Faculty Bibliography

Cholinergic basal forebrain (CBF) nucleus basalis (NB) neurons display neurofibrillary tangles (NFTs) during Alzheimer's disease (AD) progression, yet the mechanisms underlying this selective vulnerability are currently unclear. Rac1, a member of the Rho family of GTPases, may interact with the proapoptotic pan-neurotrophin receptor p75(NTR) to induce neuronal cytoskeletal abnormalities in AD NB neurons. Herein, we examined the expression of Rac1b, a constitutively active splice variant of Rac1, in NB cholinergic neurons during AD progression. CBF tissues harvested from people who died with a clinical diagnosis of no cognitive impairment (NCI), mild cognitive impairment, or AD were immunolabeled for both p75(NTR) and Rac1b. Rac1b appeared as cytoplasmic diffuse granules, loosely aggregated filaments, or compact spheres in p75(NTR)-positive NB neurons. Although Rac1b colocalized with tau cytoskeletal markers, the percentage of p75(NTR)-immunoreactive neurons expressing Rac1b was significantly increased only in AD compared with both mild cognitive impairment and NCI. Furthermore, single-cell gene expression profiling with custom-designed microarrays showed down-regulation of caveolin 2, GNB4, and lipase A in AD Rac1b-positive/p75(NTR)-labeled NB neurons compared with Rac1b-negative/p75(NTR)-positive perikarya in NCI. These proteins are involved in Rac1 pathway/cell cycle progression and lipid metabolism. These data suggest that Rac1b expression acts as a modulator or transducer of various signaling pathways that lead to NFT formation and membrane dysfunction in a subgroup of CBF NB neurons in AD.

The hTau mouse model of tauopathy was utilized to assess gene expression changes in vulnerable hippocampal CA1 neurons. CA1 pyramidal neurons were microaspirated via laser capture microdissection followed by RNA amplification in combination with custom-designed microarray analysis and qPCR validation in hTau mice and nontransgenic (ntg) littermates aged 11-14months. Statistical analysis revealed ~8% of all the genes on the array platform were dysregulated, with notable downregulation of several synaptic-related markers including synaptophysin (Syp), synaptojanin, and synaptobrevin, among others. Downregulation was also observed for select glutamate receptors (GluRs), Psd-95, TrkB, and several protein phosphatase subunits. In contrast, upregulation of tau isoforms and a calpain subunit were found. Microarray assessment of synaptic-related markers in a separate cohort of hTau mice at 7-8months of age indicated only a few alterations compared to the 11-14month cohort, suggesting progressive synaptic dysfunction occurs as tau accumulates in CA1 pyramidal neurons. An assessment of SYP and PSD-95 expression was performed in the hippocampal CA1 sector of hTau and ntg mice via confocal laser scanning microscopy along with hippocampal immunoblot analysis for protein-based validation of selected microarray observations. Results indicate significant decreases in SYP-immunoreactive and PSD-95-immunoreactive puncta as well as downregulation of SYP-immunoreactive and PSD-95-immunoreactive band intensity in hTau mice compared to age-matched ntg littermates. In summary, the high prevalence of downregulation of synaptic-related genes indicates that the moderately aged hTau mouse may be a model of tau-induced synaptodegeneration, and has profound effects on how we perceive progressive tau pathology affecting synaptic transmission in AD

To evaluate molecular signatures of an individual cell type in comparison to the associated region relevant towards understanding the pathogenesis of Alzheimer's disease (AD), CA1 pyramidal neurons and the surrounding hippocampal formation were microaspirated via laser capture microdissection (LCM) from neuropathologically confirmed AD and age-matched control (CTR) subjects as well as from wild type mouse brain using single population RNA amplification methodology coupled with custom-designed microarray analysis with real-time quantitative polymerase-chain reaction (qPCR) validation. CA1 pyramidal neurons predominantly displayed downregulation of classes of transcripts related to synaptic transmission in AD versus CTR. Regional hippocampal dissections displayed downregulation of several overlapping genes found in the CA1 neuronal population related to neuronal expression, as well as upregulation of select transcripts indicative of admixed cell types including glial-associated markers and immediate-early and cell death genes. Gene level distributions observed in CA1 neurons and regional hippocampal dissections in wild type mice paralleled expression mosaics seen in postmortem human tissue. Microarray analysis was validated in qPCR studies using human postmortem brain tissue and CA1 sector and regional hippocampal dissections obtained from a mouse model of AD/Down syndrome (Ts65Dn mice) and normal disomic (2N) littermates. Classes of transcripts that have a greater percentage of the overall hybridization signal intensity within single neurons tended to be genes related to neuronal communication. The converse was also found, as classes of transcripts such as glial-associated markers were under represented in CA1 pyramidal neuron expression profiles relative to regional hippocampal dissections. These observations highlight a dilution effect that is likely to occur in conventional regional microarray and qPCR studies. Thus, single population studies of specific neurons and intrinsic circuits will likely yield informative gene expression profile data that may be subthreshold and/or underrepresented in regional studies with an admixture of cell types

