Technical Reference #1621

1621.

Examination of Potential Mechanisms of Amyloid-induced Defects in Neuronal Transport Sameer B. Shah a; Rhiannon Nolan b; Emily Davis b; Gorazd B. Stokin c; Ingrid Niesman b; Isabel Canto d; 4 Charles Glabe d; Lawrence S.B. Goldstein b;e;⁎, University of Maryland, Neurobiology of Disease, 36(1621), (2009)
Link To Paper

Abstract:
Microtubule-based neuronal transport pathways are impaired during the progression of Alzheimer's disease 27and other neurodegenerative conditions. However mechanisms leading to defects in transport remain to be 28determined. We quantified morphologic

Keywords:
Alzheimer's disease; Amyloid; Axonal transport; Oxidative stress; Aggregation; Morphology

Materials & Methods:
Cell culture 126 We grew Neuro-2a (N2a) mouse neuroblastoma cells (ATCC) at 127 37 °C/5% CO2 and passaged them in DMEM with L-glutamine (Gibco 128 11965) containing 10% fetal bovine serum and penicillin/streptomycin 129 (Gibco 15140). Prior to imaging we plated cells on poly-D-lysine 130 coated glass coverslips (Carolina Supply Company 633029) Therma- 131 nox coverslips (EMS 72280) or poly-D-lysine coated live-imaging 132 dishes (Mattek P35G-020-C). To induce differentiation cells were 133 incubated overnight in Optimem (Gibco 31985) with 1 mM di- 134 butyryl-cAMP. We harvested hippocampi from P1 C57/Bl6 mouse 135 neonates per standard protocols and plated triturated cells at a 136 density of ~1×106 cells/ml on poly-D-lysine coated glass coverslips in 137 DMEM containing 10% fetal bovine serum. After 3 h we changed the 138 media to Neurobasal media (Gibco 12348) containing B27 supple- 139 ment (Gibco 17504) and 1 mM L-glutamine. After five days we 140 replaced this media with Neurobasal containing B27 supplement. All 141 animal work was performed in accordance with the approval of UCSD 142 IACUC.

Microscopic Technique
Fluorescence Micrscopy

Cell Type(s)
N2A