The long term objectives of the investigator are to understand the precise regulation of cyclic nucleotide synthesis and their function in normal tissues and various disease states. The work proposed will emphasis the regulation and role of cyclic GMP. Model systems will be developed to examine hormonal and E. Coli heat-stable enterotoxin regulation of guanylate cyclase activity and cyclic GMP accumulation in both intact cell and cell-free systems. The mechanisms of coupling of E. Coli heat-stable enterotoxin binding with cyclic GMP synthesis intestinal mucosa and the effects of endothelial dependent vasodilators on vascular smooth muscle cyclic GMP synthesis will be used as example of two model systems for guanylate cyclase regulation. Furthermore experiments will be conducted to reconstitute hormonal regulation of guanylate cyclase in cell-free systems using model cell culture lines and tissues. In addition red cell ghosts will be loaded with various macromolecules involved in cyclic nucleotide synthesis, hydrolysis and function. These loaded ghosts will be fused to cell lines and tissues of interest to modify cyclic nucleotide metabolism and function in order to assess their role in various processes and/or phosphorylation events. Thus, several approaches will be conducted simultaneously to provide new techniques and information to examine the regulation of cyclic nucleotide formation and action.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK030787-06
Application #
3229641
Study Section
Pharmacology A Study Section (PHRA)
Project Start
1981-09-01
Project End
1988-06-30
Budget Start
1986-07-01
Budget End
1987-06-30
Support Year
6
Fiscal Year
1986
Total Cost
Indirect Cost
Name
Stanford University
Department
Type
Schools of Medicine
DUNS #
800771545
City
Stanford
State
CA
Country
United States
Zip Code
94305
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Nakane, M; Schmidt, H H; Pollock, J S et al. (1993) Cloned human brain nitric oxide synthase is highly expressed in skeletal muscle. FEBS Lett 316:175-80
Pollock, J S; Nakane, M; Buttery, L D et al. (1993) Characterization and localization of endothelial nitric oxide synthase using specific monoclonal antibodies. Am J Physiol 265:C1379-87
Sheng, H; Gagne, G D; Matsumoto, T et al. (1993) Nitric oxide synthase in bovine superior cervical ganglion. J Neurochem 61:1120-6
Schmidt, H H; Gagne, G D; Nakane, M et al. (1992) Mapping of neural nitric oxide synthase in the rat suggests frequent co-localization with NADPH diaphorase but not with soluble guanylyl cyclase, and novel paraneural functions for nitrinergic signal transduction. J Histochem Cytochem 40:1439-56
Pollock, J S; Klinghofer, V; Forstermann, U et al. (1992) Endothelial nitric oxide synthase is myristylated. FEBS Lett 309:402-4
Mitchell, J A; Kohlhaas, K L; Matsumoto, T et al. (1992) Induction of NADPH-dependent diaphorase and nitric oxide synthase activity in aortic smooth muscle and cultured macrophages. Mol Pharmacol 41:1163-8
Warner, T D; Budzik, G P; Matsumoto, T et al. (1992) Regional differences in endothelin converting enzyme activity in rat brain: inhibition by phosphoramidon and EDTA. Br J Pharmacol 106:948-52
Klabunde, R E; Kimber, N D; Kuk, J E et al. (1992) NG-methyl-L-arginine decreases contractility, cGMP and cAMP in isoproterenol-stimulated rat hearts in vitro. Eur J Pharmacol 223:1-7
Schmidt, H H; Smith, R M; Nakane, M et al. (1992) Ca2+/calmodulin-dependent NO synthase type I: a biopteroflavoprotein with Ca2+/calmodulin-independent diaphorase and reductase activities. Biochemistry 31:3243-9

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