Abstract
Dopamine circuit has been shown to associate with locomotion, sleep and arousal, decision making, and associative learning in mammals and fly. To understand the mechanism of dopaminergic-associated behaviors and disorders, we aim to visualize dopamine release in vivo. Currently, monitoring the dynamics and distribution of dopamine released in live is unattainable; the traditional ways to measure dopamine level in neurons is cumbersome and lacks of temporal and spatial resolutions. Our goal is to develop a probe that can switch fluorescence upon detecting dopamine release from dopaminergic neurons in vivo. By modifying our previously established cytosolic probe MMG1, in which we used intrinsic spectral properties of MAO B to switch GFP emission upon dopamine binding (we have coined this phenomena as “shield effect”) as an intracellular dopamine sensor, I have developed different membrane dopamine sensors that could potentially use for detecting dopamine release. Current results show that two full-length constructs, MSG11 (MAO B-ste2p-sfGFP11) and MSG (MAO B-ste2p-GFP), and two MAO B C-terminal truncated probes could be embedded on the cell membrane with expected topology. Additional tests are currently ongoing to validate the shield effect and enzyme activity of these sensors. My ultimate goal is to make the transgenic flies and test the probes in the brain. With this type of molecular probe on hand, we may have a better way to determine how brain process information in dopamine circuits under different environmental stimulations and behavioral responses.