The overall goals of this research proposal are: (a) to investigate the role of phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis and its derived second messengers in agonist-induced muscle contraction in the sphincter and dilator smooth muscles of the iris-ciliary body, (b) to seek novel interactions between the cAMP and IP3-DG-CA2+ second messenger systems and the tension responses in these muscles, and (c) to investigate the involvement of PIP2 hydrolysis and its derived second messengers in the mechanism of receptor desensitization in the smooth muscles of the iris. Specifically: (I) To understand the biochemical-functional interactions between IP3-Ca2+-DG and cAMP signalling pathways in the smooth muscles of the iris-ciliary body we will: 1. Investigate the relationships between PG-induced IP3 accumulation, DG production, MLC phosphorylation, cAMP formation and muscle tension in the iris sphincter of different species. 2. Investigate the relationships between the effects of various agonists and antagonists, including adrenergic, muscarinic cholinergic, peptidergic and PGs, on the biochemical and functional responses in the bovine sphincter and dilator muscles. 3. Investigate the interaction between the PIP2 pathway and the adenylate cyclase pathway that is mediated through the activation of protein kinase C. 4. Investigate the effects of cAMP and/or its analogues on agonist- induced IP3 accumulation in iris sphincter membrane fractions. (11) To elucidate the role of PIP2 hydrolysis in the molecular mechanisms underlying receptor desensitization of the smooth muscles of the rabbit and bovine iris-ciliary body we will: (1) Investigate alterations in receptor-induced IP3 accumulation, DG production, MLC phosphorylation, cAMP formation and contraction due to in vitro and/or in vivo cholinergic, adrenergic, peptidergic and PG desensitization. 2. Investigate the possibility that protein kinase C-mediated phosphorylation of G proteins and/or receptors could be involved in the mechanism of desensitization. 3. Investigate the possibility that in vivo cholinergic and adrenergic desensitization of the eye may be reversed by administration of myo- inositol. 4. Characterize, through binding studies, the IP3 receptors in the iris muscle membranes and investigate the properties of IP3-induced Ca2+ fluxes in membrane fractions from normal and desensitized muscle. (III) We will investigate the role of MLC phosphatase in the mechanism of agonist-induced muscle relaxation in the iris sphincter. (IV) We will isolate and characterize the smooth muscle cells of the iris sphincter and dilator and investigate the properties of receptor-mediated PIP2 hydrolysis, Ca2+ mobilization, MLC phosphorylation and cAMP formation. Interaction between the cAMP and IP3-CA2+-DG signalling systems represent an important focal point for pharmacological manipulation and the proposed studies will lead to better understanding of aqueous humor dynamics and to the development of more effective and antiglaucoma drugs. In addition, our proposed studies will yield important information on: (a) Properties and function of muscarinic, alpha1-adrenergic, PG and peptidergic receptors in the anterior segment; (b) mechanisms of receptor desensitization in ocular tissues; (c) mechanisms of action of pharmacological agents in the eye; and (d) mechanistic insights into the role of PIP2 and its derived second messengers in smooth muscle function.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37EY004171-12
Application #
3484004
Study Section
Visual Sciences A Study Section (VISA)
Project Start
1981-09-30
Project End
1994-11-30
Budget Start
1992-12-01
Budget End
1993-11-30
Support Year
12
Fiscal Year
1993
Total Cost
Indirect Cost
Name
Medical College of Georgia (MCG)
Department
Type
Schools of Medicine
DUNS #
City
Augusta
State
GA
Country
United States
Zip Code
30912
Sharif, Naj A; Crider, Julie Y; Husain, Shahid et al. (2003) Human ciliary muscle cell responses to FP-class prostaglandin analogs: phosphoinositide hydrolysis, intracellular Ca2+ mobilization and MAP kinase activation. J Ocul Pharmacol Ther 19:437-55
Abdel-Latif, A A; Husain, S; Yousufzai, S Y (2000) Role of protein kinase C alpha and mitogen-activated protein kinases in endothelin-1-stimulation of cytosolic phospholipase A2 in iris sphincter smooth muscle. J Cardiovasc Pharmacol 36:S117-9
Husain, S; Abdel-Latif, A A (1999) Endothelin-1 activates p38 mitogen-activated protein kinase and cytosolic phospholipase A2 in cat iris sphincter smooth muscle cells. Biochem J 342 ( Pt 1):87-96
Yousufzai, S Y; Ali, N; Abdel-Latif, A A (1999) Effects of adrenomedullin on cyclic AMP formation and on relaxation in iris sphincter smooth muscle. Invest Ophthalmol Vis Sci 40:3245-53
Ding, K H; Latimer, A J; Abdel-Latif, A A (1999) Activation of particulate guanylyl cyclase by endothelins in cultured SV-40 transformed cat iris sphincter smooth muscle cells. Life Sci 64:161-74
Ding, K H; Ali, N; Abdel-Latif, A A (1999) Atrial natriuretic peptide provokes a dramatic increase in cyclic GMP formation and markedly inhibits muscarinic-stimulated Ca2+ mobilisation in SV-40 transformed cat iris sphincter smooth muscle (SV-CISM-2) cells. Cell Signal 11:87-94
Yousufzai, S Y; Abdel-Latif, A A (1998) Tyrosine kinase inhibitors suppress prostaglandin F2alpha-induced phosphoinositide hydrolysis, Ca2+ elevation and contraction in iris sphincter smooth muscle. Eur J Pharmacol 360:185-93
Husain, S; Abdel-Latif, A A (1998) Role of protein kinase C alpha in endothelin-1 stimulation of cytosolic phospholipase A2 and arachidonic acid release in cultured cat iris sphincter smooth muscle cells. Biochim Biophys Acta 1392:127-44
Yousufzai, S Y; Abdel-Latif, A A (1998) Calcitonin gene-related peptide relaxes rabbit iris dilator smooth muscle via cyclic AMP-dependent mechanisms: cross-talk between the sensory and sympathetic nervous systems. Curr Eye Res 17:197-204
Ding, K H; Abdel-Latif, A A (1997) Actions of C-type natriuretic peptide and sodium nitroprusside on carbachol-stimulated inositol phosphate formation and contraction in ciliary and iris sphincter smooth muscles. Invest Ophthalmol Vis Sci 38:2629-38

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