular level. Further, no effective treatment exists to combat intellectual disability and basal forebrain cholinergic neurodegeneration seen in DS. To further our understanding of gene expression changes before and following cholinergic degeneration in a well-established mouse model of DS/AD, the Ts65Dn mouse, we assessed RNA expression levels from CA1 pyramidal neurons at two adult ages (∼6 months of age and ∼11 months of age) in both Ts65Dn and their normal disomic (2N) littermates. We further examined a therapeutic intervention, maternal choline supplementation (MCS), which has been previously shown to lessen dysfunction in spatial cognition and attention, and have protective effects on the survival of basal forebrain cholinergic neurons in the Ts65Dn mouse model. Results indicate that MCS normalized expression of several genes in key gene ontology categories, including synaptic plasticity, calcium signaling, and AD-associated neurodegeneration related to amyloid-beta peptide (Aß) clearance. Specifically, normalized expression levels were found for endothelin converting enzyme-2 (Ece2), insulin degrading enzyme (Ide), Dyrk1a, and calcium/calmodulin-dependent protein kinase II (Camk2a), among other relevant genes. Single population expression profiling of vulnerable CA1 pyramidal neurons indicates that MCS is a viable therapeutic for long-term reprogramming of key transcripts involved in neuronal signaling that are dysregulated in the trisomic mouse brain which have translational potential for DS and AD.

Early involvement of synapsin III in neural progenitor cell development in the adult hippocampus.

Synapsin III is a synaptic vesicle-associated protein that is expressed in cells of the subgranular layer of the hippocampal dentate gyrus, a brain region known to sustain substantial levels of neurogenesis into adulthood. Here we tested the hypothesis that synapsin III plays a role in adult neurogenesis with synapsin III knockout and wild-type mice. Immunocytochemistry of the adult hippocampal dentate gyrus revealed that synapsin III colocalizes with markers of neural progenitor cell development (nestin, PSA-NCAM, NeuN, and Tuj1) but did not colocalize with markers of mitosis (Ki67 and PCNA). Because neurogenesis consists of a number of stages, the proliferation, survival, and differentiation of neural progenitor cells were systematically quantitated in the hippocampal dentate gyrus of adult synapsin III knockout and wild-type mice. We found a 30% decrease in proliferation and a 55% increase in survival of neural progenitor cells in synapsin III knockout mice. We also observed a 6% increase in the number of neural progenitor cells that differentiated into neurons. No difference in the volume of the dentate gyrus was observed between synapsin III knockout and wild-type mice. Collectively, our results demonstrate a novel role for synapsin III in regulating the proliferation of neural progenitor cells in the adult hippocampal dentate gyrus. These findings suggest a distinct function for this synaptic vesicle protein, in addition to its role in neurotransmission.

BDNF Val66Met variant and age of onset in schizophrenia.

Brain-derived neurotrophic factor (BDNF) has been advanced as a candidate gene for schizophrenia by virtue of its effects on neurotransmitter systems that are dysregulated in psychiatric disorder and its involvement in the response to antipsychotic drugs. The extensively examined BDNF gene Val66Met (or rs6265) variant has been associated with schizophrenia, and studies have linked this polymorphism to brain morphology, cognitive function, and psychiatric symptoms in schizophrenia. Moreover the BDNF Val66Met variant has been reported to be associated with age of onset in schizophrenia. Genotyping of African-American subjects with schizophrenia for five BDNF coding region single nucleotide polymorphisms revealed variance only at the Val66Met allele. The results of statistical analyses indicate a relationship between the BDNF Val66Met genotype and the ages of first psychiatric hospitalization and first schizophrenia symptoms.

Investigation of the phenylalanine hydroxylase gene and tardive dyskinesia.

