I have demonstrated that neurotrophins (NTs) protect cultured embryonic rat basal forebrain cholinergic neurons during hypoglycemic stress. The goal of the proposed study is to identify important second messenger and early effector pathways involved in this protection. Many of the experiments will utilize cholinergic neurons purified by immunopanning. I have preliminary evidence that the following contribute importantly to NT-mediated protection of cholinergic neurons during hypoglycemic stress: (1) activation of phosphotidyl inositol (PI) 3-kinase, and (2) activation of NF-kB (perhaps) via the p75 NT receptor (p75NTR). Receptor-blocking antibodies, neurons from receptor-deficient mice, and mutant forms of nerve growth factor (NGF) will be used to test the involvement of the p75 NTR and TrkA NGF receptors. Assays of kinase activity and immunohistochemical staining for activated forms of signalling molecules will be used to clarify the stress-protective pathways activated by NTs under hypoglycemic conditions. Inhibitors and activators of specific kinases and introduction (by transfection) of mutant kinases will be tested to determine whether they block, mimic or occlude NT-mediated protection. To investigate early effector pathways by which NTs might produce their stress-protection, imaging techniques will be used to measure NT effects on cytosolic [Na+] and [Ca2+], mitochondrial [Ca2+] and mitochondrial membrane potential. Alternative energy substrates, respiration measurements and inhibitors of specific mitochondrial complexes will be tested to determine whether NT-mediated protection requires up-regulation of plasma membrane glucose transporters and/or alterations in mitochondrial energy and/or Ca2+ metabolism. I have recently discovered that certain bone morphogenetic proteins (BMPs) robustly enhance NT-mediated protection during hypoglycemic stress, and the proposed experimental plan integrates studies to uncover basic mechanisms underlying this synergistic effect.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS012207-25
Application #
6639351
Study Section
Special Emphasis Panel (ZRG1-MDCN-2 (01))
Program Officer
Oliver, Eugene J
Project Start
1986-07-01
Project End
2005-03-31
Budget Start
2003-07-01
Budget End
2005-03-31
Support Year
25
Fiscal Year
2003
Total Cost
$337,500
Indirect Cost
Name
University of Miami School of Medicine
Department
Physiology
Type
Schools of Medicine
DUNS #
052780918
City
Miami
State
FL
Country
United States
Zip Code
33146
White, Michael G; Saleh, Osama; Nonner, Doris et al. (2012) Mitochondrial dysfunction induced by heat stress in cultured rat CNS neurons. J Neurophysiol 108:2203-14
Chaverneff, Florence; Barrett, John (2009) Casein kinase II contributes to the synergistic effects of BMP7 and BDNF on Smad 1/5/8 phosphorylation in septal neurons under hypoglycemic stress. J Neurochem 109:733-43
Panickar, Kiran S; Nonner, Doris; White, Michael G et al. (2008) Overexpression of Cdk5 or non-phosphorylatable retinoblastoma protein protects septal neurons from oxygen-glucose deprivation. Neurochem Res 33:1852-8
Talbot, Janet D; Barrett, John N; Barrett, Ellen F et al. (2008) Rapid, stimulation-induced reduction of C12-resorufin in motor nerve terminals: linkage to mitochondrial metabolism. J Neurochem 105:807-19
White, Michael G; Luca, Luminita E; Nonner, Doris et al. (2007) Cellular mechanisms of neuronal damage from hyperthermia. Prog Brain Res 162:347-71
Talbot, Janet; Barrett, John N; Barrett, Ellen F et al. (2007) Stimulation-induced changes in NADH fluorescence and mitochondrial membrane potential in lizard motor nerve terminals. J Physiol 579:783-98
Panickar, K S; Nonner, D; Barrett, J N (2005) Overexpression of Bcl-xl protects septal neurons from prolonged hypoglycemia and from acute ischemia-like stress. Neuroscience 135:73-80
White, Michael G; Emery, Michael; Nonner, Doris et al. (2003) Caspase activation contributes to delayed death of heat-stressed striatal neurons. J Neurochem 87:958-68
Vila, Lizette; Barrett, Ellen F; Barrett, John N (2003) Stimulation-induced mitochondrial [Ca2+] elevations in mouse motor terminals: comparison of wild-type with SOD1-G93A. J Physiol 549:719-28
Nonner, D; Barrett, E F; Barrett, J N (2000) Brief exposure to neurotrophins produces a calcium-dependent increase in choline acetyltransferase activity in cultured rat septal neurons. J Neurochem 74:988-99

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