ABSTRACT

Purpose: Optogenetics is an emerging alternative to traditional electrical stimulation to initiate action potentials in activatable cells both ex vivo and in vivo. Optogenetics has been commonly used in mammalian neurons and more recently, it has been adapted for activation of cardiomyocytes and skeletal muscle. Therefore, the aim of this study was to evaluate the stimulation feasibility and sustain isometric muscle contraction and limit decay for an extended period of time (1s), using non-invasive transdermal light activation of skeletal muscle (triceps surae) in vivo.

Materials and Methods

 We used inducible Cre recombination to target expression of Channelrhodopsin-2 (ChR2(H134R)-EYFP) in skeletal muscle (Acta1-Cre) in mice. Fluorescent imaging confirmed that ChR2 expression is localized in skeletal muscle and does not have specific expression in sciatic nerve branch, therefore, allowing for non-nerve mediated optical stimulation of skeletal muscle. We induced muscle contraction using transdermal exposure to blue light and selected 10 Hz stimulation after controlled optimization experiments to sustain prolonged muscle contraction.

Results

Increasing the stimulation frequency from 10 Hz to 40 Hz increased the muscle contraction decay during prolonged 1s stimulation, highlighting frequency dependency and importance of membrane repolarization for effective light activation. Finally, we showed that optimized pulsed optogenetic stimulation of 10 Hz resulted in comparable ankle torque and contractile functionality to that of electrical stimulation.

Conclusions

 Our results demonstrate the feasibility and repeatability of non-invasive optogenetic stimulation of muscle in vivo and highlight optogenetic stimulation as a powerful tool for non-invasive in vivo direct activation of skeletal muscle.

中文翻译：

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体内肌肉收缩的光遗传学激活。

摘要

目的：光遗传学是传统电刺激的新兴替代方案，可在体外和体内启动可激活细胞的动作电位。光遗传学已普遍用于哺乳动物神经元，最近，它已适用于心肌细胞和骨骼肌的激活。因此，本研究的目的是评估刺激的可行性，并使用体内非侵入性透皮光激活骨骼肌（小腿三头肌）来长时间（1秒）维持等长肌肉收缩并限制衰减。

材料和方法

 我们使用诱导型 Cre 重组来靶向小鼠骨骼肌 (Acta1-Cre) 中通道视紫红质-2 (ChR2(H134R)-EYFP) 的表达。荧光成像证实ChR2表达位于骨骼肌中，并且在坐骨神经分支中没有特异性表达，因此，允许对骨骼肌进行非神经介导的光刺激。我们通过透皮暴露于蓝光来诱导肌肉收缩，并在受控优化实验后选择 10 Hz 刺激来维持长时间的肌肉收缩。

结果

将刺激频率从 10 Hz 增加到 40 Hz 增加了长时间 1 秒刺激期间的肌肉收缩衰减，突出了频率依赖性和膜复极化对于有效光激活的重要性。最后，我们证明优化的 10 Hz 脉冲光遗传学刺激可产生与电刺激相当的踝关节扭矩和收缩功能。

结论

 我们的结果证明了体内非侵入性肌肉光遗传学刺激的可行性和可重复性，并强调光遗传学刺激作为骨骼肌非侵入性体内直接激活的有力工具。