4.5. Electropharyngeogram

To perform electrical recordings of pharynx pumping (electropharyngeogram, EPG), we placed the Nemametrix Screen Chip System on an inverted Zeiss Axiovert 200 microscope equipped with an LD A-Plan 20× objective. In order to minimize interfering electrical noise during the recordings, we used a system grounding shield. Baseline noise was typically between 10 and 40 µV. The experiments were performed at 20 °C.

For each experiment, we picked 100 worms from the culture plate and washed them in 1.5 mL of 0.2 µm filtered M9 buffer +0.1% Tween. Worms were then washed 4× with 0.2 µm filtered M9 buffer, then once in M9 buffer containing 2.3 mM serotonin, and they were finally suspended in 1 mL of M9 buffer containing 2.3 mM serotonin and allowed to settle for 15 min. All the experiments were performed between 15 and 120 min of the initial serotonin exposure. We loaded a NemaMetrix screen chip system (NemaMetrix, Eugene OR; Cat # SK100) with a fresh SC40 screen chip (NemaMetrix, Eugene OR; Cat # SKU: 0002) and added M9 buffer containing 2.3 mM serotonin. After initiating the NemAcquire software and recording the basal power line noise, worms were vacuum-loaded onto the Nemametrix Screen Chip SC40 to start the experiment. The 1-hz high-pass and 50-hz notch filter settings were selected. A 180-s EPG recording was made for each animal, and records from 20–25 animals were obtained for each replicate. Records were analyzed with the NemAnalysis v0.2 software. All the experiments with a frequency of less than 0.1 Hz or pump duration coefficient of variation bigger than 50% were rejected.