Using 31P NMR spectroscopy at 14.1 Tesla to investigate PARP-1 associated energy failure and metabolic rescue in cerebrocortical slices.
PARP-1 activation by H(2)O(2) in an acute preparation of superfused, respiring, neonatal cerebrocortical slices was assessed from PAR-polymer formation detected with immunohistochemistry and Western blotting. (31)P NMR spectroscopy at 14.1 Tesla of perchloric acid slice extracts was used to assess energy failure in a 1-h H(2)O(2) exposure as well as in a subsequent 4-h recovery period where the superfusate had no H(2)O(2) and specifically chosen metabolic substrates. Although more data are needed to fully characterize different bioenergetic responses, a high NMR spectral resolution (PCr full-width at half-max approximately.01 ppm) and narrow widths for most metabolites (<.2 ppm) permitted accurate quantifications of spectrally resolved resonances for ADP, ATP, NAD(+)/NADH, and other high energy phosphates. It appears possible to use brain slices to quantitatively study PARP-related, NAD-associated energy failure, and rescue with TCA metabolites.
Zeng
Jianying
J
Department of Anesthesia, University of California at San Francisco, San Francisco, California 94143-0648, USA.
Hirai
Kiyoshi
K
Yang
Guo-Yuan
GY
Ying
Weihai
W
Swanson
Raymond A
RA
Kelly
Mark
M
Mayer
Moriz
M
James
Thomas L
TL
Litt
Lawrence
L
D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology; D004734:Energy Metabolism / Q000187:drug effects / Q000502:physiology; D006861:Hydrogen Peroxide / Q000008:administration & dosage; D008279:Magnetic Resonance Imaging / Q000379:methods*; D008280:Magnetics; D009243:NAD / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D010759:Phosphorus Isotopes; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D046509:Tissue Culture Techniques / Q000379:methods
D010759:Phosphorus Isotopes; D009243:NAD; D006861:Hydrogen Peroxide; C499321:Parp1 protein, mouse; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases