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. 2021 Dec 6;11:23424. doi: 10.1038/s41598-021-02832-9

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Kinetics of intrinsic dim-flash responses from various arrestin knock-out M1-ipRGCs. (A) Left: Grand average (black trace) of normalized mean dim-flash responses (grey traces, 5–15 pA) from seven Arr1,4^−/− M1-ipRGCs, seven βArr1^−/− M1-ipRGCs, eight βArr2^−/− M1-ipRGCs, and twenty-nine Opn4-Cre;βArr1,2^fl/fl M1-ipRGCs. In each panel, the red trace is the fit to the black trace by the convolution of two single-exponential declines, $e^{- t / τ_{1}} * e^{- t / τ_{2}}$ . Arr1,4^−/−: τ₁ = 1.0 s, τ₂ = 22.3 s. βArr1^−/−: τ₁ = 1.5 s, τ₂ = 21.1 s. βArr2^−/−: τ₁ = 2.0 s, τ₂ = 18.9 s. Opn4-Cre;βArr1,2^fl/fl: τ₁ = 1.1 s, τ₂ = 12.5 s. Right: Superposition of grand-averaged dim-flash responses from Arr1,4^−/−, βArr1^−/−, βArr2^−/−, Opn4-Cre;βArr1,2^fl/fl M1-ipRGCs (the black traces in Left panel), as well as WT M1-ipRGCs for comparison of the response waveforms. (B) Extraction of unitary response amplitude from an Opn4-Cre;βArr1,2^fl/fl M1-ipRGC. Left: Stationarity of the dim-flash response over time. Right: Similar waveforms shown by the square of the ensemble mean of a series of responses to identical dim flashes and the ensemble variance of the dim-flash responses. Unitary response = 0.43 pA. (C) Comparison between the unitary-response amplitudes from WT and Opn4-Cre;βArr1,2^fl/fl M1-ipRGCs. Bars and error bars represent mean ± SD. WT: 0.57 ± 0.22 pA (n = 7). Opn4-Cre;βArr1,2^fl/fl: 0.68 ± 0.23 pA (n = 7). p = 0.62, Wilcoxon test. (n.s., not statistically significant.) (D) Collected data on τ₁ and τ₂ from individual cells of each genotype. Bars and error bars represent mean ± SD. WT: τ₁ = 2.7 ± 1.4 s and τ₂ = 21.5 ± 4.1 s. Arr1,4^−/−: τ₁ = 1.4 ± 0.8 and τ₂ = 19.2 ± 3.0 s. βArr1^−/−: τ₁ = 1.8 ± 1.3 s and τ₂ = 19.4 ± 3.1 s. βArr2^−/−: τ₁ = 1.8 ± 0.6 s and τ₂ = 19.7 ± 3.4 s. Opn4-Cre;βArr1,2^fl/fl: τ₁ = 1.2 ± 0.8 s and τ₂ = 11.0 ± 5.9 s. Adjusted p-values for pairwise comparisons in τ₁ using Benjamini–Hochberg procedure are 0.06 for WT vs. Arr1,4^−/−, 0.18 for WT vs. βArr1^−/−, 0.56 for WT vs. βArr2^−/−, 1.0 × 10^–4 for WT vs. Opn4-Cre;βArr1,2^fl/fl. Adjusted p-values for pairwise comparisons in τ₂ using Benjamini–Hochberg procedure are 0.44 for WT vs. Arr1,4^−/−, 0.44 for WT vs. βArr1^−/−, 0.46 for WT vs. βArr2^−/−, 1.6 × 10^–4 for WT vs. Opn4-Cre;βArr1,2^fl/fl. (***, p < 0.001; n.s., not statistically significant).