Faculty Bibliography

Clinical neuropathologic studies suggest that the selective vulnerability of hippocampal CA1 pyramidal projection neurons plays a key role in the onset of cognitive impairment during the early phases of Alzheimer's disease (AD). Disruption of this neuronal population likely affects hippocampal pre- and postsynaptic efficacy underlying episodic memory circuits. Therefore, identifying perturbations in the expression of synaptic gene products within CA1 neurons prior to frank AD is crucial for the development of disease modifying therapies. Here we used custom-designed microarrays to examine progressive alterations in synaptic gene expression within CA1 neurons in cases harvested from the Rush Religious Orders Study who died with a clinical diagnosis of no cognitive impairment (NCI), mild cognitive impairment (MCI, a putative prodromal AD stage), or mild/moderate AD. Quantitative analysis revealed that 21 out of 28 different transcripts encoding regulators of synaptic function were significantly downregulated (1.4-1.8 fold) in CA1 neurons in MCI and AD compared to NCI, whereas synaptic transcript levels were not significantly different between MCI and AD. The downregulated transcripts encoded regulators of presynaptic vesicle trafficking, including synaptophysin and synaptogyrin, regulators of vesicle docking and fusion/release, such as synaptotagmin and syntaxin 1, and regulators of glutamatergic postsynaptic function, including PSD-95 and synaptopodin. Clinical pathologic correlation analysis revealed that downregulation of these synaptic markers was strongly associated with poorer antemortem cognitive status and postmortem AD pathological criteria such as Braak stage, NIA-Reagan, and CERAD diagnosis. In contrast to the widespread loss of synaptic gene expression observed in CA1 neurons in MCI, transcripts encoding beta-amyloid precursor protein (APP), APP family members, and regulators of APP metabolism were not differentially regulated in CA1 neurons across the clinical diagnostic groups. Taken together, these data suggest that CA1 synaptic gene dysregulation occurs early in the cascade of pathogenic molecular events prior to the onset of AD, which may form the basis for novel pharmacological treatment approaches for this dementing disorder. This article is part of a Special Issue entitled 'Neurodegenerative Disorders'.

In the Down syndrome (DS) population, there is an early incidence of dementia and neuropathology similar to that seen in sporadic Alzheimer's disease (AD), including dysfunction of the basal forebrain cholinergic neuron (BFCN) system. Using Ts65Dn mice, a model of DS and AD, we examined differences in the BFCN system between male and female segmentally trisomic (Ts65Dn) and disomic (2N) mice at ages 5-8 months. Quantitative stereology was applied to BFCN subfields immunolabeled for choline acetyltransferase (ChAT) within the medial septum/vertical limb of the diagonal band (MS/VDB), horizontal limb of the diagonal band (HDB) and nucleus basalis of Meynert/substantia innominata (NBM/SI). We found no sex differences in neuron number or subregion area measurement in the MS/VDB or HDB. However, 2N and Ts65Dn females showed an average 34% decrease in BFCN number and an average 20% smaller NBM/SI region area compared with genotype-matched males. Further, relative to genotype-matched males, female mice had smaller BFCNs in all subregions. These findings demonstrate that differences between the sexes in BFCNs of young adult Ts65Dn and 2N mice are region and genotype specific. In addition, changes in post-processing tissue thickness suggest altered parenchymal characteristics between male and female Ts65Dn mice.

The signature sequence amplification method (SSAM) described herein is an approach for amplifying noncoding RNA (ncRNA), microRNA (miRNA), and small polynucleotide sequences. A key point of the SSAM technology is the generation of signature sequences. The signature sequences include target sequences (miRNA, ncRNA, and/or any small polynucleotide sequence) flanked by two DNA fragments. Target sequences can be amplified through DNA synthesis, RNA synthesis, or the combination of DNA and RNA synthesis. The amplification of signature sequences provides an efficient and reproducible mechanism to determine the presence or absence of the target miRNAs/ncRNAs, to analyze the quantities of the miRNAs in biological samples, and for miRNA/ncRNA profiling.

In addition to intellectual disability, individuals with Down syndrome (DS) exhibit dementia by the third or fourth decade of life, due to the early onset of neuropathological changes typical of Alzheimer's disease (AD). Deficient ontogenetic neurogenesis contributes to the brain hypoplasia and hypocellularity evident in fetuses and children with DS. A murine model of DS and AD (the Ts65Dn mouse) exhibits key features of these disorders, notably deficient ontogenetic neurogenesis, degeneration of basal forebrain cholinergic neurons (BFCNs), and cognitive deficits. Adult hippocampal (HP) neurogenesis is also deficient in Ts65Dn mice and may contribute to the observed cognitive dysfunction. Herein, we demonstrate that supplementing the maternal diet with additional choline (approximately 4.5 times the amount in normal rodent chow) dramatically improved the performance of the adult trisomic offspring in a radial arm water maze task. Ts65Dn offspring of choline-supplemented dams performed significantly better than unsupplemented Ts65Dn mice. Furthermore, adult hippocampal neurogenesis was partially normalized in the maternal choline supplemented (MCS) trisomic offspring relative to their unsupplemented counterparts. A significant correlation was observed between adult hippocampal neurogenesis and performance in the water maze, suggesting that the increased neurogenesis seen in the supplemented trisomic mice contributed functionally to their improved spatial cognition. These findings suggest that supplementing the maternal diet with additional choline has significant translational potential for DS.

