Dual Near Infrared Two-Photon Microscopy for Deep-Tissue Dopamine Nanosensor Imaging

Bonis-O’Donnell, Jackson Travis Del; Page, Ralph H.; Beyene, Abraham G.; Tindall, Eric G.; McFarlane, Ian R.; Landry, Markita P.

doi:10.1101/145912

Cited by 7 publications

(6 citation statements)

References 45 publications

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“…Two-photon absorption relies on the essentially simultaneous absorption of two lower-energy photons to promote a chromophore to a singlet excited state. Due to the use of longer-wavelength light, 29 , 30 two-photon microscopy enables imaging in thick tissue samples such as brain slices and intact brains. We showed previously that rhodol-based voltage indicators display voltage sensitivity in both traditional single-photon and two-photon microscopy contexts.…”

Section: Resultsmentioning

confidence: 99%

In Vivo Two-Photon Voltage Imaging with Sulfonated Rhodamine Dyes

et al. 2018

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Optical methods that rely on fluorescence for mapping changes in neuronal membrane potential in the brains of awake animals provide a powerful way to interrogate the activity of neurons that underlie neural computations ranging from sensation and perception to learning and memory. To achieve this goal, fluorescent indicators should be bright, highly sensitive to small changes in membrane potential, nontoxic, and excitable with infrared light. We report a new class of fluorescent, voltage-sensitive dyes: sulfonated rhodamine voltage reporters (sRhoVR), synthetic fluorophores with high voltage sensitivity, excellent two-photon performance, and compatibility in intact mouse brains. sRhoVR dyes are based on a tetramethyl rhodamine fluorophore coupled to a phenylenevinylene molecular wire/diethyl aniline voltage-sensitive domain. When applied to cells, sRhoVR dyes localize to the plasma membrane and respond to membrane depolarization with a fluorescence increase. The best of the new dyes, sRhoVR 1, displays a 44% ΔF/F increase in fluorescence per 100 mV change, emits at 570 nm, and possesses excellent two-photon absorption of approximately 200 GM at 840 nm. sRhoVR 1 can detect action potentials in cultured rat hippocampal neurons under both single- and two-photon illumination with sufficient speed and sensitivity to report on action potentials in single trials, without perturbing underlying physiology or membrane properties. The combination of speed, sensitivity, and brightness under two-photon illumination makes sRhoVR 1 a promising candidate for in vivo imaging in intact brains. We show sRhoVR powerfully complements electrode-based modes of neuronal activity recording in the mouse brain by recording neuronal transmembrane potentials from the neuropil of layer 2/3 of the mouse barrel cortex in concert with extracellularly recorded local field potentials (LFPs). sRhoVR imaging reveals robust depolarization in response to whisker stimulation; concurrent electrode recordings reveal negative deflections in the LFP recording, consistent with the canonical thalamocortical response. Importantly, sRhoVR 1 can be applied in mice with chronic optical windows, presaging its utility in dissecting and resolving voltage dynamics using two-photon functional imaging in awake, behaving animals.

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Section: Resultsmentioning

confidence: 99%

In Vivo Two-Photon Voltage Imaging with Sulfonated Rhodamine Dyes

et al. 2018

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“…Second, SWNT-based nanosensors rely on nIR fluorescence, which greatly reduces the impact of tissue scattering in the emission window and therefore may enable through-cranium imaging ( 25 ). nIRNSs are compatible with multiphoton imaging with an excitation of 1600 nm ( 50 ) and, as such, could permit nanoscale imaging of intact neuronal structures pending parallel developments in all-infrared microscopy, as has been shown with visible wavelength-emitting fluorophores ( 51 ). Third, nIRNSs exhibit robust nonphotobleaching photostability, allowing their use in long-term imaging experiments ( 52 ).…”

Section: Discussionmentioning

confidence: 99%

Imaging striatal dopamine release using a nongenetically encoded near infrared fluorescent catecholamine nanosensor

et al. 2019

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Neuromodulation plays a critical role in brain function in both health and disease, and new tools that capture neuromodulation with high spatial and temporal resolution are needed. Here, we introduce a synthetic catecholamine nanosensor with fluorescent emission in the near infrared range (1000–1300 nm), near infrared catecholamine nanosensor (nIRCat). We demonstrate that nIRCats can be used to measure electrically and optogenetically evoked dopamine release in brain tissue, revealing hotspots with a median size of 2 µm. We also demonstrated that nIRCats are compatible with dopamine pharmacology and show D2 autoreceptor modulation of evoked dopamine release, which varied as a function of initial release magnitude at different hotspots. Together, our data demonstrate that nIRCats and other nanosensors of this class can serve as versatile synthetic optical tools to monitor neuromodulatory neurotransmitter release with high spatial resolution.

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“…1560 nm pulsed laser irradiation) as is common in high resolution microscopy applications. 34 Such approaches when utilized with the NIR photocaging strategy describe here may find strong utility in intravital imaging applications 24-26 as well as ex vivo studies of organoids and embryo-like body development where high spatial control of protein delivery is necessary for proper cell organization. 27, 28…”

Section: Discussionmentioning

confidence: 99%

Multicolor Light-Induced Immune Activation via Polymer Photocaged Cytokines

Birnbaum

Sullivan

et al. 2022

Preprint

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Cytokines act as potent, extracellular signals of the human immune system and can elicit striking treatment responses in patients with autoimmune disease, tissue damage, and cancer. Yet despite their therapeutic potential, recombinant cytokine-mediated immune responses remain difficult to control as their administration is often systemic whereas their intended sites of action are localized. To address the challenge of spatially and temporally constraining cytokine signals, we recently devised a strategy whereby recombinant cytokines are reversibly inactivated via chemical modification with photo-labile polymers that respond to visible LED light. Extending this approach to enable both in vivo and multicolor immune activation, here we describe a strategy whereby cytokines appended with heptamethine cyanine-polyethylene glycol are selectively re-activated ex vivo using tissue-penetrating near-infrared (NIR) light. We show that NIR LED light illumination of caged, pro-inflammatory cytokines restores cognate receptor signaling and potentiates the activity of T cell-engager cancer immunotherapies ex vivo. Using combinations of visible- and NIR- responsive cytokines, we further demonstrate multi-wavelength optical control of T cell cytolysis ex vivo, as well as the ability to perform Boolean logic using multicolored light and orthogonally photocaged cytokine pairs as inputs, and T cell activity as outputs. Together, this work demonstrates a novel approach to control extracellular immune cell signals using light, a strategy that in the future may improve our understanding of and ability to treat cancer and other diseases.

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Dual Near Infrared Two-Photon Microscopy for Deep-Tissue Dopamine Nanosensor Imaging

Cited by 7 publications

References 45 publications

In Vivo Two-Photon Voltage Imaging with Sulfonated Rhodamine Dyes

In Vivo Two-Photon Voltage Imaging with Sulfonated Rhodamine Dyes

Imaging striatal dopamine release using a nongenetically encoded near infrared fluorescent catecholamine nanosensor

Multicolor Light-Induced Immune Activation via Polymer Photocaged Cytokines

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