Stroke is a major cause of morbidity and mortality in the United States and presents challenges in the development of disease prevention strategies. Deprivation of nutrients and oxygen supply to brain cells produces immediate death in the most severely affected cells and delayed cell death in some cells that are placed at risk at the first onset of ischemia. The latter form of neuronal death is orchestrated by BCL-2 family proteins, which also regulate cell death during nervous system development. BCL-2 family proteins are present in mitochondrial membranes and become activated to form large conductance ion channel activity in response to many death stimuli, including a low concentration of cellular oxygen. The relationship of the channel activity to the onset of cell death is as yet poorly understood. We have developed patch clamp techniques to record mitochondrial ion channel activity within living neurons and to study the ion channel activity of BCL-2 family proteins in mitochondria isolated from brain. In comparing the channel activity of recombinant BCL-2 family proteins to endogenous activity, we find that the recombinant N-truncated form of BCL-xL, (?N BCL-xL) that is produced by acute proteolytic processing of full length BCL-xL in response to death stimuli, has biophysical and pharmacological similarities to an endogenous mitochondrial channel that appears during transient global ischemia in mammalian brain. The onset of channel activity of ?N BCL-xL may initiate a series of molecular events that leads to selective, delayed cell death in vulnerable CA1 neurons of the hippocampus. We hypothesize that inhibition of the ?N BCL-xL channel activity with the specific inhibitor of BCL-xL, ABT-737, will block cell death in these sensitive neurons after ischemia. Furthermore, if proteolytic processing of full length BCL-xL is necessary for cell death, then hippocampal neurons of a mouse that lacks a cleavable form of BCL-xL may fail to form ?N BCL-xL and fail to die after ischemic insult. These studies will test the hypothesis that ?N BCL-xL is the key regulator of cell death in hippocampal neurons. In addition, these studies will attempt to elucidate the molecular mechanisms underlying cell death in neurons after ischemic brain injury with an eye to developing strategies to combat stroke.

Public Health Relevance

Stroke is a major cause of morbidity and mortality in the United States. We have found that mitochondrial ion channel activity of the pro- and anti-death BCL-2 protein family contributes to the physiological and pathological function of neurons. We hypothesize that in the setting of brain ischemia, such channel activity leads to cell death in the brain and that this death can be prevented by a pharmacological inhibitor of BCL-2-induced mitochondrial ion channels.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Neural Oxidative Metabolism and Death Study Section (NOMD)
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Bosetti, Francesca
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Yale University
Internal Medicine/Medicine
Schools of Medicine
New Haven
United States
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Mnatsakanyan, Nelli; Beutner, Gisela; Porter, George A et al. (2017) Physiological roles of the mitochondrial permeability transition pore. J Bioenerg Biomembr 49:13-25
Alavian, Kambiz N; Dworetzky, Steven I; Bonanni, Laura et al. (2015) The mitochondrial complex V-associated large-conductance inner membrane current is regulated by cyclosporine and dexpramipexole. Mol Pharmacol 87:1-8
Jonas, Elizabeth A; Porter Jr, George A; Beutner, Gisela et al. (2015) Cell death disguised: The mitochondrial permeability transition pore as the c-subunit of the F(1)F(O) ATP synthase. Pharmacol Res 99:382-92
Jonas, Elizabeth A (2014) Contributions of Bcl-xL to acute and long term changes in bioenergetics during neuronal plasticity. Biochim Biophys Acta 1842:1168-78
Li, Hongmei; Park, Han-A; Jonas, Elizabeth A (2014) Fluorescent Measurement of Synaptic Activity Using SynaptopHluorin in Isolated Hippocampal Neurons. Bio Protoc 4:
Li, Hongmei; Alavian, Kambiz N; Lazrove, Emma et al. (2013) A Bcl-xL-Drp1 complex regulates synaptic vesicle membrane dynamics during endocytosis. Nat Cell Biol 15:773-85
Sacchetti, Silvio; Alavian, Kambiz N; Lazrove, Emma et al. (2013) F1FO ATPase vesicle preparation and technique for performing patch clamp recordings of submitochondrial vesicle membranes. J Vis Exp :e4394
Pontarelli, Fabrizio; Ofengeim, Dimitry; Zukin, R Suzanne et al. (2012) Mouse Transient Global Ischemia Two-Vessel Occlusion Model. Bio Protoc 2:
Gray, Joshua P; Alavian, Kambiz N; Jonas, Elizabeth A et al. (2012) NAD kinase regulates the size of the NADPH pool and insulin secretion in pancreatic ?-cells. Am J Physiol Endocrinol Metab 303:E191-9
Hardwick, J Marie; Chen, Ying-bei; Jonas, Elizabeth A (2012) Multipolar functions of BCL-2 proteins link energetics to apoptosis. Trends Cell Biol 22:318-28

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