Core - Brain Microdissection, RNA and Protein Analysis

Hyman, Bradley T.

Abstract

This core will provide a central resource for reagent synthesis, tissue preparation, and quantitative RNA and protein analysis for all 5 projects. DNA oligonucleotides will be synthesized using an Applied Biosciences 392 DNA synthesizer and purified by gel electrophoresis. These will be used for in situ hybridization protocols in projects 1,2, and 4. RNA and protein will be isolated from tissue samples used in each of the projects (human brain, experimental animals, or tissue culture material). Brains will be microdissected to allow analysis of discrete anatomical regions. RNA and protein separation will be carried out by RNAse protection assays and Western blots. Quantification will be by autoradiography and densitometry using an LKB Ultrascan Laser Densitometer. Providing these quantitative assays as a core service will enhance each project, and be cost effective compared to duplicate analytical setups in each laboratory. Particularly with regard to the human material, the core will allow for optimal use of scarce resources. Furthermore, in the course of these studies, the core will generate RNA and protein samples from carefully microdissected brain regions of clinically well characterized patients and young and old experimental animals. This will be an invaluable resource and will be of great use as these studies of aging and Alzheimer disease evolve.

Funding Agency

• Agency: National Institute of Health (NIH)
• Institute: National Institute on Aging (NIA)
• Type: Research Program Projects (P01)
• Project #: 5P01AG011337-03
• Application #: 5204893
• Support Year: 3
• Fiscal Year: 1996

Publications

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Liu, Qing; Waltz, Shannon; Woodruff, Grace et al. (2014) Effect of potent ?-secretase modulator in human neurons derived from multiple presenilin 1-induced pluripotent stem cell mutant carriers. JAMA Neurol 71:1481-9

Young, Anne B (2009) Four decades of neurodegenerative disease research: how far we have come! J Neurosci 29:12722-8

Swerdlow, N R; Young, A B (2001) Neuropathology in Tourette syndrome: an update. Adv Neurol 85:151-61

Cha, J H; Farrell, L A; Ahmed, S F et al. (2001) Glutamate receptor dysregulation in the hippocampus of transgenic mice carrying mutated human amyloid precursor protein. Neurobiol Dis 8:90-102

Montine, T J; Shinobu, L; Montine, K S et al. (2000) No difference in plasma or urinary F2-isoprostanes among patients with Huntington's disease or Alzheimer's disease and controls. Ann Neurol 48:950

Chin, J Y; Knowles, R B; Schneider, A et al. (2000) Microtubule-affinity regulating kinase (MARK) is tightly associated with neurofibrillary tangles in Alzheimer brain: a fluorescence resonance energy transfer study. J Neuropathol Exp Neurol 59:966-71

Standaert, D G; Friberg, I K; Landwehrmeyer, G B et al. (1999) Expression of NMDA glutamate receptor subunit mRNAs in neurochemically identified projection and interneurons in the striatum of the rat. Brain Res Mol Brain Res 64:11-23

Knowles, R B; Chin, J; Ruff, C T et al. (1999) Demonstration by fluorescence resonance energy transfer of a close association between activated MAP kinase and neurofibrillary tangles: implications for MAP kinase activation in Alzheimer disease. J Neuropathol Exp Neurol 58:1090-8

Montine, T J; Beal, M F; Robertson, D et al. (1999) Cerebrospinal fluid F2-isoprostanes are elevated in Huntington's disease. Neurology 52:1104-5