Journal: Neurophotonics

Article Title: Desensitization of opsin responses during all-optical interrogation depends on imaging parameters

doi: 10.1117/1.NPh.13.1.015007

Figure Lengend Snippet: Determining the impact of 920 nm two-photon imaging on 1035 nm photostimulation responses in targeted cells under common all-optical experimental conditions. (a) The opsin AAV-C1V1-Kv2.1 was injected into layer 2/3 of either the right somatosensory cortex ( n = 3 ) or right visual cortex ( n = 4 ) of mice transgenic for GCaMP6s in excitatory neurons. Calcium activity was imaged with a 920 nm laser over a 1.4 mm × 1.4 mm view around the opsin-expressing area (somatosensory cortex shown). Sixteen photostimulation targets were equally divided between two SLM phase masks. (b) Schematic showing the different raster-scanning imaging durations centered around 1035 nm photostimulation with a 15 s ISI. Each group of cells targeted by one SLM phase mask received ten blocks of 10 ms spiraling stimulation at 6 mW with a 20 ms interval between spirals. (c) Left: grand average response (over imaging conditions and then animals, n = 7 mice) ± SEM fit with a two-term exponential model. Right: cells from an example session color-coded by their individual responses to stimulation with the same holograms as in (a). (d) Responses on trials 1 and 10 as a function of imaging dose (* = p < 0.05 , Wilcoxon signed-rank test, n = 7 mice). Box plots show the median, upper quartile, lower quartile, maximum, and minimum, excluding outliers (< lower quartile − 1.5 × inter-quartile range OR > upper quartile + 1.5 × inter-quartile range). Each dot represents an animal. Grey lines connect data from the same animal. (e) ΔResponse (trial 10 − trial 1 response) as a function of imaging dose (n.s = p > 0.05 , Kruskal–Wallis test, n = 7 mice).

Article Snippet: The microscope used in this study was a Bruker Ultima 2Pplus equipped with a tunable wavelength laser for imaging (Chameleon Ultra II, Coherent, Saxonburg, Pennsylvania 16056, USA) and a 1035 nm fixed wavelength laser for photostimulation (Monaco 1035, Coherent).

Techniques: Imaging, Injection, Transgenic Assay, Activity Assay, Expressing

Journal: Neurophotonics

Article Title: Desensitization of opsin responses during all-optical interrogation depends on imaging parameters

doi: 10.1117/1.NPh.13.1.015007

Figure Lengend Snippet: Two-photon imaging at 920 nm increases 1035 nm photostimulation response desensitization in targeted cells at shorter ISIs. (a) Schematic showing the two 920 nm imaging conditions (either imaging constantly or in a 2 s window centered around photostimulation) and four possible ISIs (10/20/30/40 s). Each group of cells targeted by one SLM phase mask received ten blocks of 10 ms spiraling 1035 nm stimulation at 6 mW with a 20 ms interval between spirals. (b) Responses on trials 1 and 10 as a function of imaging dose, averaged across ISIs (* = p < 0.05 , Wilcoxon signed-rank test, n = 9 mice). Box plots show the median, upper quartile, lower quartile, maximum, and minimum, excluding outliers (< lower quartile − 1.5 × inter-quartile range OR > upper quartile + 1.5 × inter-quartile range). Each dot represents an animal. Grey lines connect data from the same animal. (c) ΔResponses (trial 10 − trial 1 response) as a function of imaging dose, averaged across ISIs (* = p < 0.05 , Wilcoxon signed-rank test, n = 9 mice) (d) Normalized response (mean trial response averaged across ISIs then animals/mean trial 1 response averaged across ISIs then animals) as a function of trial number, fit with a two-term exponential model (n = 9 mice). (e) Responses on trials 1 and 10 as a function of ISI, averaged across imaging conditions (* = p < 0.05 , Wilcoxon signed-rank test, n = 9 mice). (f) Average ΔResponse ± SEM fit with a first-degree polynomial ( n = 9 mice).

Article Snippet: The microscope used in this study was a Bruker Ultima 2Pplus equipped with a tunable wavelength laser for imaging (Chameleon Ultra II, Coherent, Saxonburg, Pennsylvania 16056, USA) and a 1035 nm fixed wavelength laser for photostimulation (Monaco 1035, Coherent).

Techniques: Imaging

Journal: Neurophotonics

Article Title: Desensitization of opsin responses during all-optical interrogation depends on imaging parameters

doi: 10.1117/1.NPh.13.1.015007

Figure Lengend Snippet: Two-photon imaging at 920 nm exacerbates photostimulation response desensitization of excited non-target cells. (a) Distribution of target, excited, inhibited, and non-responder cells across the field of view from the same example session as in Figs. 1(a) and . (b) Responses on trials 1 and 10 for the different cell classifications (* = p < 0.05 , Wilcoxon signed-rank test, n = 9 mice). Box plots show the median, upper quartile, lower quartile, maximum, and minimum, excluding outliers (< lower quartile − 1.5 × inter-quartile range OR > upper quartile + 1.5 × inter-quartile range). Each dot represents an animal. Gray lines connect data from the same animal. (c) Grand average response (over conditions then animals, n = 9 mice) fit with a two-term exponential model. (d) Average ΔResponse (trial 10 − trial 1 response) ± SEM ( n = 9 mice) of excited cells, fit with a first-degree polynomial. (e) Excited cell responses on trials one and ten with a 40 s ISI under different imaging conditions (* = p < 0.05 , Wilcoxon signed-rank test, n = 9 mice with somatosensory opsin expression). (f) ΔResponse in target and excited cells as a function of imaging dose and ISI. The same cells were stimulated across imaging conditions for the 10, 20, 30, and 40 s ISIs ( n = 9 mice). For a different population of cells, the same cells were stimulated across imaging conditions for the 15 s ISI ( n = 7 mice).

Article Snippet: The microscope used in this study was a Bruker Ultima 2Pplus equipped with a tunable wavelength laser for imaging (Chameleon Ultra II, Coherent, Saxonburg, Pennsylvania 16056, USA) and a 1035 nm fixed wavelength laser for photostimulation (Monaco 1035, Coherent).

Techniques: Imaging, Expressing