Assessment of Newly Synthesized Mitochondrial DNA Using BrdU Labeling in Primary Neurons from Alzheimer's Disease Mice: Implications for Impaired Mitochondrial Biogenesis and Synaptic Damage

Marcus J. Calkins (1) , P. Hemachandra Reddy (1, 2)

1 Division of Neuroscience
2 Department of Physiology and Pharmacology

The purpose our study was to assess mitochondrial biogenesis and distribution in murine primary neurons. Using 5-bromo-2-deoxyuridie (BrdU) incorporation and primary neurons, we studied the mitochondrial biogenesis and mitochondrial distribution in hippocampal neurons from amyloid beta precursor protein (AβPP) transgenic mice and wild-type (WT) neurons treated with oxidative stressors, rotenone and HO. We found that after 20hr of labeling, BrdU incorporation was specific to porin-positive mitochondria. The proportion of mitochondrial area that labeled with BrdU was 40.3±6.3% at 20hr. The number of mitochondria with newly synthesized DNA was higher in AβPP neuronal cell bodies than in the cell bodies of WT neurons (AβPP, 45.23±2.67 BrdU-positive/cell body; WT, 32.92±2.49 BrdU-positive/cell body; p=0.005). In neurites, the number of BrdU-positive mitochondria decreased in AβPP cultures compared to WT neurons (AβPP, 0.105±0.008 BrdU-positive/μm neurite; WT, 0.220±0.036 BrdU-positive−/−μm neurite; p=0.010). Further, BrdU in the cell body increased when neurons were treated with low doses of HO (49.6±2.7 BrdU-positive/cell body, p=0.0002 compared to untreated cells), while the neurites showed decreased BrdU staining (0.122±0.010 BrdU-positive/μm neurite, p=0.005 compared to the untreated). BrdU labeling was increased in the cell body under rotenone treatment. Additionally, under rotenone treatment, the content of BrdU labeling decreased in neurites. These findings suggest that Aβ and mitochondrial toxins enhance mitochondrial fragmentation in cell body, and may cause impaired axonal transport of mitochondria leading to synaptic degeneration.

Mots clés: AβPP; AD; ATP; BrdU; Drp1; Fis1; Primary neurons; Alzheimer's disease; Mitochondrial DNA synthesis; 5-bromo-2-deoxyuridine; Cell body mitochondria; Mitochondrial toxins; Mitochondria-targeted antioxidant

Domaines: Biochimie, Biologie Moléculaire

Marcus J. Calkins, P. Hemachandra Reddy. Assessment of Newly Synthesized Mitochondrial DNA Using BrdU Labeling in Primary Neurons from Alzheimer's Disease Mice: Implications for Impaired Mitochondrial Biogenesis and Synaptic Damage. Biochimica et Biophysica Acta - Molecular Basis of Disease, 2011, 1812 (9), pp.1182. ⟨10.1016/j.bbadis.2011.04.006⟩. ⟨hal-00719833⟩