Mild cognitive impairment (MCI) is rapidly becoming one of the most common clinical manifestations affecting the elderly. The pathologic and molecular substrate of people diagnosed with MCI is not well established. Since MCI is a human specific disorder and neither the clinical nor the neuropathological course appears to follow a direct linear path, it is imperative to characterize neuropathology changes in the brains of people who came to autopsy with a well-characterized clinical diagnosis of MCI. Herein, we discuss findings derived from clinical pathologic studies of autopsy cases who died with a clinical diagnosis of MCI. The heterogeneity of clinical MCI imparts significant challenges to any review of this subject. The pathologic substrate of MCI is equally complex and must take into account not only conventional plaque and tangle pathology but also a wide range of cellular, biochemical and molecular deficits, many of which relate to cognitive decline as well as compensatory responses to the progressive disease process. The multifaceted nature of the neuronal disconnection syndrome associated with MCI suggests that there is no single event which precipitates this prodromal stage of AD. In fact, it can be argued that neuronal degeneration initiated at different levels of the central nervous system drives cognitive decline as a final common pathway at this stage of the dementing disease process.

Background: Neurotrophin signaling of cholinergic basal forebrain (CBF) neurons is critical for survival and plasticity. Microaspiration of identified CBF neurons from postmortem human brain revealed a shift in balance of neurotrophin receptors toward cell death pathways during the progression of Alzheimer's disease (AD). Methods: In this study transcriptomic data from mouse basal forebrain cholinergic neurons (BFCNs; NCBI GEO GSE13379) were used to derive parameters for a deterministic kinetic model of the nerve growth factor (NGF) signaling pathway from Reactome, with TrkB receptor mechanisms added. This method is called Transcriptome-To-Reactome (TTR)-. The biosimulation was performed using COPASI software and included 11 compartments 435 species, and 263 reactions; 245 genes were used to determine initial values of species and kinetic values of reactions. The mouse BFCN model was considered baseline and a biosimulation was run with two doses of NGF, 500 m M and 10 mM, delivered as a bolus and for a 10 and 240 second duration, respectively. This approach tested selectively for p75 NTR and TrkA receptor mediated mechanisms. A second biosimulation test used a combination of 25 mM brain derived neurotrophic factor (BDNF) and 10 m M NGF as a continuous exposure for 60 min duration; this approach evaluated stimulation of p75 NTR TrkA, and TrkB. Based on the human microarray results demonstrating downregulation of TrkA (50%) and TrkB (60%), the corresponding parameters in the TTR biosimulation were decreased by the same amount. Results: Baseline results were validated from published literature on neuronal calcium levels mediated via the phospholipase C-g and inositol- 3-phosphate pathway at both bolus doses of NGF alone. With the corresponding parameters decreased in the TTR biosimulation, Figure 1: A) The reaction flux for c-RAF1 phosphorylation of MEK1 was delayed to peak value by 1.5 min from exposure, but the peak value was increased to 5 times the baseline value; B) Moreover, a slight shift t!

Background: Reductions in brain-derived neurotrophic factor (BDNF) have been implicated in the pathophysiology of Alzheimer's disease (AD). Nevertheless, the factors influencing central and peripheral BDNF levels are still poorly understood. Cerebral microvascular endothelial cells are known to be a major source of BDNF with a rate of production by far exceeding that of cortical neurons. Exposure of these cells to amyloid beta (Ab), results in cell death or injury with significant reductions in BDNF secretion. Moreover, in rodents, infusion of Ab40 into the carotid resulted in a disruption of endothelial cells, which was not observed with Ab42. Plasma Ab40 levels have also been associated with white matter hyperintense lesions (WMHI) on MRI scans in AD, an effect that may be mediated by the toxic effects of soluble Ab40 on small cerebral blood vessels and endothelial cells. Therefore, we hypothesized that concentrations of plasma Ab40, but not Ab42, would have a negative effect on plasma BDNF and on measures of white matter integrity as determined by Diffusion Tensor Imaging (DTI). Methods: To test this hypothesis, we examined BDNF and Ab levels in plasma from 119 subjects with intact cognition (no dementia and a Mini-Mental State Exam score of at least 28) and no gross MRI abnormalities other than white matter hyperintensities. Of these, 88 subjects also had BDNF in plasma determined. Results: Consistent with our prediction, Ab40 was inversely correlated with BDNF concentrations (P <.001), whereas Ab42 was independent (P = .231). Fractional anisotropy (FA; a measure of white matter integrity in DTI) was also inversely correlated with Ab40 (P = .001) and so was performance in delayed recall (P = .029). Conclusions: In cognitively intact individuals, circulating Ab40 results in reduction in plasma BDNF, white matter integrity (FA), and memory performance. As such, it may have prognostic significance