The objective of this research proposal is to investigate the mechanisms of excitation-contraction coupling in the gallbladder smooth muscle and in doing so, prepare the applicant for a long term career as an independent investigator. Excitation-contraction coupling of gallbladder smooth muscle is particularly important because abnormal gallbladder contractility may play a primary pathophysiologic role in the formation of gallstones. Investigation of excitation-contraction coupling in feline gallbladder smooth muscle will provide a background for studies in human gallbladder smooth muscle in gallstone disease.
The specific aims of this project are: 1) to characterize the utilization of intracellular and extracellular calcium sources in excitation-contraction coupling in isolated gallbladder myocytes stimulated by three agonists - acetylcholine, cholecystokinin, and potassium; 2) to determine the transmembrane electrical properties of gallbladder smooth muscle in muscle strips under basal conditions and to determine the effects of classical neurotransmitters and hormones on electromechanical coupling by evaluating the voltage-tension relationships of graded concentrations of extracellular potassium, acetylcholine, and cholecystokinin; 3) to evaluate the effects of endogenous neurotransmitters on intracellular electrical activity and muscle tension of gallbladder smooth muscle using electric field stimulation to activate intrinsic enteric nerves of the gallbladder; and 4) to characterize the excitation-contraction coupling abnormalities in human gallbladder smooth muscle from patients with gallstones. Several sophisticated research techniques will be employed in the conduct of this project: simultaneous intracellular electrical and isometric muscle tension recordings will be performed to investigate the electromechanical coupling of gallbladder smooth muscle; freshly dispersed isolated gallbladder myocytes will be used to measure cell shortening; cytosolic calcium concentrations will be evaluated using the calcium-sensitive fluorescent dye indo-1; and cell membrane calcium channels will be characterized using the whole cell patch clamp technique. In the process of elucidating the mechanisms of gallbladder excitation-contraction coupling, the relationship between cell membrane potential, calcium utilization, membrane calcium currents and gallbladder smooth muscle contraction will be better understood. The candidate is a physician-scientist who is determined to develop a career committed to the study of gastrointestinal motility. To achieve this goal, the candidate will continue to work with and learn from two respected basic scientists in gastrointestinal smooth muscle and cardiac muscle physiology. Interaction with these investigators will provide a stimulating and productive environment, so that the candidate can conduct productive research and position himself to compete successfully for independent extramural funding in the future.
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