Faculty Bibliography

Background: Biomarkers such as amyloid beta (e.g. Ab42) and hyperphosphorylated tau (e.g. pTau181) in cerebral spinal fluid (CSF) and hippocampal volume loss measured by magnetic resonance imaging (MRI) are useful for identifying cognitively normal elderly likely to have "preclinical" Alzheimer's disease (AD), but such methods are invasive and/or expensive. We investigated whether cognitive tasks dependent on brain regions affected in early AD can serve as proxies of AD biomarkers. Research indicates that the hippocampal formation (Hipp), particularly CA3/dentate gyrus (CA3/DG) and the entorhinal cortex (EC) are affected in preclinical AD. Therefore, we hypothesized that performance on a CA3/DG-dependent spatial pattern separation task (PST) and a Hipp/ EC-dependent discrimination and generalization task (DGT) would be impaired in cognitively normal individuals with biomarker evidence for AD. Methods: We collected initial data on our tasks from 31 cognitively normal NYU Alzheimer's Disease Center/Center for Brain Health participants who had MRI and who also provided CSF for longitudinal studies. In the PST, participants discriminated between two identical dots, one in a previously viewed location and one in a new location. In the DGT, participants learned to discriminate pairs of stimuli determined by shapes or colors in a discrimination phase, then had to generalize the "preferred" shapes and colors to novel stimuli in a generalization phase. Results: Linear regression analyses (with age and years of education as covariates) were used to determine whether task performance correlates with bilateral Hipp volume (used as a surrogate for CA3/DG and controlled for total intracranial volume) and CSF biomarkers. Performance on the PST correlates with bilateral Hipp volume (n = 31; R 2 = 0.151, P = 0.004) and CSF Ab42/pTau181 ratio (n = 26; R 2 = 0.182, P = 0.026). Performance on generalization correlates with Ab42 (R 2 = 0.182, P = 0.026) and marginally with Ab42/pTau181 ratio (R 2 = 0.119, P = 0.079). Performance on discrimination correlates with Ab42/ pTau181 ratio only (R 2 = 0.159, P = 0.039). A standard memory test (NYU Paragraph Recall) shows no significant correlations. Conclusions: These preliminary results are consistent with our hypothesis that cognitive tasks dependent on brain regions affected by early AD pathology may provide a non-invasive and cost-effective method to identify and track change in clinically normal individuals at high risk for progressing to theMCI and dementia stages of AD

Tauopathy in the brain of patients with Alzheimer's disease starts in the entorhinal cortex (EC) and spreads anatomically in a defined pattern. To test whether pathology initiating in the EC spreads through the brain along synaptically connected circuits, we have generated a transgenic mouse model that differentially expresses pathological human tau in the EC and we have examined the distribution of tau pathology at different timepoints. In relatively young mice (10-11 months old), human tau was present in some cell bodies, but it was mostly observed in axons within the superficial layers of the medial and lateral EC, and at the terminal zones of the perforant pathway. In old mice (>22 months old), intense human tau immunoreactivity was readily detected not only in neurons in the superficial layers of the EC, but also in the subiculum, a substantial number of hippocampal pyramidal neurons especially in CA1, and in dentate gyrus granule cells. Scattered immunoreactive neurons were also seen in the deeper layers of the EC and in perirhinal and secondary somatosensory cortex. Immunoreactivity with the conformation-specific tau antibody MC1 correlated with the accumulation of argyrophilic material seen in old, but not young mice. In old mice, axonal human tau immunoreactivity, especially at the endzones of the perforant pathway, was greatly reduced. Relocalization of tau from axons to somatodendritic compartments and propagation of tauopathy to regions outside of the EC correlated with mature tangle formation in neurons in the EC as revealed by thioflavin-S staining. Our data demonstrate propagation of pathology from the EC and support a trans-synaptic mechanism of spread along anatomically connected networks, between connected and vulnerable neurons. In general, the mouse recapitulates the tauopathy that defines the early stages of AD and provides a model for testing mechanisms and functional outcomes associated with disease progression.

