Abstract

Contractile function of cardiac muscle is strongly dependent on fine regulation of intracellular ion levels. The primary aim of this project is to increase the understanding of the transsarcolemmal transport of sodium and calcium by the Na-Ca exchange mechanism. The physiological significance of Na-Ca exchange is controversal, and the long range objective is to relate molecular findings to intact preparations. Five approaches are proposed as follows: 1. Identification and isolation of the Na-Ca exchange protein. Molecular characterization of the Na-Ca exchanger requires that the protein first be isolated. Excellent progress towards this goal has been achieved, and several approaches to complete the purification procedure are planned. 2. Expression of Na-Ca exchange in Xenopus oocytes. To begin to examine the molecular biology of Na-Ca exchange, cardiac mRNA is being injected into oocytes to induce expression of exchange activity. Experiments are then planned to fractionate mRNA and, eventually, to clone the Na-Ca exchange protein. 3. Interaction of the Na-Ca exchanger with the membrane lipid environment. Na-Ca exchange activity is very sensitive to changes in the lipid environment of native sarcolemmal vesicles. Lipid-exchanger interactions are now being explored in more detail using solubilization/reconstitution techniques. 4. Kinetics of Na-Ca exchange. Much basic characterization of Na-Ca exchange is planned using both native and reconstituted cardiac sarcolemmal vesicles. For example, stoichiometry, effects of intravesicular Ca, and Na-Cd exchange will be examined. 5. Long term regulation of Na-Ca exchange. The response of cultured neonatal heart cells to chronic changes in intracellular ion levels will be examined. For examples, Na-Ca exchange will be measured in sarcolemmal vesicles isolated from control cells and cells which have been exposed to ouabain for 24 hours. It is speculated that the cells will adjust rates of synthesis and degradation of different ion transporters in response to interventions.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL027821-08
Application #
3339345
Study Section
Cardiovascular Study Section (CVA)
Project Start
1981-08-01
Project End
1993-03-31
Budget Start
1989-04-01
Budget End
1990-03-31
Support Year
8
Fiscal Year
1989
Total Cost
Indirect Cost

  Institution

Name
University of California Los Angeles
Department
Type
Schools of Medicine
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095

  Publications

Porzig, H; Philipson, K D (1996) Epitope mapping using random fragment expression libraries in lambda phages. Methods Mol Biol 66:257-68
Porzig, H; Li, Z; Nicoll, D A et al. (1993) Mapping of the cardiac sodium-calcium exchanger with monoclonal antibodies. Am J Physiol 265:C748-56
Li, Z P; Burke, E P; Frank, J S et al. (1993) The cardiac Na+-Ca2+ exchanger binds to the cytoskeletal protein ankyrin. J Biol Chem 268:11489-91
Philipson, K D; Nicoll, D A; Li, Z (1993) The cardiac sodium-calcium exchanger. Soc Gen Physiol Ser 48:187-91
Matsuoka, S; Nicoll, D A; Reilly, R F et al. (1993) Initial localization of regulatory regions of the cardiac sarcolemmal Na(+)-Ca2+ exchanger. Proc Natl Acad Sci U S A 90:3870-4
Hryshko, L V; Nicoll, D A; Weiss, J N et al. (1993) Biosynthesis and initial processing of the cardiac sarcolemmal Na(+)-Ca2+ exchanger. Biochim Biophys Acta 1151:35-42
Shieh, B H; Xia, Y; Sparkes, R S et al. (1992) Mapping of the gene for the cardiac sarcolemmal Na(+)-Ca2+ exchanger to human chromosome 2p21-p23. Genomics 12:616-7
Yip, R K; Blaustein, M P; Philipson, K D (1992) Immunologic identification of Na/Ca exchange protein in rat brain synaptic plasma membrane. Neurosci Lett 136:123-6
Komuro, I; Wenninger, K E; Philipson, K D et al. (1992) Molecular cloning and characterization of the human cardiac Na+/Ca2+ exchanger cDNA. Proc Natl Acad Sci U S A 89:4769-73
Li, Z; Smith, C D; Smolley, J R et al. (1992) Expression of the cardiac Na(+)-Ca2+ exchanger in insect cells using a baculovirus vector. J Biol Chem 267:7828-33

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