This project proposes that adenyl purines, compounds known to cause contraction of uterus, participate in the regulation of human parturition.
The specific aims of this research are to discover the origins, binding sites and molecular signaling events that subserve the contractile action of adenyl purines in uterine smooth muscle. The working hypothesis is that adenyl purines, both adenosine and ATP from one or more sources stimulate purinergic receptors in myometrial smooth muscle leading to uterine contraction. We propose that adenyl purines participate in, or initiate labor and as such, may provide a focal point for understanding preterm labor in humans. Studies will be conducted with tissues and freshly isolated cells from both pregnant guinea pigs and humans. The research plan will be separated into three areas of investigation. (1) We will determine the origins of adenyl purines in the uterus. HPLC and radiolabeling studies will investigate the role of putative purinergic nerves as well as the co-release of purines from adrenergic and cholinergic nerves in myometrium. Paracrine release of purines from the smooth muscle of myometrium itself, or from endometrium will also be tested. (2) The receptor targets for adenyl purines will be investigated with radioligand binding studies in cells and tissues and will determine the presence and distribution of purinergic receptors of the P1 and P2 subtype in circular and longitudinal smooth muscles. These methods will also be used to determine the nature of regulation of receptor affinity for agonist and the role of coupling proteins such as GTP-binding proteins in the molecular action of these receptors.(3) Pharmacological studies will characterize the actions of adenyl purines in pregnant uterus. Mechanical contraction of tissues, electrophysiological measurements of cellular ionic currents and biochemical measurements of receptor action will be made. Biochemical and electrophysiological studies will employ tissue segments and purified smooth muscle cells and will concentrate on the role of purines in the regulation of cellular Ca+2. Biochemical studies will include measurement of inositol phosphates, arachidonic acid release, cyclic nucleotide metabolism and the action of protein kinases. Cellular imaging using fluorescent pH dyes will be used to determine the regulation of phospholipase A2 through stimulation of N+/H+ exchange. Intracellular calcium fluctuations will be studied in cells and tissues using intracellular fluorescent dyes.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD026227-03
Application #
3327614
Study Section
Special Emphasis Panel (SRC (17))
Project Start
1989-09-01
Project End
1994-05-31
Budget Start
1991-06-01
Budget End
1992-05-31
Support Year
3
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Nevada Reno
Department
Type
Schools of Medicine
DUNS #
146515460
City
Reno
State
NV
Country
United States
Zip Code
89557
Zhang, L; Buxton, I L (1998) Measurement of phosphoinositols and phosphoinositides using radio high-performance liquid chromatography flow detection. Methods Mol Biol 105:47-63
Bradley, K K; Buxton, I L; Barber, J E et al. (1998) Nitric oxide relaxes human myometrium by a cGMP-independent mechanism. Am J Physiol 275:C1668-73
Kuenzli, K A; Bradley, M E; Buxton, I L (1996) Cyclic GMP-independent effects of nitric oxide on guinea-pig uterine contractility. Br J Pharmacol 119:737-43
Yang, S; Buxton, I L; Probert, C B et al. (1996) Evidence for a discrete UTP receptor in cardiac endothelial cells. Br J Pharmacol 117:1572-8
Zhang, L; Bradley, M E; Buxton, I L (1995) Inositolpolyphosphate binding sites and their likely role in calcium regulation in smooth muscle. Int J Biochem Cell Biol 27:1231-48
Shuttleworth, C W; Weinert, J S; Sanders, K M et al. (1995) Detection of nitric oxide release from canine enteric neurons. J Auton Nerv Syst 56:61-8
Yang, S; Cheek, D J; Westfall, D P et al. (1994) Purinergic axis in cardiac blood vessels. Agonist-mediated release of ATP from cardiac endothelial cells. Circ Res 74:401-7
Duckles, S P; Buxton, I L (1994) Neuropeptide Y potentiates norepinephrine-stimulated inositol phosphate production in the rat tail artery. Life Sci 55:103-9
Bradley, M E; Kuenzli, K A; Buxton, I L (1993) Adenosine-stimulated contraction in nonpregnant guinea pig myometrium does not involve cyclooxygenase. J Pharmacol Exp Ther 264:1033-9
Smith, T K; Ward, S M; Zhang, L et al. (1993) Beta-adrenergic inhibition of electrical and mechanical activity in canine colon: role of cAMP. Am J Physiol 264:G708-17

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