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

Abstract

Neuromodulation plays a critical role in brain function in both health and disease. New optical tools are needed that can image neuromodulation with high spatial and temporal resolution, which will add an important new dimension of information to neuroscience research. Here, we demonstrate the use of a catecholamine nanosensor with fluorescent emission in the 1000-1300 nm near-infrared window to measure dopamine transmission in ex vivo brain slices. These near-infrared catecholamine nanosensors (nIRCats) represent a broader class of nanosensors that can be synthesized from non-covalent conjugation of single wall carbon nanotubes (SWNT) with single strand oligonucleotides. We show that nIRCats can be used to detect catecholamine efflux in brain tissue driven by both electrical stimulation or optogenetic stimulation. Spatial analysis of electrically evoked signals revealed dynamic regions of interest approximately 2 microns in size in which transients scaled with simulation intensity and lasted 5-10 seconds. Optogenetic stimulation of dopaminergic terminals produced similar transients, while optogenetic stimulation of glutamatergic terminals showed no effect on nIRCat signal. Furthermore, bath application of nomifensine prolonged nIRCat fluorescence signal, consistent with reuptake blockade of dopamine. These nanosensors may be advantageous for future use because they i) do not require virus delivery, gene delivery, or protein expression, ii) their near-infrared fluorescence facilitates imaging in optically scattering brain tissue and is compatible for use in conjunction with other optical neuroscience tool sets, and iii) the broad availability of unique near-infrared colors have the potential for simultaneous detection of multiple neurochemical signals. Together, these data suggest nIRCats and other nanosensors of this class can serve as versatile new optical tools to report dynamics of extracellular neuromodulation in the brain.All rights reserved. No reuse allowed without permission.was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.

Fluorescent Sp3 Defect-Tailored Carbon Nanotubes Enable NIR-II Single Particle Imaging in Live Brain Slices at Ultra-Low Excitation Doses

“…We quantitatively demonstrated single nanotube imaging in the NIR-II region at high signal-to-noise ratios using unprecedentedly low excitation intensities (100 W/cm 2 ) that are an order of magnitude lower than those previously reported for SWCNTs. This work paves the way toward the application of sp 3 defect-tailored nanotubes as single-molecule probes [39] and chemical/molecular sensors [15,40] in live biological tissues.…”

“…For this, we chose live brain tissues as an archetypical system because single SWCNT detection and tracking were shown to provide unique knowledge about the brain extracellular space environment. [10,15,39] Organotypic rat brain tissue slices were prepared from P5-7 rat pups and maintained in culture medium at 35 °C/5% CO 2 until imaging. Unf-or f-SWCNTs were delivered identically into brain tissues by incubation (120 minutes) and mounted in a ludin chamber containing the artificial cerebrospinal fluid under the microscope for imaging.…”

“…In addition, light excitation necessary to achieve efficient single SWCNT PL must be compatible with live tissue integrities. [13] Due to these constraints, imaging low concentrations of SWCNTs, [14,15] notably single SWCNTs [10] into live tissue remains challenging. Note that reduced sensitivity and higher noise of detectors at NIR wavelengths complicates the task.…”

High Throughput Evolution of Near Infrared Serotonin Nanosensors

“…The resulting ssDNA-SWNT moiety can then selectively bind a molecular analyte for its optical detection [23][24][25][26] . The process of generating ssDNA-SWNT nanosensor candidates, and their screening against target analytes, has since revealed optical nanosensors for neurotransmitters including a (GT)6-SWNT nanosensor for dopamine 25 which was subsequently used to image dopamine dynamics in the brain striatum and capture the influence of dopamine receptor drugs on dopamine dynamics at the level of individual synapses 27 . Despite the nascent utility of SWNT-based nanosensor technology, the nanosensor screening process involves a heuristic approach in which each nanosensor candidate must be synthesized individually before screening, limiting candidate nanosensor libraries to a few dozen, thus restricting the throughput of nanosensor discovery.…”

“…nIRHT nanosensors were immobilized onto a (3-aminopropyltriethoxysilane) (APTES)-treated glass slide, and placed in a flow chamber with a syringe pump. The nanosensor chamber was exposed to three cycles of 100 µM 5-HT in PBS subsequently rinsed with PBS only, where the nearinfrared fluorescence response was imaged on an inverted microscope as previously described 27 (Figure 3d-f, and Supplementary Movie 1). We observe nIRHT fluorescence increase and decrease as predicted by the influx and washing of 5-HT, respectively.…”