ABSTRACT: Hippocampal precursor of nerve growth factor (proNGF)/NGF signaling occurs in conjunction with beta-amyloid (Abeta) accumulations in Alzheimer disease (AD). To assess the involvement of this pathway in AD progression, we quantified these proteins and their downstream pathway activators in postmortem tissues from the brains of subjects with no cognitive impairment (NCI), mild cognitive impairment (MCI), and AD using immunoblotting and ELISA. Hippocampal proNGF was significantly greater in AD cases compared with those in NCI and MCI cases. TrkA was significantly reduced in MCI compared with those in NCI and AD, whereas p75 neurotrophin receptor, sortilin, and neurotrophin receptor homolog 2 remained stable. Akt decreased from NCI to MCI to AD, whereas phospho-Akt and phospho-Akt-to-Akt ratio were elevated in AD compared with those in MCI and NCI. No differences were found in phospho-Erk, Erk, or their ratio across groups. Although c-jun kinase (JNK) remained stable across groups, phospho-JNK and the phospho-JNK-to-JNK ratio increased significantly in AD compared with those in NCI and MCI. Expression levels of Abeta1-40, Abeta1-42, and Abeta40/42 ratio were stable. Statistical analysis revealed a strong positive correlation between proNGF and phospho-JNK, although only proNGF was negatively correlated with cognitive function and only TrkA was negatively associated with pathologic criteria. These findings suggest that alterations in the hippocampal NGF signaling pathway in MCI and AD favor proNGF-mediated proapoptotic pathways, and that this is independent of Abeta accumulation during AD progression.

Cysteine string protein alpha (CSPalpha), a presynaptic cochaperone for Hsc70, is required for synapse maintenance. Deletion of CSPalpha leads to neuronal dysfunction, synapse loss, and neurodegeneration. We utilized unbiased, systematic proteomics to identify putative CSPalpha protein clients. We found 22 such proteins whose levels are selectively decreased in CSPalpha knockout synapses. Of these putative CSPalpha protein clients, two directly bind to the CSPalpha chaperone complex and are bona fide clients. They are the t-SNARE SNAP-25 and the GTPase dynamin 1, which are necessary for synaptic vesicle fusion and fission, respectively. Using hippocampal cultures, we show that CSPalpha regulates the stability of client proteins and synaptic vesicle number. Our analysis of CSPalpha-dynamin 1 interactions reveals unexpectedly that CSPalpha regulates the polymerization of dynamin 1. CSPalpha, therefore, participates in synaptic vesicle endocytosis and may facilitate exo- and endocytic coupling. These findings advance the understanding of how synapses are functionally and structurally maintained.

OBJECTIVES/SPECIFIC AIMS: Th ere is no successful treatment or preventative measure for Alzheimer's disease (AD), a disorder characterized by altered amyloidbeta precursor (APP) processing, amyloid-beta (Abeta) aggregation, cell-type specific vulnerability, and cognitive deficits. Caloric restriction (CR) is a noninvasive dietary intervention that may prevent AD pathology. Previous studies have documented CRinduced AD pathology prevention through regional, low-resolution analysis. Th e effects of CR in selectively vulnerable hippocampal CA1 pyramidal neurons have not been demonstrated. METHODS/STUDY POPULATION: Th rough laser capture microdissection (LCM) and microarray analysis, we will investigate gene expression profiles of CA1 neurons isolated during pathology onset and later progression (5 and 15 mos.) from Tg2576 AD mice and nontransgenic littermates maintained on a 30% CR regimen versus ad libitum (AL) feeding. Abeta burden and APP metabolism are assessed as indicators of pathology and for correlation with high-throughput gene expression changes. RESULTS/ANTICIPATED RESULTS: We have identified CR-induced decreases in transcripts implicated in autophagy, synaptic transmission, transcription, and metabolism, as well as CR-specific increases in several kinase and phosphatase transcripts and qualitative reduction in Abeta peptides in CR-Tg2576 mice. We expect further characterization to support the hypothesis that CR induces increased neuronal efficiency, up-regulation of neuroprotective pathways, and decreased Abeta deposition in aged AL-Tg2576 mice. DISCUSSION/SIGNIFICANCE OF IMPACT: Alterations identified by these experiments may serve as targets in the development of therapeutic interventions. Additionally, this work may serve as the basis for further studies of CR-based dietary regimens as an AD pathology prevention option

Functional genomics and proteomics approaches are being employed to evaluate gene and encoded protein expression changes with the tacit goal to find novel targets for drug discovery. Genome-wide association studies (GWAS) have attempted to identify valid candidate genes through single nucleotide polymorphism (SNP) analysis. Furthermore, microarray analysis of gene expression in brain regions and discrete cell populations has enabled the simultaneous quantitative assessment of relevant genes. The ability to associate gene expression changes with neuropsychiatric disorders, including bipolar disorder (BP), and their response to therapeutic drugs provides a novel means for pharmacotherapeutic interventions. This review summarizes gene and pathway targets that have been identified in GWAS studies and expression profiling of human postmortem brain in BP, with an emphasis on glutamate receptors (GluRs). Although functional genomic assessment of BP is in its infancy, results to date point towards a dysregulation of GluRs that bear some similarity to schizophrenia (SZ), although the pattern is complex, and likely to be more complementary than overlapping. The importance of single population expression profiling of specific neurons and intrinsic circuits is emphasized, as this approach provides informative gene expression profile data that may be underappreciated in regional studies with admixed neuronal and non-neuronal cell types