Abstract
Background and PurposeOptogenetic control of electromechanical coupling in vascular smooth muscle cells (VSMCs) is emerging as a powerful research tool with potential applications in drug discovery and therapeutics. However, the precise ionic mechanisms involved in this control remain unclear.Experimental ApproachCell imaging, patch‐clamp electrophysiology and muscle tension recordings were used to define these mechanisms over a wide range of light stimulations.Key ResultsTransgenic mice expressing a channelrhodopsin‐2 variant [ChR2(H134R)] selectively in VSMCs were generated. Isolated VSMCs obtained from these mice demonstrated blue light‐induced depolarizing whole‐cell currents. Fine control of artery tone was attained by varying the intensity of the light stimulus. This arterial response was sufficient to overcome the endogenous, melanopsin‐mediated, light‐evoked, arterial relaxation observed in the presence of contractile agonists. Ca2+ entry through voltage‐gated Ca2+ channels, and opening of plasmalemmal depolarizing channels (TMEM16A and TRPM) and intracellular IP3 receptors were involved in the ChR2(H134R)‐dependent arterial response to blue light at intensities lower than ~0.1 mW·mm−2. Light stimuli of greater intensity evoked a significant Ca2+ influx directly through ChR2(H134R) and produced marked intracellular alkalinization of VSMCs.Conclusions and ImplicationsWe identified the range of light intensity allowing optical control of arterial tone, primarily by means of endogenous channels and without substantial alteration to intracellular pH. Within this range, mice expressing ChR2(H134R) in VSMCs are a powerful experimental model for achieving accurate and tuneable optical voltage‐clamp of VSMCs and finely graded control of arterial tone, offering new approaches to the discovery of vasorelaxant drugs.
Highlights
Optogenetics is an experimental approach involving the use of light in conjunction with microbial opsins to control cell, tissue or organ function (Fenno et al, 2011; Lin, 2011; Entcheva, 2013)
We systematically examined the effects of a wide range (0.01–12.1 mW·mmÀ2) of blue light intensities on both the magnitude of ChR2(H134R) current in isolated vascular smooth muscle cell (VSMC) and the extent of contraction of artery rings
To assess the utility of vascular optogenetics in different vascular beds, we determined the degree of ChR2(H134R) expression in various artery types obtained from ChR2(H134R)SM mice, as this may influence the arterial sensitivity to blue light stimuli
Summary
Optogenetics is an experimental approach involving the use of light in conjunction with microbial opsins (light-sensitive proteins) to control cell, tissue or organ function (Fenno et al, 2011; Lin, 2011; Entcheva, 2013). ChR2 and its variants have been used extensively as experimental tools in the field of neuroscience to define brain circuitry (Boyden et al, 2005; Zhang et al, 2007; Yizhar et al, 2011) These opsins have been harnessed to control the function of non-neuronal cell types. Optogenetics has been used to gain insights into the role of arterioles and pericyte-covered capillaries in the control of cerebral blood flow (Hill et al, 2015) These important studies have demonstrated the utility and potential of optogenetics in vascular research. Important mechanistic aspects, need to be addressed to enable a complete interpretation of vascular optogenetics experiments and to exploit this technology in an in vivo setting This technology has the potential of offering new approaches to the identification and testing of new compounds that could relieve vasoconstriction. Patch-clamp electrophysiology and muscle tension recordings were used to define these mechanisms over a wide range of light stimulations
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