Multispectral near Infrared Imaging of Catecholamine Neurotransmitters with Fluorescent Nanosensors

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The aim of this project is to create near infrared (nIR) fluorescent nanosensors for neurotransmitters, measure and understand their kinetics/selectivity and use them for multispectral imaging of chemical communication between cells. This project is motivated by the importance of chemical communication in multicellular organisms. Neuronal networks such as those in the human brain are the most intriguing example for the complexity of this process. So far, there are not many analytical tools available to image small molecules such as neurotransmitters on small length scales and fast time scales in complex biological environments. For that reason, fast chemical communication between cells and most notably neurotransmitter release and diffusion in neural networks remains largely unexplored. We have recently introduced semiconducting single-walled carbon nanotubes (SWCNTs) as versatile building blocks for (molecular) sensors or probes. SWCNTs are hollow cylinders of one-atom-thick sheets of carbon. They possess an intrinsic size-dependent bandgap, which results in non-bleaching near infrared (nIR) fluorescence (900 nm - 1700 nm), a very beneficial spectral region for optical imaging because of reduced scattering and background. Our previous results indicate that SWCNTs functionalized non-covalently with certain single stranded DNA sequences are able to detect important catecholamine neurotransmitters including dopamine. The organic DNA phase (corona) around the SWCNTs interacts with dopamine, which affects and changes their fluorescence. The goal of this project is to create different sensors for catecholamines by changing the immobilized DNA sequence around SWCNTs. Most importantly, we will measure sensor kinetics in single-molecule experiments (on- and off-rates). For that purpose, we will make use of a nIR microscope that can image single nanosensors of different emission wavelength ("color"). Our theoretical simulations imply that only sensors with certain kinetic properties are useful for imaging of fast processes such as neurotransmitter release. The planned experiments will provide fundamental insights into SWCNT photophysics and how the corona (DNA sequence) affects rate constants and selectivity to structurally very similar catecholamine neurotransmitters (dopamine, epinephrine, norepinephrine). Furthermore, these experiments will reveal candidates with suitable properties for different envisioned sensing applications and kinetic simulations are used to predict their spatiotemporal resolution. The major idea of this project is that combining multiple sensors enhances sensitivity or selectivity. To this end SWCNTs of different color (emission wavelength) and different kinetics/selectivity will be imaged at the same time in a multiplexing approach (multispectral imaging). Finally, we want to demonstrate the versatility of this approach by resolving catecholamine release from cells.