DOI
https://doi.org/10.25772/ATMF-GA72
Defense Date
2011
Document Type
Dissertation
Degree Name
Doctor of Philosophy
Department
Physiology
First Advisor
Murthy Karnam
Abstract
Gastrointestinal (GI) smooth muscle possesses distinct regional and functional properties that distinguish it from other types of visceral and vascular smooth muscle. On the basis of electrical properties and contractile phenotype, GI smooth muscles have been classified into phasic (non-sphinteric) and tonic (sphinteric) smooth muscles. The biochemical basis of phasic and tonic phenotypes of smooth muscle is not clear and is the major question of inquiry of the present study. Phosphorylation of Ser19 on the 20 kDa myosin light chain (MLC) is essential for acto-myosin interaction and contraction in both phasic and tonic muscles. The levels of MLC20 phosphorylation are regulated by Ca2+/calmodulin-dependent MLC kinase (MLCK) and MLC phosphatase (MLCP), and the activity of these enzymes are in turn regulated by various signaling molecules whose expression and activity are important in determining the strength and duration of their activity. The signaling proteins are AMP kinase (MLCK activity), Rho kinase, zipper-interacting protein kinase (ZIPK), CPI-17 and telokin (MLCP activity), phosphodiesterase 5 (PDE5) and multi-drug resistance protein 5 (MRP5). The overarching goal of the dissertation is to identify the differences in the signaling pathways that regulate MLCK and MLCP activities, and thus MLC20 phosphorylation and muscle function. Using biochemical, molecular and functional approaches, and antrum (distal stomach) and fundus (proximal stomach) of rabbit stomach as models of phasic and tonic smooth muscles, respectively, the present study characterized important differences in the signaling pathways that highly correlate with the contractile phenotype. These include: 1) tissue-specific expression of contractile proteins such as myosin heavy chain isoforms, actin, caldesmon, calponin, - and β-tropomyosin, smoothelin-A and -B; 2) higher expression of AMPK, selective feedback inhibition of MLCK activity via AMPK-mediated phosphorylation, and higher expression of telokin and activation of MLCP correlate with the rapid cyclical contractile function in phasic muscle; 3) higher expression and activation of Rho kinase/ZIPK/MYPT1 and PKC/CPI-17 pathways, preferential inhibition of MLCP activity, and sustained phosphorylation of MLC20 correlate with the sustained contraction in tonic muscle; and 4) rapid termination of cGMP signal and muscle relaxation by preferential degradation and efflux of cGMP via higher expression of PDE5 and MRP5, respectively, correlate with the brief relaxation and rapid restoration of contraction in tonic muscle. It is anticipated that these findings could be important in providing the underlying mechanisms involved in the pathophysiology of smooth muscle function and new insights for the development of therapeutic agents that should act on smooth muscle in the gut to treat motility disorders as well as in other regions such as airways and vascular smooth muscle where similar intracellular mechanisms may prevail.
Rights
© The Author
Is Part Of
VCU University Archives
Is Part Of
VCU Theses and Dissertations
Date of Submission
May 2011