Two-photon activation of endogenous store-operated calcium channels without optogenetics

Pan Cheng; Wanyi Tang; Hao He

doi:10.1117/12.2286586

23 February 2018 Two-photon activation of endogenous store-operated calcium channels without optogenetics

Pan Cheng, Wanyi Tang, Hao He

Proceedings Volume 10498, Multiphoton Microscopy in the Biomedical Sciences XVIII; 1049828 (2018) https://doi.org/10.1117/12.2286586
Event: SPIE BiOS, 2018, San Francisco, California, United States

Abstract

Store-operated calcium (SOC) channels, regulated by intracellular Ca²⁺ store, are the essential pathway of calcium signaling and participate in a wide variety of cellular activities such as gene expression, secretion and immune response¹. However, our understanding and regulation of SOC channels are mainly based on pharmacological methods. Considering the unique advantages of optical control, optogenetic control of SOC channels has been developed². However, the process of genetic engineering to express exogenous light-sensitive protein is complicated, which arouses concerns about ethic difficulties in some research of animal and applications in human. In this report, we demonstrate rapid, robust and reproducible two-photon activation of endogenous SOC channels by femtosecond laser without optogenetics. We present that the short-duration two-photon scanning on subcellular microregion induces slow Ca²⁺ influx from extracellular medium, which can be eliminated by removing extracellular Ca2+. Block of SOC channels using various pharmacological inhibitors or knockdown of SOC channels by RNA interference reduce the probability of two-photon activated Ca²⁺ influx. On the contrary, overexpression of SOC channels can increase the probability of Ca²⁺ influx by two-photon scanning. These results collectively indicate Ca²⁺ influx through two-photon activated SOC channels. Different from classical pathway of SOC entry activated by Ca²⁺ store depletion, STIM1, the sensor protein of Ca²⁺ level in endoplasmic reticulum, does not show any aggregation or migration in this two-photon activated Ca²⁺ influx, which rules out the possibility of intracellular Ca²⁺store depletion. Thereby, we propose this all-optical method of two-photon activation of SOC channels is of great potential to be widely applied in the research of cell calcium signaling and related biological research.

Citation Download Citation

Pan Cheng, Wanyi Tang, and Hao He "Two-photon activation of endogenous store-operated calcium channels without optogenetics", Proc. SPIE 10498, Multiphoton Microscopy in the Biomedical Sciences XVIII, 1049828 (23 February 2018); https://doi.org/10.1117/12.2286586

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