FIGURE 7.

Robust in vitro and in vivo optogenetic control of the heart. (A) Adenoviral (AdV)-mediated ChR2-eYFP expression and functional measurements in NRVMs (MOI 25) and hiPSC-CMs (MOI 250) 2 days post-infection. Functional measurements were acquired using voltage- (di-4-ANBDQBS) and calcium- (Rhod4) sensitive dyes and example traces with optical pacing are shown. Cell nuclei were labeled with DAPI (blue) and AdV-infected cells expressed eYFP (green). Alpha-actinin staining (red) showed the cardiospecificity of the ChR2-eYFP infection. (B) Strength-duration curves for AAV9-mediated ChR2 expression in hiPSC-CMs. Conditions for infection included MOIs of 50,000–100,000 and NM applications of 500 mU/mL. Black arrows show the effect of NM treatment on lowering irradiance (mW/mm²) requirements; shown are voltage and calcium traces for the case of using MOI 50,000 without and with NM treatment. Data are presented as mean ± SEM (n = 3 per group). (C) AAV9-mediated ChR2-mCherry expression in the intact adult rat heart after 4 weeks results in optically sensitive myocardium in situ (left panel). A 0.8 mm diameter optical fiber was used to optically control electrical activity from the LV free wall as recorded using ECG (middle panel). Optical pacing resulted in an increased heart rate, as well as significant morphological changes in the QRS complex (right panel); the irradiance needed for this point stimulation was substantially higher than in vitro. Spatial scale bars are 50 μm in B,C. Temporal scale bars are as indicated.