Yi, Bumjun Kang, Bok Eum Lee, Sungmoo Braubach, Sophie Baker, Bradley J. 2024-01-19T21:33:00Z 2024-01-19T21:33:00Z 2021-09-04 2018-10-12 2045-2322 https://pubs.kist.re.kr/handle/201004/120795 A bright, red-shifted Genetically Encoded Voltage Indicator (GEVI) was developed using a modified version of the fluorescent protein, tdTomato. Dimerization of the fluorescent domain for ArcLight-type GEVIs has been shown to affect the signal size of the voltage-dependent optical signal. For red-shifted GEVI development, tdTomato was split fusing a single dTomato chromophore to the voltage sensing domain. Optimization of the amino acid length and charge composition of the linker region between the voltage sensing domain and the fluorescent protein resulted in a probe that is an order of magnitude brighter than FlicRl at a resting potential of -70 mV and exhibits a ten-fold larger change in fluorescence (Delta F) upon 100 mV depolarization of the plasma membrane in HEK 293 cells. Unlike ArcLight, the introduction of charged residues to the exterior of dTomato did not substantially improve the dynamic range of the optical signal. As a result, this new GEVI, Ilmol, yields a 3-fold improvement in the signal-to-noise ratio compared to FlicRl despite a smaller fractional change in fluorescence of 4% per 100 mV depolarization of the plasma membrane. Ilmol expresses well in neurons resolving action potentials in neuronal cultures and reporting population signals in mouse hippocampa I acute brain slice recordings. Ilmol is the brightest red-shifted GEVI to date enabling imaging with 160-fold less light than Archonl for primary neuron recordings (50 mW/cm(2) versus 8W/cm(2)) and 600-fold less light than QuasAr2 for mouse brain slice recordings (500 mW/cm(2) versus 300W/cm(2)). This new GEVI uses a distinct mechanism from other approaches, opening an alternate engineering path to improve sensitivity and speed. English NATURE PUBLISHING GROUP ALL-OPTICAL ELECTROPHYSIOLOGY ENCODED VOLTAGE INDICATOR ACTION-POTENTIALS NEURONS TRANSMISSION SENSITIVITY DYNAMICS PH A dimeric fluorescent protein yields a bright, red-shifted GEVI capable of population signals in brain slice Article 10.1038/s41598-018-33297-y 1 SCIENTIFIC REPORTS, v.8 SCIENTIFIC REPORTS 8 scie scopus 000447125400021 2-s2.0-85054892055 Multidisciplinary Sciences Science & Technology - Other Topics Article ALL-OPTICAL ELECTROPHYSIOLOGY ENCODED VOLTAGE INDICATOR ACTION-POTENTIALS NEURONS TRANSMISSION SENSITIVITY DYNAMICS PH