Molecular Recognition of Dopamine with Near Infrared Dual Excitation-Emission Two-Photon Microscopy of Nanosensors

Neuromodulatory neurotransmitters in the brain are implicated in a wide variety of psychiatric and neurological disorders, yet their quantification in the biologically-relevant milieu of the living brain remains inaccessible. Non-invasive, optical methods to directly quantify neurotransmission will transform our ability to study and understand psychiatric disorders. Recently, functionalized single walled carbon nanotubes (SWNTs) have emerged as promising near-infrared (NIR) fluorescent nanosensors for the sensitive and rapid detection of neurotransmitter dopamine. These sensors fluorescence in the NIR-II window (1000-1700 nm) where absorption and scattering of light is minimal, making them optimally suited for non-invasive imaging in brain tissue. However, the photoexcitation of SWNT dopamine nanosenors is currently accomplished with highly-scattering visible light. To further optimize dopamine nanosensors for in vivo imaging, we show that non-linear photoexcitation of our SWNT nanosensors can be achieved using a 1560 nm laser excitation source, falling within the optimal NIR-II window. Using a femtosecond pulsed erbium laser, we observe two-photon induced fluorescence of SWNT dopamine nanosensors and measure nanosensor quantum yield and 2-photon cross sections. Furthermore, we demonstrate SWNT-based molecular recognition of dopamine using two-photon excitation, confirming that the molecular recognition principle of our sensors is compatible with non-linear excitation. Finally, we show significantly improved fluorescence spatial resolution attained by two-photon excitation when imaging in strongly scattering tissue phantoms, motivating future work using these sensors for in vivo, real-time sensing of dopamine in brain tissue.