Jennifer Kalb with Robert Harvey (School of Medicine, Dept. of Physiology and Biophysics)

Function of Beta_1 and Beta_2 Adrenergic Receptors in Cardiac Myocytes

The goal of this project is to determine whether beta_2 adrenergic receptor stimulation results in compartmentalized production of cAMP in intact cardiac ventricular myocytes. This goal will be accomplished by measuring the changes in cAMP activity in intact, isolated rat ventricular myocytes using PKA- and Epac-based FRET (Fluorescence Resonance Energy Transfer) biosensors. These will be introduced into the rat cardiac myocytes using adenovirus-based expression systems. FRET will be measured using images obtained from an inverted microscope (Olympus IX70) with camera (Hamamatsu Orca ER). The images will be acquired and analyzed with Simple PCI software. The drugs used will be Isoproterenol, a nonselective beta receptor activator, ICI-118,551, a selective beta_1 antagonist, and CGP 20712A, a selective beta_2 antagonist. Another component of this project is computational; after preliminary experiments have been run to obtain necessary parameters, a simulation will be run (in C++ or Java) and compared with experimental results. This project is motivated by the desire to obtain a clearer view of the selective, possibly differently compartmentalized, responses of beta_1 and beta_2 receptors, though they both utilize a common cAMP-dependent pathway, as well as the functioning of the FRET-based biosensors.