Figure 1. ChR2 expressed in pharyngeal muscle cells (PMCs) enables photostimulation and light-induced pumping with high reproducibility.

(a) Expression of ChR2(H134R)::mCherry in PMCs, compared to a non-transgenic control. Dashed lines indicate pharynx position, dotted line indicates pharynx lumen, structural features of the pharynx are named. Scale bars: 20 μm. (b) The pharynx pump rate on food (visually counted) is unaffected by ChR2 expressed in PMCs (compared to wild type—wt), regardless of cultivation in absence or presence of 10 μM ATR (n = 10 animals each). (c) EPG recordings of activity in pharynxes of animals expressing ChR2::mCherry in PMCs. Compared are spontaneous activity (upper trace), activity stimulated by addition of serotonin (5-HT, 1 μM; middle trace), or in response to 1 Hz optical pacing (lower panel). (d) Comparison of overlaid original EPG recordings of spontaneous (black) and 2 Hz (10 ms, 470 nm) stimulated (grey) EPGs from four pharynxes expressing ChR2::mCherry. E (excitation/ contraction of pharynx) and R (relaxation/repolarization) peaks are indicated, traces have been aligned to the E spike. (e) Averaged, E-peak normalized EPG traces of spontaneous (14 traces) or 2 Hz paced pumps (20 traces) from different pharynxes (n = 4–5). A peak preceding the E-spike is observed in averaged 2 Hz paced pump traces, likely elicited by ChR2-photocurrents in PMCs, indicated by a blue circle. (f) Pump delay (time between start of stimulation pulse and occurrence of E-peak) is consistent during long recording periods, and was averaged from the indicated number of pumps/animals, for seconds 5–10 or 55–60 of one minute paced pumping.