: It is becoming increasingly apparent that mitochondria play a critical role in a wide range of acute and chronic neurodegenerative diseases. Recent studies from this laboratory and many others have suggested that mitochondria may be the source of a signal that kills neurons. Perhaps the best candidate for this signal is mitochondrial generation of reactive oxygen species (ROS). The mechanisms of ROS generation by brain mitochondria, however, are poorly understood. Moreover, little is known about the regulation of the ROS signal. We propose to investigate the characteristics of ROS generation by brain mitochondria and cultured neurons.
In Specific Aim 1 we will determine the site(s) in the electron transport chain (ETC) that are the source of superoxide, and will determine the consequence of ETC inhibition on ROS generation in isolated brain mitochondria. Thus we will be able to model neurodegenerative disease based on prior reports of ETC inhibition in disorders such as Parkinson's disease and Alzheimer's disease.
In Specific Aim 2 we will investigate the mechanisms that regulate ROS generation and will focus on agents that alter the mitochondrial membrane potential and agents that uncouple oxygen consumption from ATP synthesis. These agents include calcium and substrates for uncoupling proteins.
In Specific Aim 3 we will use a series of neuron preparations to establish the properties of ROS generation by mitochondria in their intact cellular environment. We will use standard neuronal cultures, permeabilized neurons and acutely dissociated cells, along with fluorescence imaging of ROS, to establish neuron-specific mechanisms of mitochondrial ROS generation. In the final Specific Aim we will investigate whether the retrograde transport of mitochondria by neurons in vivo is altered by oxidative injury. This novel approach will allow us to investigate the properties of cellular mitochondrial homeostasis which will provide a long-term view of the role of mitochondria in neurodegeneration. These studies will illuminate a critical role of mitochondria and ROS signaling in neuronal injury, and will provide novel targets for intervention in a wide range of neurodegenerative disease.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Project (R01)
Project #
5R01AG020899-05
Application #
6944710
Study Section
Special Emphasis Panel (ZNS1-SRB-S (01))
Program Officer
Wise, Bradley C
Project Start
2001-09-30
Project End
2008-08-31
Budget Start
2005-09-15
Budget End
2008-08-31
Support Year
5
Fiscal Year
2005
Total Cost
$350,942
Indirect Cost
Name
University of Pittsburgh
Department
Pharmacology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Selivanov, Vitaly A; Votyakova, Tatyana V; Pivtoraiko, Violetta N et al. (2011) Reactive oxygen species production by forward and reverse electron fluxes in the mitochondrial respiratory chain. PLoS Comput Biol 7:e1001115
Devinney, Michael J; Malaiyandi, Latha M; Vergun, Olga et al. (2009) A comparison of Zn2+- and Ca2+-triggered depolarization of liver mitochondria reveals no evidence of Zn2+-induced permeability transition. Cell Calcium 45:447-55
Selivanov, Vitaly A; Votyakova, Tatyana V; Zeak, Jennifer A et al. (2009) Bistability of mitochondrial respiration underlies paradoxical reactive oxygen species generation induced by anoxia. PLoS Comput Biol 5:e1000619
Zhao, Jinfu; Bertoglio, Bryan A; Devinney Jr, Michael J et al. (2009) The interaction of biological and noxious transition metals with the zinc probes FluoZin-3 and Newport Green. Anal Biochem 384:34-41
Van Laar, Victor S; Mishizen, Amanda J; Cascio, Michael et al. (2009) Proteomic identification of dopamine-conjugated proteins from isolated rat brain mitochondria and SH-SY5Y cells. Neurobiol Dis 34:487-500
Dukes, April A; Van Laar, Victor S; Cascio, Michael et al. (2008) Changes in endoplasmic reticulum stress proteins and aldolase A in cells exposed to dopamine. J Neurochem 106:333-46
Van Laar, Victor S; Dukes, April A; Cascio, Michael et al. (2008) Proteomic analysis of rat brain mitochondria following exposure to dopamine quinone: implications for Parkinson disease. Neurobiol Dis 29:477-89
Selivanov, Vitaly A; Zeak, Jennifer A; Roca, Josep et al. (2008) The role of external and matrix pH in mitochondrial reactive oxygen species generation. J Biol Chem 283:29292-300
Dineley, Kirk E; Devinney 2nd, Michael J; Zeak, Jennifer A et al. (2008) Glutamate mobilizes [Zn2+] through Ca2+ -dependent reactive oxygen species accumulation. J Neurochem 106:2184-93
Gusdon, Aaron M; Votyakova, Tatyana V; Reynolds, Ian J et al. (2007) Nuclear and mitochondrial interaction involving mt-Nd2 leads to increased mitochondrial reactive oxygen species production. J Biol Chem 282:5171-9

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