Transgenic mice are used increasingly to model brain amyloidosis, mimicking the pathogenic processes involved in Alzheimer's disease (AD). In this chapter, an in vivo strategy is described that has been successfully used to map amyloid-beta deposits in transgenic mouse models of AD with magnetic resonance imaging (MRI), utilizing both the endogenous contrast induced by the plaques attributed to their iron content and by selectively enhancing the signal from amyloid-beta plaques using molecular-targeting vectors labeled with MRI contrast agents. To obtain sufficient spatial resolution for effective and sensitive mouse brain imaging, magnetic fields of 7-Tesla (T) or more are required. These are higher than the 1.5-T field strength routinely used for human brain imaging. The higher magnetic fields affect contrast agent efficiency and dictate the choice of pulse sequence parameters for in vivo MRI, all addressed in this chapter. Two-dimensional (2D) multi-slice and three-dimensional (3D) MRI acquisitions are described and their advantages and limitations are discussed. The experimental setup required for mouse brain imaging is explained in detail, including anesthesia, immobilization of the mouse's head to reduce motion artifacts, and anatomical landmarks to use for the slice alignment procedure to improve image co-registration during longitudinal studies and for subsequent matching of MRI with histology.

Reports an error in "Epac2-dependent mobilization of intracellular Ca2+ by glucagon-like peptide-1 receptor agonist exendin-4 is disrupted in beta -cells of phospholipase C-epsilon knockout mice" by Igor Dzhura, Oleg G. Chepurny, Grant G. Kelley, Colin A. Leech, Michael W. Roe, Elvira Dzhura, Parisa Afshari, Sundeep Malik, Michael J. Rindler, Xin Xu, Youming Lu, Alan V. Smrcka and George G. Holz (The Journal of Physiology, 2010[Dec][15], Vol 588[24], 4871-4889). In the original article, there was an error in the Methods section entitled 'Generation of Epac2 knockout mice' on page 4873. The first sentence of that section should read 'Epac2 KO mice with global disruption of RAPGEF4 gene expression (NCBI GeneID 56508) were generated by the Texas A&M Institute for Genomic Medicine through customized service for Dr. Lu at Louisiana State University Health Sciences Center'. (The following abstract of the original article appeared in record 2011-11969-007). Calcium can be mobilized in pancreatic beta -cells via a mechanism of Ca2+-induced Ca2+ release (CICR), and cAMP-elevating agents such as exendin-4 facilitate CICR in beta -cells by activating both protein kinase A and Epac2. Here we provide the first report that a novel phosphoinositide-specific phospholipase C-epsilon (PLC-epsilon ) is expressed in the islets of Langerhans, and that the knockout (KO) of PLC-epsilon gene expression in mice disrupts the action of exendin-4 to facilitate CICR in the beta -cells of these mice. Thus, in the present study, in which wild-type (WT) C57BL/6 mouse beta -cells were loaded with the photolabile Ca2+ chelator NP-EGTA, the UV flash photolysis-catalysed uncaging of Ca2+ generated CICR in only 9% of the beta -cells tested, whereas CICR was generated in 82% of the beta -cells pretreated with exendin-4. This action of exendin-4 to facilitate CICR was reproduced by cAMP analogues that activate protein kinase A(6-Bnz-cAMP-AM)orEpac2 (8-pCPT-2'-O-Me-cAMP-AM)selectively. However, in beta -cells of PLC-epsilon KO mice, and also Epac2 KO mice, these test substances exhibited differential efficacies in the CICR assay such that exendin-4 was partly effective, 6-Bnz-cAMP-AM was fully effective, and 8-pCPT-2'-O-Me-cAMP-AM was without significant effect. Importantly, transduction of PLC-epsilon KO beta -cells with recombinant PLC-epsilon rescued the action of 8-pCPT-2'-O-Me-cAMP-AM to facilitate CICR, whereas a K2150E PLC-epsilon with amutated Ras association (RA) domain, or a H1640L PLC-epsilon that is catalytically dead, were both ineffective. Since 8-pCPT-2'-O-Me-cAMP-AM failed to facilitate CICR in WT beta -cells transduced with a GTPase activating protein (RapGAP) that downregulates Rap activity, the available evidence indicates that a signal transduction 'module' comprised of Epac2, Rap and PLC-epsilon exists in beta -cells, and that the activities of Epac2 and PLC-epsilon are key determinants of CICR in this cell type.

