Two-Photon Neuronal and Astrocytic Stimulation with Azobenzene-Based Photoswitches

Izquierdo-Serra, Mercè; Gascón-Moya, Marta; Hirtz, Jan J.; Pittolo, Silvia; Poskanzer, Kira E.; Ferrer, Èric; Alibés, Ramón; Busquè, Félix; Yuste, Rafael; Hernando, Jordi; Gorostiza, Pau

doi:10.1021/ja5026326

Cited by 108 publications

(173 citation statements)

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“…Similarly, the slower thermal decay of L-MAG0 460 suggests that this PTL should be relatively insensitive to the method of 2P stimulation. Indeed, our findings of efficient 2P photoswitching of L-MAG0 460 by 2P-DH complement the recent report of two antenna-bearing MAGs with similar thermal relaxation, where LiGluR photocurrents were evoked by 2P laser scanning (32). Notably, in contrast to the study in ref.…”

Section: Discussionsupporting

confidence: 90%

“…2D, Lower, green circles), qualitatively consistent with a cis isomer (42). However, the 1-ns absorption spectrum extends to longer wavelengths than observed in photostationary measurements on related compounds (32), and may represent a vibrationally "hot" cis-L-MAG0 460 . Transient absorption following 2P excitation with the laser fundamental (800 nm, 100 fs, 16 mW) produced a similar photoproduct (Fig.…”

Section: Resultssupporting

confidence: 52%

“…Two-photon digital holography (2P-DH) is an effective method to flexibly target light to many regions simultaneously, or to produce an extended image to excite large patches of membrane for summation of single-channel ionic currents (13). It was recently shown that 2P scanning excitation of MAG 2p could depolarize neurons, but scans of nearly 1 s were necessary (32). We asked whether faster photoactivation might be attained with 2P-DH.…”

Section: Resultsmentioning

confidence: 99%

“…It was recently reported that the asymmetric substitution in second generation PTLs enabled efficient 2P excitation in two antennae-bearing PTLs, MAG 2p , and MAGA 2p (32). However, that study concluded that the bistable nature of a first generation MAG could ultimately generate a larger effect, albeit at a slower rate, by building up a population of cis isomers.…”

Section: Significancementioning

confidence: 99%

“…Two families of MAGs have been synthesized to date: First generation MAGs have a symmetrically-substituted azobenzene core, e.g., L-MAG0 (21) and D-MAG0 (30). Second generation MAGs have an asymmetrically-substituted azobenzene core, e.g., L-MAG0 460 (31) and MAG 2p (32). The latter MAGs are single-color photoswitches incorporating an electron-donating amine and an electron-withdrawing amide substitution, creating a "push-pull" system that both red-shifts the trans isomer absorption spectrum and reduces the thermal stability of the cis isomer (31,33).…”

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confidence: 99%

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Two-photon brightness of azobenzene photoswitches designed for glutamate receptor optogenetics

Carroll

Berlin

Levitz

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

113

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Mammalian neurotransmitter-gated receptors can be conjugated to photoswitchable tethered ligands (PTLs) to enable photoactivation, or photoantagonism, while preserving normal function at neuronal synapses. "MAG" PTLs for ionotropic and metabotropic glutamate receptors (GluRs) are based on an azobenzene photoswitch that is optimally switched into the liganding state by blue or near-UV light, wavelengths that penetrate poorly into the brain. To facilitate deep-tissue photoactivation with near-infrared light, we measured the efficacy of two-photon (2P) excitation for two MAG molecules using nonlinear spectroscopy. Based on quantitative characterization, we find a recently designed second generation PTL, , to have a favorable 2P absorbance peak at 850 nm, enabling efficient 2P activation of the GluK2 kainate receptor, LiGluR. We also achieve 2P photoactivation of a metabotropic receptor, LimGluR3, with a new mGluR-specific PTL, D-MAG0 460 . 2P photoswitching is efficiently achieved using digital holography to shape illumination over single somata of cultured neurons. Simultaneous Ca 2+ -imaging reports on 2P photoswitching in multiple cells with high temporal resolution. The combination of electrophysiology or Ca 2+ imaging with 2P activation by optical wavefront shaping should make second generation PTL-controlled receptors suitable for studies of intact neural circuits.optogenetics | pharmacology | multiphoton | photoswitch | azobenzene M odern neurobiology relies heavily on optical microscopy to observe, and, increasingly, to manipulate (1, 2), biological processes in live tissue. Among these methods, 2-photon-excited fluorescence microscopy (2PM) with near-infrared (NIR) light has emerged as an important technique for extending optical microscopy to highly scattering tissue (3, 4). Remarkably, barely 25 years after the first 2P-excited fluorescence image was published (5), 2PM is now performed in awake, behaving animals (4, 6-8). Naturally, optical manipulations in the brain can benefit from the many advantages of 2PM (9). In particular, the inherent spatial confinement of 2P absorption is critical for optical manipulation of individual cells (10) in the intact mammalian brain, where it is difficult to control the spatial extent of gene expression or confine soluble reagents. However, 2P-excited optogenetics is not yet as widespread in adoption as 2PM, being widely perceived to require sophisticated optical techniques (11-13) or reagent concentrations that compromise pharmacological specificity (2, 14).The rapid time to adoption of 2PM owes at least some credit to the availability of spectroscopic data on the 2P-excited efficacy, or brightness, of synthetic and genetically encoded fluorophores (15-17). Brightness, defined for fluorophores as the product of absorption cross-section and fluorescence quantum yield, gives the experimenter an objective metric to assess fluorescent reporters and identify appropriate optical parameters, such as the optimal excitation wavelength and range of light intensities (18). By ...

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Section: Discussionsupporting

confidence: 90%

Section: Resultssupporting

confidence: 52%

Section: Resultsmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

mentioning

confidence: 99%

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Two-photon brightness of azobenzene photoswitches designed for glutamate receptor optogenetics

Carroll

Berlin

Levitz

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

113

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Bridged Azobenzenes and Their Biological Applications

Warner

Keane

Saint-Louis

et al. 2019

Patai's Chemistry of Functional Groups

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Capable of rapid and reversible conformational change in response to irradiation, azobenzene (AB) derivatives have been used in chemical systems to introduce on/off switchability and allow for photoregulation of the properties of such systems. Following a brief introduction of tuning AB absorption via substitution, this chapter summarizes the selected recent reports of applications of bridged ABs in biological systems or systems having potential biomedical utility. ABs have been used to achieve photocontrol of drug‐delivery systems, molecular actuators, deoxyribonucleic acid (DNA) nanomachines, enzyme activity, peptide structure and activity, and ion channels. This chapter describes how researchers have incorporated AB into biological systems as a photoresponsive switch to afford photocontrol of the chemical activity in such systems. The various works detailed in this chapter showcase the versatility of AB for use as a light‐responsive molecular switch in the photoregulation of biological processes. Additionally, light‐responsive AB‐derived macroscopic materials are also discussed, given their biomedical potential for use as artificial muscle.

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Photoswitching of Ion Channels

Fehrentz¹,

Broichhagen²

2022

Molecular Photoswitches

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Two-Photon Neuronal and Astrocytic Stimulation with Azobenzene-Based Photoswitches

Cited by 108 publications

References 50 publications

Two-photon brightness of azobenzene photoswitches designed for glutamate receptor optogenetics

Two-photon brightness of azobenzene photoswitches designed for glutamate receptor optogenetics

Bridged Azobenzenes and Their Biological Applications

Photoswitching of Ion Channels

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