The basic underlying hypothesis of this proposal is that (Na+)i plays a role in regulating the amount of contractile proteins and the number of Na+ pump sites in cultured vascular smooth muscle cells. We will grow both arterial and venous smooth muscle cells in culture and quantitate contractile protein content (actin and myosin) by gel electrophoresis, Na+ pump site number by H3-ouabain binding and Na+ pump turnover rate by ouabain sensitive Rb86 uptake. After control characteristics have been established for these protein complexes, the cells will be grown under 4 specific conditions which will alter (Na+)i by significantly varied mechanisms: 1) O K+ or ouabain, which will inhibit the Na+ pump and increase (Na+)i; 2) monensin, aldosterone, angiotensin II and veratridine, which will increase (Na+)i and stimulate the pump; 3) amiloride which will decrease (Na+)i and inhibit the pump; and 4) cyclical stretch. The contractile protein content and Na+ pump site number will then be assessed after the appropriate growth period. In addition to these specific parameters, important additional parameters will be assessed to characterize the affect of these altered conditions. These will include determining the cell content of cAMP, DNA, Na+, K+, Ca++, the measurement of cell volume, intermediate filament content and H3-thymidine incorporation into DNA and morphological characterization (scanning and TEM). If an increase in contractile protein content or Na+ pump site number occurs, the role of Ca++ and amino acids in this process will be assessed by determining the amont of Na+i-Ca++o exchange and H3-alpha aminoisobutyric acid uptake. In addition, the effect of protein synthesis inhibitors (anisomycin, cyclohexamide) will be determined. Assessment of the amount of contractile protein and the number of Na+ pump sites under these systematically controlled conditions will lead to a better understanding concerning the role of the (Na+)i in regulating the amount of these proteins. Because of the similarity between these growth conditions and certain models of hypertension which involve increased (Na+)i, increased strain, and Na+ pump alterations, this work may lead to a hypothesis regarding the etiology of this disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL034280-07
Application #
3347030
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Project Start
1985-01-01
Project End
1992-12-31
Budget Start
1991-01-01
Budget End
1991-12-31
Support Year
7
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Baylor College of Medicine
Department
Type
Schools of Medicine
DUNS #
074615394
City
Houston
State
TX
Country
United States
Zip Code
77030
Kahn, A M; Allen, J C; Seidel, C L et al. (1994) Insulin inhibits serotonin-induced Ca2+ influx in vascular smooth muscle. Circulation 90:384-90
Feltes, T F; Seidel, C L; Dennison, D K et al. (1993) Relationship between functional Na+ pumps and mitogenesis in cultured coronary artery smooth muscle cells. Am J Physiol 264:C169-78
Kahn, A M; Seidel, C L; Allen, J C et al. (1993) Insulin reduces contraction and intracellular calcium concentration in vascular smooth muscle. Hypertension 22:735-42
Kahn, A M; Bishara, M; Cragoe Jr, E J et al. (1992) Effects of serotonin on intracellular pH and contraction in vascular smooth muscle. Circ Res 71:1294-304
Seidel, C L; Rickman, D; Steuckrath, H et al. (1991) Control and function of alterations in contractile protein isoform expression in vascular smooth muscle. Adv Exp Med Biol 304:315-25
Navran, S S; Allain, G; Garcia, H F et al. (1991) Serotonin-induced Na+/K+ pump stimulation in vascular smooth muscle cells. Evidence for coupling to multiple receptor mechanisms. J Pharmacol Exp Ther 256:297-303
Kahn, A M; Cragoe Jr, E J; Allen, J C et al. (1991) Effects of pHi on Na(+)-H+, Na(+)-dependent, and Na(+)-independent C1(-)-HCO3-exchangers in vascular smooth muscle. Am J Physiol 261:C837-44
Kahn, A M; Cragoe Jr, E J; Allen, J C et al. (1990) Na(+)-H+ and Na(+)-dependent Cl(-)-HCO3- exchange control pHi in vascular smooth muscle. Am J Physiol 259:C134-43
Allen, J C; Navran, S S; Seidel, C L et al. (1989) Intracellular Na+ regulation of Na+ pump sites in cultured vascular smooth muscle cells. Am J Physiol 256:C786-92
Seidel, C L; Wallace, C L; Dennison, D K et al. (1989) Vascular myosin expression during cytokinesis, attachment, and hypertrophy. Am J Physiol 256:C793-8

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