Inappropriate electrical activity gives rise to devastating brain disorders including, epilepsies and cell death following ischemia. These diseases trigger damaging inflammatory responses that elevate free arachidonic acid (AA) levels. Under these pathological conditions, excessive Ca2+ influx leads to cell death. By understanding the role that AA serves in modulating nerve cell excitability and survival, we hope to identify therapeutic targets that will minimize nerve cell death due to inflammatory signaling. My lab has found that AA inhibits two Ca2+ currents called L- and N-currents in superior cervical ganglion (SCG) neurons by promoting channel closing. AA also increases N-current amplitude by acting at a distinct site. As with inhibition, enhancement is mimicked by stimulating M1 muscarinic receptors and requires phospholipase A2, indicating functional relevance of AA's dual actions. We have measured increased AA release from individual SCG following muscarinic stimulation using gas chromatography (GC) and mass spectroscopy (MS). Three additional, distinct peaks with a mass/charge ratio (m/z) equal to that of AA are also released suggesting that SCGs acutely synthesize AA isoforms with novel double bond patterns. AA normally has four cis double bonds. Our findings appear to be the first study documenting AA isoforms in neurons. However a recent report described a peroxidation process in endothelial cells that generates AAs with one of the bonds in the trans- conformation (TAAs). Moreover TTAs appear to mediate oxidative stress-induced microvascular degeneration. These findings raise many questions surrounding the roles of AA versus its isoforms in normal and pathological nerve cell functioning. Because of the possible clinical relevance of AA isoforms as therapeutic targets for the treatment of ischemia, the following specific aims are proposed:
Aim I. Define the chemical structures of the AA isoforms released from SCG using GC-MS and nanospray ionization MS-MS. Characterizing the changes in AA double bond geometry and/or location will allow us to identify new potential proinflammatory signaling molecules.
Aim II. Separate, collect and test the four AA isoforms found in SCG neurons for their ability to modulate whole-cell Ca2+ currents of SCG neurons and recombinant channels transiently transfected into HEK293 cells.
Aim III. Determine whether any of the AA isoforms alter action potential firing. Whether particular AA isoforms promote nerve cell growth or death will also be tested. These studies will document at the cellular level the cumulative effect of AA (or AA isoforms) on membrane excitability and viability. If one or more of the isoforms modulates channel activity, action potential firing and/or cell survival, we will have identified a new signaling molecule(s) that may serve as a novel therapeutic target for treating ischemia. Inappropriate electrical activity gives rise to devastating brain disorders including, epilepsies and cell death following ischemia. These diseases trigger damaging inflammatory responses that elevate free arachidonic acid (AA) levels. We have discovered what appear to be novel isoforms of AA in a peripheral ganglion that regulates blood flow to the brain. By understanding the roles that AA and the novel AA isoforms serve in modulating nerve cell excitability, growth and survival, we hope to identify therapeutic targets that will minimize nerve cell death due to inflammatory signaling. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21NS057473-02
Application #
7422313
Study Section
Neural Degenerative Disorders and Glial Biology Study Section (NDGB)
Program Officer
Silberberg, Shai D
Project Start
2007-07-01
Project End
2010-06-30
Budget Start
2008-07-01
Budget End
2010-06-30
Support Year
2
Fiscal Year
2008
Total Cost
$177,734
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Physiology
Type
Schools of Medicine
DUNS #
603847393
City
Worcester
State
MA
Country
United States
Zip Code
01655
Roberts-Crowley, Mandy L; Mitra-Ganguli, Tora; Liu, Liwang et al. (2009) Regulation of voltage-gated Ca2+ channels by lipids. Cell Calcium 45:589-601