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

Coronal mouse brain slices were prepared as described previously. 28 Slices were subsequently incubated with 2 mg/L nIRCats for 15 minutes to enable sensors to diffuse into the brain tissue ( Figure 2a ). Slices were subsequently rinsed to remove excess nIRCats and incubated in standard artificial cerebrospinal fluid (ACSF) for another 15 minutes before imaging. This method, modified from Godin et al. 29 who demonstrated that SWNT localize in the ECS of acute brain slices, enabled even and widespread labeling of the coronal slice, including the dorsal striatum ( Figure 2c ). Imaging of nIRCat fluorescence modulation in dorsal striatum was accomplished with a custom-built visible and nearinfrared microscope to enable concurrent imaging of both visible (400 nm -750 nm) and near-infrared (750 nm -1700 nm) wavelengths on the same detector ( Figure 2b ). Briefly, a 785 nm laser for excitation of nIRCats and mercury bulb for generating brightfield images were directed onto the back focal plane of an epifluorescence upright microscope, and imaging channels were selected using a sliding mirror. Serially, either brightfield or nearinfrared images were collected on a Ninox Vis-SWIR 640 broadband camera (Raptor Photonics) with appropriate dichroic filters (Methods) and a 60X water dipping objective (Nikon) providing an imaging field of 178 µm by 142 µm, containing hundreds of dopaminergic terminals. Experimental schematic depicting preparation of acute brain slices and subsequent incubation in 2 mg/L nIRCat solution to load the nanosensors into brain tissue. (b) Schematic of visible/near-infrared microscope. A 785 nm CW laser is beam-expanded and co-aligned with a mercury vapor lamp and directed into the objective with dichroic filter cubes. Emitted photons are filtered through a 900 nm long-bass filter and are relayed onto the sensor of a broadband InGaAs camera that is sensitive to both visible and near-infrared wavelengths. (c) Dorsomedial striatum from mouse acute slice imaged in brightfield (top) and near-infrared (bottom) after tissue nanosensor loading. Scale bars = 10 µm.