A disturbance of the mechanism for intracellular Ca2+ homeostasis due to cerebral ischemic insult has been implicated in the irreversible damage leading to neuronal cell death. Although several known factors may contribute to the initial increase in [Ca2+]i during the early phase of an ischemic insult, the mechanism(s) leading to the progressive neuronal damage after prolonged ischemia and recirculation is not known. Due to the important role of Ins(1,4,5)P3 as second messenger for intracellular [Ca2+]i mobilization, it is hypothesized that prolonged ischemia- reperfusion results in irreversible modifications, either on the enzymes responsible for metabolism of Ins(1,4,5)P3 or its binding to the intracellular receptor. In turn, changes in Ins(1,4,5)P3 metabolism and/or binding may be an important mechanism leading to the altered cellular Ca2+ homeostasis associated with neuronal cell death. this hypothesis will be tested using a well established rat focal ischemia model (occlusion of middle cerebral artery) that resembles stroke in humans.
Specific aims are: (1) In vivo and in vitro experiments to determine the effects of transient (15 min) or prolonged (60 min) ischemic insult on poly- phosphoinositide breakdown and release of inositol phosphates. (2) Experiments to examine the effects of ischemia-reperfusion on metabolism of Ins(1,4,5)P3 by the 5-phosphatase and 3-kinase. Since there is strong evidence that Ins(1,4,5)P3 3-kinase, a key regulatory enzyme for Ins(1,4,5)P3 metabolism, may be irreversibly modified by a number of factors including Ca2+/calmodulin and Ca2+-proteases (calpain), studies will focus on effects of ischemia on the extent of proteolysis and levels of mRNA for this enzyme. (3) To examine the effect of ischemia on Ins(1,4,5)P3 receptor binding activity and the levels of mRNA encoding this receptor protein. (4) Upon establishing these experimental protocols, we will examine the effects of non-NMDA antagonists (NBQX) and protease inhibitors (leupeptin) on their ability to inhibit the ischemia-induced changes in Ins(1,4,5)P3 metabolism and binding. The long term goal is to understand the effects of focal ischemic insult on molecular and cellular mechanisms leading to the changes in intracellular Ca2+ homeostasis through alteration of the poly-phosphoinositide signaling pathways. Information resulted from this project will be important in designing strategies for therapeutic intervention towards alleviating the pathophysiology of ischemic tissue injury.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS030178-02
Application #
3417138
Study Section
Neurology A Study Section (NEUA)
Project Start
1992-08-01
Project End
1995-07-31
Budget Start
1993-08-01
Budget End
1994-07-31
Support Year
2
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Missouri-Columbia
Department
Type
Schools of Medicine
DUNS #
112205955
City
Columbia
State
MO
Country
United States
Zip Code
65211
Tong, W; Shah, D; Xu, J et al. (1999) Involvement of lipid mediators on cytokine signaling and induction of secretory phospholipase A2 in immortalized astrocytes (DITNC). J Mol Neurosci 12:89-99
Farwell, W; Simonyi, A; Scott, H et al. (1998) Effects of ischemic tolerance on mRNA levels of IP3R1, beta-actin, and neuron-specific enolase in hippocampal CA1 area of the gerbil brain. Neurochem Res 23:539-42
Wu, J M; Sun, G Y (1997) Effects of IL-1 beta on receptor-mediated poly-phosphoinositide signaling pathway in immortalized astrocytes (DITNC). Neurochem Res 22:1309-15
Sun, G Y; Hsu, C Y (1996) Poly-phosphoinositide-mediated messengers in focal cerebral ischemia and reperfusion. J Lipid Mediat Cell Signal 14:137-45
Simonyi, A; Zhang, J P; Sun, A Y et al. (1996) Chronic ethanol on mRNA levels of IP3R1, IP3 3-kinase and mGluR1 in mouse Purkinje neurons. Neuroreport 7:2115-8
Diehl, J A; Tong, W; Sun, G et al. (1995) Tumor necrosis factor-alpha-dependent activation of a RelA homodimer in astrocytes. Increased phosphorylation of RelA and MAD-3 precede activation of RelA. J Biol Chem 270:2703-7
Hersey, K; Hu, Z Y; Zhang, J P et al. (1995) In utero hypoxic ischemia decreases the cholinergic agonist-stimulated poly-phosphoinositide turnover in the developing rat brain. Neurochem Res 20:1477-82
Zhang, J P; Sun, G Y (1995) Free fatty acids, neutral glycerides, and phosphoglycerides in transient focal cerebral ischemia. J Neurochem 64:1688-95
Tong, W; Hu, Z Y; Sun, G Y (1995) Stimulation of group II phospholipase A2 mRNA expression and release in an immortalized astrocyte cell line (DITNC) by LPS, TNF alpha, and IL-1 beta. Interactive effects. Mol Chem Neuropathol 25:1-17
Sun, G Y; Zhang, J P; Lin, T A et al. (1995) Inositol trisphosphate, polyphosphoinositide turnover, and high-energy metabolites in focal cerebral ischemia and reperfusion. Stroke 26:1893-900

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