The goal of the proposed research is to explain the role of extracellular adenine nucleotides, particularly adenosine 5'- triphosphate (ATP), in normal cardiac endothelial cell (EC) function where release of factors that participate in the regulation of blood flow may be important in normal cardiac function and in cardiovascular disease. ECs have been shown to release ATP and autacoids such as endothelium-dependent relaxing factor and prostacyclin in response to various stimuli, including ATP itself. In addition, we propose that ECs can convert adenosine 5'-diphosphate (ADP) to ATP extracellularly to maintain the presence of ATP-mediated relaxation of the coronary and resistance vessels down-stream from the site of ATP release. We suggest that control of ATP appearance outside the EC and its eventual conversion to adenosine is an important component of regional blood flow control in the normal heart. As it is not clear how release of ATP or conversion of ADP to ATP by ECs is regulated, specific aims have been designed as follows: 1. Test the hypothesis that the release of ATP from ECs and the conversion of ADP to ATP by ECs can be measured in an intact coronary artery and that ATP release can support nucleoside triphosphate formation. Furthermore, to test the hypothesis that conversion of ADP to ATP can be inhibited in situ and determine the affect of hypoxia on extracellular generation of ATP. 2. Test the hypothesis that cardiac EC release of ATP in response to agonists occurs both luminally and abluminally and test the effect of hypoxia and re-oxygenation on this process. 3. Explore the manner in which ECs release ATP and test the hypothesis that the elaboration of ATP by stimulated ECs is associated, mechanistically, with the ability of ECs to release nitric oxide. 4. Assess the role of ecto-nucleoside diphosphate kinase and ecto- adenylate kinase in the extracellular production of ATP by ECs and test effects of hypoxia and reoxygenation on the activity of these enzymes. We will approach these aims with biochemical, pharmacological, molecular, cell biological and microscopic methods employing intact blood vessels, cells in primary culture and subcellular fractions.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL056422-02
Application #
2609377
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Program Officer
Sigmon, Hilary D
Project Start
1996-12-01
Project End
2001-11-30
Budget Start
1998-02-13
Budget End
1998-11-30
Support Year
2
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Nevada Reno
Department
Pharmacology
Type
Schools of Medicine
DUNS #
146515460
City
Reno
State
NV
Country
United States
Zip Code
89557
Buxton, Iain L O; Yokdang, Nucharee; Matz, Robert M (2010) Purinergic mechanisms in breast cancer support intravasation, extravasation and angiogenesis. Cancer Lett 291:131-41
Buxton, Iain L O; Anzinger, Joshua J (2008) Agonist-specific regulation of inositol phosphate metabolism in cardiac endothelial cells. Proc West Pharmacol Soc 51:23-6
Oxhorn, Brian C; Sanguinetti, Amy R; Mastick, Cynthia Corley et al. (2005) c-Abl is required for staurosporine-induced caspase activity. Proc West Pharmacol Soc 48:110-7
Oxhorn, Brian C; Buxton, Iain L O (2003) Caveolar compartmentation of caspase-3 in cardiac endothelial cells. Cell Signal 15:489-96
Tichenor, Stephen D; Malmquist, Nicholas A; Buxton, Iain L O (2003) Dissociation of cGMP accumulation and relaxation in myometrial smooth muscle: effects of S-nitroso-N-acetylpenicillamine and 3-morpholinosyndonimine. Cell Signal 15:763-72
Tichenor, Stephen D; Buxton, Iain L O; Johnson, Paul et al. (2002) Excitatory motor innervation in the canine rectoanal region: role of changing receptor populations. Br J Pharmacol 137:1321-9
Oxhorn, Brian C; Hirzel, Douglas J; Buxton, Iain L O (2002) Isolation and characterization of large numbers of endothelial cells for studies of cell signaling. Microvasc Res 64:302-15
Oxhorn, B C; Wadia, R; Buxton, I L (2001) Caspase-3 is localized to endothelial caveolar domains. Proc West Pharmacol Soc 44:45-8
Buxton, I L; Kaiser, R A; Oxhorn, B C et al. (2001) Evidence supporting the Nucleotide Axis Hypothesis: ATP release and metabolism by coronary endothelium. Am J Physiol Heart Circ Physiol 281:H1657-66