Homeostatic scaling of synaptic strengths is essential for maintenance of network "gain", but also poses a risk of losing the distinctions among relative synaptic weights, which are possibly cellular correlates of memory storage. Multiplicative scaling of all synapses has been proposed as a mechanism that would preserve the relative weights among them, because they would all be proportionately adjusted. It is crucial for this hypothesis that all synapses be affected identically, but whether or not this actually occurs is difficult to determine directly. Mathematical tests for multiplicative synaptic scaling are presently carried out on distributions of miniature synaptic current amplitudes, but the accuracy of the test procedure has not been fully validated. We now show that the existence of an amplitude threshold for empirical detection of miniature synaptic currents limits the use of the most common method for detecting multiplicative changes. Our new method circumvents the problem by discarding the potentially distorting subthreshold values after computational scaling. This new method should be useful in assessing the underlying neurophysiological nature of a homeostatic synaptic scaling transformation, and therefore in evaluating its functional significance.

the molecular characterization of melanoma has ex- panded to include studies of microRNA (miRNA) ex- pression. As miR-16 has been utilized as a normalizer in serum-based miRNA studies in several cancers, we evaluated miR-16 expression as a potential reference for normalization of serum miRNA expression in melanoma patients. Methods: 143 primary cutaneous melanoma patients who presented to New York Uni- versity (NYU) Langone Medical Center for surgical resection of AJCC stage I-III disease were studied. In addition, sera samples from 60 control subjects were utilized including 22 healthy volunteers, 13 rheuma- toid arthritis patients, 20 non-melanoma cancer pa- tients (10 renal cell carcinoma and 10 bladder cancer), and 5 Atypical Mole Syndrome patients. The Kruskal- Wallis test (k = 6) or Wilcoxon test (k = 2) with Bon- ferroni correction was used for analyses of miR-16 expression in melanoma patients compared to various control groups, using raw Ct values directly. The Kruskal-Wallis test was used to compare miR-16 ex- pression across stages of melanoma. The equivalence test for independent samples was used to test the equivalence of miR-16 expression among different groups. Results: No significant differential expression of miR-16 was observed between melanoma patients and healthy volunteers (Wilcoxon test, p = 0.37). How- ever, miR-16 did show a significant difference in ex- pression as it related to stage of melanoma (p = 0.015). Additionally, the equivalence test was unable to con- firm equivalent expression of miR-16 in any melanoma versus control group pair. Conclusion: Our data in- dicate that miR-16 cannot be used as a universal normalizer in sera studies of melanoma patients

BACKGROUND: Mutations in any of the five subunits of eukaryotic translation initiation factor 2B (eIF2B) can lead to an inherited chronic-progressive fatal brain disease of unknown aetiology termed leucoencephalopathy with vanishing white matter (VWM). VWM is one of the most prevalent childhood white matter disorders, which markedly deteriorates after inflammation or exposure to other stressors. eIF2B is a major housekeeping complex that governs the rate of global protein synthesis under normal and stress conditions. A previous study demonstrated that Eif2b5(R132H/R132H) mice suffer delayed white matter development and fail to recover from cuprizone-induced demyelination, although eIF2B enzymatic activity in the mutant brain is reduced by merely 20%. PRINCIPAL FINDINGS: Poor astrogliosis was observed in Eif2b5(R132H/R132H) mice brain in response to systemic stress induced by peripheral injections of lipopolysaccharide (LPS). Even with normal rates of protein synthesis under normal conditions, primary astrocytes and microglia isolated from mutant brains fail to adequately synthesise and secrete cytokines in response to LPS treatment despite proper induction of cytokine mRNAs. CONCLUSIONS: The mild reduction in eIF2B activity prevents the appropriate increase in translation rates upon exposure to the inflammatory stressor LPS. The data underscore the importance of fully-functional translation machinery for efficient cerebral inflammatory response upon insults. It highlights the magnitude of proficient translation rates in restoration of brain homeostasis via microglia-astrocyte crosstalk. This study is the first to suggest the involvement of microglia in the pathology of VWM disease. Importantly, it rationalises the deterioration of clinical symptoms upon exposure of VWM patients to physiological stressors and provides possible explanation for their high phenotypic variability.

The increased availability of transgenic mouse models for studying human diseases has shifted the focus of many laboratories from in vitro to in vivo assays. Herein, methods are described to allow investigators to obtain well-preserved mouse tissue to be stained with the standard histological dyes for amyloid, Congo Red, and Thioflavin S. These sections can as well be used for immunohistological procedures that allow detection of tissue amyloid and pre-amyloid, such as those composed of the amyloid-beta peptide, the tau protein, and the islet amyloid polypeptide.