Phenylketonuria (PKU), an inborn error of phenylalanine metabolism, has been shown to be a risk factor for tardive dyskinesia (TD). In male psychiatric patients there was a significant relationship between TD and measures of plasma phenylalanine following ingestion of a standardized phenylalanine dose that was indicative of higher brain availability of phenylalanine in patients with TD. In addition, a medical food formulation consisting of branched chain amino acids, which compete with phenylalanine for transport across the blood-brain barrier, has been demonstrated to be an efficacious treatment for TD. Cumulatively these findings suggested that TD was related to phenylalanine metabolism and thus that sequence variants in the gene for phenylalanine hydroxylase (PAH), the rate-limiting enzyme in the catabolism of phenylalanine, could be associated with TD susceptibility. Genetic screening of PAH in a group of 123 psychiatric patients revealed ten sequence polymorphisms and two mutations, but none appeared to be a significant risk factor for TD.

Evidence for a tetrahydrobiopterin deficit in schizophrenia.

Tetrahydrobiopterin (BH(4)) is a vital cofactor maintaining availability of the amine neurotransmitters [dopamine (DA), noradrenaline (NA), and serotonin (5-HT)], regulating the synthesis of nitric oxide (NO) by nitric oxide synthase (NOS), and stimulating and modulating the glutamatergic system (directly and indirectly). These BH(4) properties and their potential relevance to schizophrenia led us to investigate the hypothesis of a study group (healthy controls, n=37; schizophrenics, n=154) effect on fasting plasma total biopterin levels (a measure of BH(4)). Study analysis showed a highly significant deficit of total biopterins for the schizophrenic sample after partialling out the effects of potential confounds of gender, age, ethnicity, neuroleptic use history and dose of current use, 24-hour dietary phenylalanine/protein ratio (a dietary variable relevant to BH(4) synthesis), and plasma phenylalanine (which stimulates BH(4) synthesis). A mean decrement of 34% in plasma total biopterins for schizophrenics from control values supports clinical relevance for the finding. In a subsample (21 controls and 23 schizophrenics), sequence analysis was done of the GTP cyclohydrolase I feedback regulatory gene and no mutations were found in the coding region of the gene. A deficiency of BH(4) could lead to hypofunction of the systems of DA, NA, 5-HT, NOS/NO, and glutamate, all of which have been independently implicated in schizophrenia psychopathology. Further, evidence has been accumulating which implicates the critical interdependence of these neurotransmitter systems in schizophrenia; this concept, along with the present study finding of a biopterin deficit, suggests that further study of the BH(4) system in schizophrenia is warranted and desirable.

Branched chain amino acid treatment of tardive dyskinesia in children and adolescents.

BACKGROUND: A series of studies had demonstrated that deficient clearance of the large neutral amino acid phenylalanine was associated with tardive dyskinesia (TD), that the administration of the branched chain amino acids (BCAA) significantly decreased TD symptoms over placebo, and that the observed TD symptom reduction was significantly correlated with a diminished availability of phenylalanine to the brain of adult men with psychosis. As part of an initiative by the National Institute of Mental Health to expand the testing of treatments that were successful in adults to children and adolescents, the present pilot study was undertaken to test whether the BCAA would also reduce TD symptoms in children and adolescents. A 2-week trial of the BCAA was thus conducted in 6 children and adolescents (age range, 10.5-16.5 years) for the treatment of TD symptoms. METHOD: A clinical diagnosis of TD was made in all subjects on the basis of a global score derived from the Simpson Abbreviated Dyskinesia Rating Scale. Subjects were videotaped for TD evaluation at baseline and after 1 and 2 weeks of BCAA treatment given in the form of a drink administered 3 times daily. TD symptom change over the trial period was evaluated by researchers blinded to the treatment status of the evaluation. RESULTS: TD symptom decreases were substantial in 5 of the 6 participants, ranging from 40% to 65%. Two of the subjects received an additional course of treatment, and further reductions in TD symptoms over those seen in the 2-week trial were observed. CONCLUSION: The substantial symptom decrease and tolerability observed suggest the use of the BCAA formulation for the treatment of TD in children and adolescents and warrant further large-scale studies.

Efficacy of the branched-chain amino acids in the treatment of tardive dyskinesia in men.

