This is a revised application for continued support to study mitochondrial contributions to Ca2+ regulation in wild type and mutant SOD1G93A mouse motor nerve terminals. Changes in cytosolic and intramitochondrial [Ca2+] evoked by repetitive stimulation will be measured by imaging the fluorescence of indicator dyes. Phasic (end-plate potentials) and asynchronous quantal transmitter release will be recorded electrophysiologically, and vesicular recycling measured using styryl dyes. We have shown that mitochondrial uptake of Ca2+ is critically important for sustaining neuromuscular transmission during high-frequency stimulation. Proposed experiments will investigate the mechanisms by which motor terminal mitochondria take up, store and extrude this Ca2+ load. Using a novel permeabilized motor terminal preparation we will test how certain cytosolic components affect the affinity of mitochondrial Ca2+ uptake. We will test the role of inorganic phosphate in intramitochondrial Ca+ buffering, and investigate the linkage between mitochondrial Ca2+ extrusion and cytosolic [Na+]. The impact of Ca2+ uptake on mitochondrial energy metabolism will be investigated using measurements of stimulation-induced changes in mitochondrial membrane potential and changes in NADH and FAD autofluorescence. Another series of experiments will investigate the linkage between abnormal mitochondrial [Ca2+] handling and abnormal transmitter release in a mouse model of familial amyotrophic lateral sclerosis (ALS, SOD1G93A). Embryonic SOD1G93A motoneurons are especially vulnerable to nitric oxide (NO)-induced death. NO is produced in active muscle, so we will investigate whether the function of SOD1G93A motor terminals is more susceptible to NO-induced disruption than that of wild-type terminals, to test the hypothesis that NO contributes to functional deficits and degeneration of motor terminals in these mice. The proposed studies will thus probe fundamental mechanisms concerning how motor terminal mitochondria handle physiological Ca2+ loads, and investigate how Ca2+ dysregulation and NO contribute to motor terminal dysfunction in SOD1G93A motor terminals.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
Project #
Application #
Study Section
Molecular, Cellular and Developmental Neurosciences 2 (MDCN)
Program Officer
Talley, Edmund M
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of Miami School of Medicine
Schools of Medicine
United States
Zip Code
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
Nguyen, Khanh T; Barrett, John N; García-Chacón, Luis et al. (2011) Repetitive nerve stimulation transiently opens the mitochondrial permeability transition pore in motor nerve terminals of symptomatic mutant SOD1 mice. Neurobiol Dis 42:381-90
Nguyen, Khanh T; García-Chacón, Luis E; Barrett, John N et al. (2009) The Psi(m) depolarization that accompanies mitochondrial Ca2+ uptake is greater in mutant SOD1 than in wild-type mouse motor terminals. Proc Natl Acad Sci U S A 106:2007-11
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
David, Gavriel; Nguyen, Khanh; Barrett, Ellen F (2007) Early vulnerability to ischemia/reperfusion injury in motor terminals innervating fast muscles of SOD1-G93A mice. Exp Neurol 204:411-20
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
Garcia-Chacon, Luis E; Nguyen, Khanh T; David, Gavriel et al. (2006) Extrusion of Ca2+ from mouse motor terminal mitochondria via a Na+-Ca2+ exchanger increases post-tetanic evoked release. J Physiol 574:663-75
Talbot, Janet D; David, Gavriel; Barrett, Ellen F (2003) Inhibition of mitochondrial Ca2+ uptake affects phasic release from motor terminals differently depending on external [Ca2+]. J Neurophysiol 90:491-502
David, Gavriel; Talbot, Janet; Barrett, Ellen F (2003) Quantitative estimate of mitochondrial [Ca2+] in stimulated motor nerve terminals. Cell Calcium 33:197-206

Showing the most recent 10 out of 25 publications