Imaging Striatal Dopamine Release Using a Non-Genetically Encoded Near-Infrared Fluorescent Catecholamine Nanosensor

Abraham G. Beyene; Kristen Delevich; Jackson Travis Del Bonis-O’Donnell; David J. Piekarski; Wan Chen Lin; A. Wren Thomas; Sarah J. Yang; Polina Kosillo; Darwin Yang; Linda Wilbrecht; Markita P. Landry

For electrical stimulation experiments, a bipolar stimulation electrode was positioned in field of view within the dorsomedial striatum identified using a 4X objective (Olympus xFluor 4x/340). Using 60X objective, the stimulation electrode was brought into contact with top surface of the brain slice and an imaging field of view was chosen at a nominal distance of 150 µm from the stimulation electrode within the dorsomedial striatum. All stimulation experiments were recorded at video frame rates of 9 frames per second (nominal) and single pulse electrical stimulations were applied after 200 frames of baseline were acquired. Each video acquisition lasted 600 frames. Stimulation amplitudes were staggered and each stimulation amplitude was repeated three times within a field of view. Slices were allowed to recover for 5 minutes between each stimulation with the excitation laser path shuttered. For optogenetic stimulation, a fiber-coupled 473 nm blue laser (OptoEngine LLC DPSS) was positioned in close proximity to the brain slice using a micromanipulator. Expression of ChR2 was confirmed via visible fluorescence imaging and an imaging field of view was chosen in dorsomedial striatum with robust expression level. Stimulation pulses (5 pulses, 5 ms duration per pulse, delivered at 25 Hz, 1 mW/mm 2 ) were delivered after acquiring 200 baseline frames and the video acquisition lasted 600 frames at nominal 9 frames per second. Drugs were bath applied to the imaging chamber through ACSF perfusion. ACSF with 10 µM of nomifensine or 1 µM of each DRD drug was used. When the effect of a drug needed to be evaluated, stimulation/imaging experiments were carried out with drug-free ACSF in an imaging field of view to collect drug-free data. Normal ACSF was then switched to ACSF prepared with the drug of interest and applied for 10 minutes before stimulation/imaging experiments resumed.