OBJECTIVE: The efficacy of the branched-chain amino acids in the treatment of tardive dyskinesia in men with psychiatric disorders was tested. METHOD: Public-sector psychiatric patients with long histories of antipsychotic treatment and presumably long-standing tardive dyskinesia were randomly assigned to receive branched-chain amino acids or placebo. Treatment frequency was three times a day, 7 days a week for 3 weeks. The efficacy measure was a frequency count of videotaped tardive dyskinesia movements. RESULTS: A robust and highly significant difference was observed between patients who received high-dose branched-chain amino acids (222 mg/kg of body weight t.i.d.) (N=18) and those who received placebo (N=18) in the percent change in tardive dyskinesia symptoms from baseline to the end of the 3-week trial. Significant and marked differences were seen between the two groups at the >/=30% and >/=60% levels of decrease in tardive dyskinesia symptoms. No clinically significant differences were seen between the pre- and posttrial results of physical examinations and laboratory screening tests. Minimal gastrointestinal symptoms occurred during the trial. The reduction in tardive dyskinesia symptoms in the amino acids group was not related to changes in antipsychotic and glucose plasma levels. A mechanism of response related to decreased amine neurotransmitter synthesis was suggested by the significant positive correlations observed between decreases in tardive dyskinesia symptoms and decreases in aromatic amino acid plasma concentrations over the course of the trial. CONCLUSIONS: Branched-chain amino acids constitute a novel, safe treatment for tardive dyskinesia, with a strong potential for providing significant improvement in the diseased physiognomy of the afflicted person.

Phenylalanine hydroxylase gene in psychiatric patients: screening and functional assay of mutations.

BACKGROUND: Reports relating phenylalanine kinetics and metabolism to psychiatric disorders led us to undertake the comprehensive screening of the phenylalanine hydroxylase (PAH) coding region and functional testing of discovered mutations in a sample of psychiatric patients and healthy control subjects. METHODS: Genomic DNA from psychiatric patients and control subjects was assayed for sequence variants in all PAH coding regions and splice junctions. In vivo functional analysis of mutations was conducted by assessing the kinetics and conversion to tyrosine of a standardized phenylalanine dose and by measuring fasting pterin levels. RESULTS: A known missense mutation was observed in a schizoaffective subject, and a novel missense mutation was discovered in four subjects with schizophrenia and one normal subject. The schizoaffective patient heterozygous for the known A403V mutation showed the lowest rate of phenylalanine kinetics and lowest conversion to tyrosine in the patient sample. The four schizophrenic patients heterozygous for the novel K274E mutation showed significantly decreased phenylalanine kinetics, reduced conversion to tyrosine, and increased synthesis of the PAH cofactor tetrahydrobiopterin compared with schizophrenic subjects without the mutation. CONCLUSIONS: The study findings suggest that larger scale studies are warranted to test the relationship of the PAH genotype with a psychiatric phenotype.

Aromatic amino acid hydroxylase genes and schizophrenia.

Phenylalanine hydroxylase (PAH), which catalyzes the conversion of phenylalanine to tyrosine, shares physical, structural and catalytic properties with tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) that catalyze the rate-limiting steps in the biosynthesis of the neurotransmitters dopamine, noradrenaline, and serotonin. Because these neurotransmitter systems have all been implicated in the pathophysiology of schizophrenia, the aromatic amino acid hydroxylases are among the likely candidates for genes associated with schizophrenia. A mutation in the functionally critical tetrahydrobiopterin cofactor binding domain of the PAH gene had been identified in African-American patients with the diagnosis of schizophrenia, and biochemical analyses suggested that this mutation has physiological consequences related to amine neurotransmitter function. DNA sequencing of the highly conserved cofactor binding domain of the PAH, TH, and TPH genes in African-American subjects with schizophrenia and unrelated, never mentally ill subjects from the NIMH Schizophrenia Genetics Initiative, was undertaken to assess the concordance of mutant genotype with psychiatric phenotype. The K274E mutation was observed in the PAH gene cofactor binding domain, and several polymorphisms were identified in adjacent intronic regions of the PAH, TH, and TPH genes. All of the genetic variants observed were represented in the schizophrenia group and in the never mentally ill group. Genetic evaluation of the family members of subjects with the PAH K274E mutation showed that all individuals with the K274E mutation also exhibited the PAH L321L polymorphism in the catalytic domain of the PAH enzyme.