The focus of this proposal is the study of oxidative stress in vivo using microdialysis sampling. Oxidative stress produces reactive oxygen species and ultimately free radicals that cause damage to DNA and other biomolecules. We propose to study the physiology of oxidative stress using in vivo microdialysis sampling at specific tissue sites during ischemia and reperfusion in the heart and brain. Microdialysis will be used to monitor oxygen free radicals (OFRs), an endogenous antioxidant that may serve as a protective agent (glutathione), and biomarkers for oxidative DNA damage (8-hydroxydeoxyguanosine) and lipid peroxidation (malondialdehye). In order to carry out the proposed studies, it will be necessary to first develop a suite of analytical methods to determine each of the target analytes in microdialysis samples. Microdialysis sampling results in samples of a few microliters or less. Therefore, methods based on capillary electrophoresis (CE), micellar electrokinetic chromatography (MEKC), and capillary electrokinetic chromatography (CEC) will be developed. The modified nucleosides and antioxidants are charged and electroactive. OFRs will be stabilized using radical trapping agents introduced through the microdialysis probe that produce electroactive products. All of these analytes will be detected using electrochemical methods. Malondialdehyde will be derivatized to a fluorescent analyte and determined by laser induced fluorescence. The role of OFR production will then be studied in vivo during ischemia and reperfusion in the heart and brain using microdialysis sampling and the newly developed methods of analysis. Studies will be aimed at elucidating the role of various proposed mechanisms of OFR formation during ischemia and reperfusion. In particular, the role xanthine oxidase plays in the production of OFRs in the brain will be investigated. OFR formation as a function of catecholamine release will be studied in both the heart and brain. Microdialysis will be used to locally administer inhibitors of xanthine oxidase and of catecholamine release and uptake in vivo to determine the role of these pathways.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL069014-03
Application #
6923672
Study Section
Special Emphasis Panel (ZRG1-SSS-6 (10))
Program Officer
Liang, Isabella Y
Project Start
2003-08-01
Project End
2007-07-31
Budget Start
2005-08-01
Budget End
2006-07-31
Support Year
3
Fiscal Year
2005
Total Cost
$210,720
Indirect Cost
Name
University of Kansas Lawrence
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
076248616
City
Lawrence
State
KS
Country
United States
Zip Code
66045
Crick, Eric W; Osorio, Ivan; Frei, Mark et al. (2014) Correlation of 3-mercaptopropionic acid induced seizures and changes in striatal neurotransmitters monitored by microdialysis. Eur J Pharm Sci 57:25-33
Whitaker, Gillian; Lunte, Craig E (2010) Investigation of microdialysis sampling calibration approaches for lipophilic analytes: doxorubicin. J Pharm Biomed Anal 53:490-6
Price, Kristin E; Lunte, Craig E; Larive, Cynthia K (2008) Development of tissue-targeted metabonomics. Part 1. Analytical considerations. J Pharm Biomed Anal 46:737-47
Whitaker, Gillian; Lillquist, Amy; Pasas, Stephanie A et al. (2008) CE-LIF method for the separation of anthracyclines: application to protein binding analysis in plasma using ultrafiltration. J Sep Sci 31:1828-33
Gilinskii, M A; Lunte, C E (2007) [Norepinephrine of the rat myocardium during repeated ischemia] Ross Fiziol Zh Im I M Sechenova 93:860-9
Crick, Eric W; Osorio, Ivan; Bhavaraju, Naresh C et al. (2007) An investigation into the pharmacokinetics of 3-mercaptopropionic acid and development of a steady-state chemical seizure model using in vivo microdialysis and electrophysiological monitoring. Epilepsy Res 74:116-25
Krisko, Ryan M; McLaughlin, Kieran; Koenigbauer, Michael J et al. (2006) Application of a column selection system and DryLab software for high-performance liquid chromatography method development. J Chromatogr A 1122:186-93
Arnett, Stacy D; Osbourn, Damon M; Moore, Kimberly D et al. (2005) Determination of 8-oxoguanine and 8-hydroxy-2'-deoxyguanosine in the rat cerebral cortex using microdialysis sampling and capillary electrophoresis with electrochemical detection. J Chromatogr B Analyt Technol Biomed Life Sci 827:16-25
Price, Kristin E; Vandaveer, Shannon S; Lunte, Craig E et al. (2005) Tissue targeted metabonomics: metabolic profiling by microdialysis sampling and microcoil NMR. J Pharm Biomed Anal 38:904-9
Hoque, Mohammed E; Arnett, Stacy D; Lunte, Craig E (2005) On-column preconcentration of glutathione and glutathione disulfide using pH-mediated base stacking for the analysis of microdialysis samples by capillary electrophoresis. J Chromatogr B Analyt Technol Biomed Life Sci 827:51-7

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