Figure 2
Automated chemical dose response of two chemosensory neurons in C. elegans to the odor diacetyl from a single experiment. (a) Schematic of the multiwell plate prepared with buffer (b) and serial dilutions of diacetyl odor, from 10−9 (11.5 nM) to 10−3 (11.5 mM). Gray dot indicates the first well position. (b) A reusable two-arena pulse microfluidic device with four fluidic inlets, two worm-loading ports, and an outlet. The device switches between the stimulus inlet (in), shown with high concentration fluorescein, and unlabeled buffer, depending on which of two control channels (c1, c2) are flowing (Supplementary Video S2). Here, c1 flows while c2 is closed, such that stimulus fluid flows through the animal arenas. Scale bar, 500 µm. (c) Cross-section schematic of a worm head expressing GCaMP in a single neuron in the microfluidic assembly. Orange arrow points to the fluorescent cell body (soma) as in d. (d) Wide-field (4X) fluorescent image of 36 animals expressing GCaMP in either ASH (green arrows, left) or AWA (blue arrows, right) chemosensory neurons in the device. (e) Magnified images from panel d of animals expressing GCaMP in either ASH (left) and AWA (right) chemosensory neurons. Arrow points to soma. Scale bar, 100 µm. (f) Time course and average normalized fluorescence (∆F/F0) dose response traces for AWA (blue) and ASH (green) neuron cell bodies. Four pulses (10 s each) were delivered once per minute at each increasing concentration (or buffer). Lines and shading represent mean and SEM across four pulses, n = 72 traces per concentration and neuron type. (g) Mean peak normalized calcium responses for AWA neurons across seven increasing odor steps and buffer (b). Data points show population average and SEM (n = 18) for each of four repeated pulses per concentration. Trace represents a third-degree polynomial fit of each concentration’s pulse average. (h) Mean peak normalized calcium responses for ASH neurons, as described in g. (i) Mean peak normalized calcium responses of AWA plotted against ASH for each individual odor pulse. Error bars represent SEM for AWA (blue) and ASH (green), per concentration (corresponding dot color). (j) Neural network wiring diagram illustrating chemical synapses (arrows) and gap junctions (flat ends) for AWA and ASH sensory neurons and four first layer interneurons. Diacetyl is detected by different affinity receptors on each sensory neuron19,27.
