Turns decrease the likelihood of mechanosensory-evoked reversals.

(A) Closed-loop optogenetic stimulation is delivered to animals as they crawl based on their current behavior. (B) Optogenetic stimulation is delivered to gentle-touch mechanosensory neurons in worms that are either moving forward (top row) or turning (bottom row). (C) The probability of a reversal is shown in response to stimulation during forward movement or turn. Responses are also shown for a low-light no-stimulation control. This figure only is a reanalysis of recordings from [19]. The number of stimulation events, from left to right: 6,002, 1,114, 5,996, and 1,050. (D) The probability of reversal in response to stimulation during turning is shown broken down further by turn subtype: escape-like turns “Esc” and isolated turns “Iso.” N = 6,002, 602, 512, 5,996, 599, and 451 stim events, from left to right. The number of plates for forward and turn context are 29 and 47, respectively. The 95% confidence intervals for population proportions are reported; *** indicates pp>0.05 via 2 proportion Z-test. Exact p values for all the statistical tests are listed in S1 Table. All data underlying this figure can be found at https://doi.org/10.25452/figshare.plus.23903202.</p

Expression pattern of <i>lim-4</i> promoter.

Fluorescence/Bright field, merged image of AML496 worms showing the expression of eGFP driven by lim-4 promoter using (Plim-4::gtACR2::SL2::eGFP) expression vector. eGFP can be seen in the neurons RIV, SMB, and SAA. (EPS)</p

Strains used.

Animals must integrate sensory cues with their current behavioral context to generate a suitable response. How this integration occurs is poorly understood. Previously, we developed high-throughput methods to probe neural activity in populations of Caenorhabditis elegans and discovered that the animal’s mechanosensory processing is rapidly modulated by the animal’s locomotion. Specifically, we found that when the worm turns it suppresses its mechanosensory-evoked reversal response. Here, we report that C. elegans use inhibitory feedback from turning-associated neurons to provide this rapid modulation of mechanosensory processing. By performing high-throughput optogenetic perturbations triggered on behavior, we show that turning-associated neurons SAA, RIV, and/or SMB suppress mechanosensory-evoked reversals during turns. We find that activation of the gentle-touch mechanosensory neurons or of any of the interneurons AIZ, RIM, AIB, and AVE during a turn is less likely to evoke a reversal than activation during forward movement. Inhibiting neurons SAA, RIV, and SMB during a turn restores the likelihood with which mechanosensory activation evokes reversals. Separately, activation of premotor interneuron AVA evokes reversals regardless of whether the animal is turning or moving forward. We therefore propose that inhibitory signals from SAA, RIV, and/or SMB gate mechanosensory signals upstream of neuron AVA. We conclude that C. elegans rely on inhibitory feedback from the motor circuit to modulate its response to sensory stimuli on fast timescales. This need for motor signals in sensory processing may explain the ubiquity in many organisms of motor-related neural activity patterns seen across the brain, including in sensory processing areas.</div

Turns decrease the likelihood of interneuron evoked reversals, except for AVA.

(A) Anatomical connectivity showing chemical (arrows) and electrical (resistor symbol) synapses among the anterior mechanosensory neurons, downstream interneurons, and turning-associated neurons. (B) Probability of a reversal response is shown for 3 s optogenetic stimulation to the listed neurons either during forward movement or immediately after the onset of turning. Strains are listed in Table 1. Illumination was 80 μW/mm2 red light to activate Chrimson in AVE or AVA, 300 μW/mm2 blue light to activate ChR2 in RIM or AIB, and 340 μW/mm2 to activate ChR2 in AIZ. Error bars indicate 95% confidence intervals for population proportions; *** indicates pp>0.05 via two-proportion Z-test, and p value for AVA stimulation group is 0.125. Exact p values for all the statistical tests are listed in S1 Table. N = 2,612, 601, 883, 107, 880, 511, 1,007, 342, 409, and 191 stimulus events, from left-to-right, measured across the following number of plates: 16, 27, 12, 19, 4, 24, 8, 16, 8, and 20. All data underlying this figure can be found at https://doi.org/10.25452/figshare.plus.23903202.</p

Red light evoked reversal responses are all-trans retinal dependent, as expected.

Animals that express chrimson in the touch receptor neurons were grown in the presence or absence of the necessary co-factor all-trans retinal (ATR) and exposed to 80 μW/mm2 intensity red light. The 95% confidence intervals for population proportions are reported. Two sample Z-test was used to calculate significance; *** indicates pp value is listed in S1 Table. The number of stimulus events for each condition (from left bar to right bar) are: 6,002 and 876. The number of assay plates for ATR + and ATR—conditions are 29, 4. Note that the ATR + condition was previously reported in [19] and also appears in Fig 1C and 1D. The ATR—condition was recorded contemporaneously, but is presented here for the first time. All data underlying this figure can be found at https://doi.org/10.25452/figshare.plus.23903202. (EPS)</p

Endogenous blue light sensitivity and baseline locomotion activity of strains used.

(A) To characterize endogenous sensitivity to blue light, blue light-evoked reversal probability is measured for different strains with and without the all-trans retinal (ATR) co-factor needed for optogenetic proteins. The 300 μW/mm2 blue light intensity used here, is less than that reported to evoke the animal’s endogenous blue light response [64]. Only those strains that express ChR2 are measured on retinal (ATR+, left, N = 2,612, 883, 880 from left to right, same as Fig 2B), while all strains, including the Chrimosn strains, are measured in the off-retinal condition (ATR-, right, N = 6,564, 3,213, 3,365, 3,867, 7,006, 993, 4,516, 3,324, 646, and 6,470 from left to right). Error bars show 95% confidence intervals for population proportions. We include a lite-1 mutant and wild-type N2 for comparison because our transgenic strains include a mix of both wild-type and lite-1 backgrounds. (B) Average speed of each strains used in this work are shown N = 1,654, 564, 1,065, 654, 983, 1,099, 1,251, 837, 1,706, and 1,952 from left to right. All data underlying this figure can be found at https://doi.org/10.25452/figshare.plus.23903202. (EPS)</p

List of optogenetic measurements performed during behavior.

List of optogenetic measurements performed during behavior.</p

Example showing behavior of a population of animals during an experiment from [19].

Middle 24 s of a 30-min recording is shown. Optogenetic stimulation is delivered in closed loop when turning of an individual animal is detected. Each yellow numbered “x” represents a tracked animal, with its track shown in yellow. Inset at top left shows detailed movements of worm number 213, denoted by a green square. The head of the worm is represented by green dot. A centerline is drawn through the worm’s body and is shown in green. The dynamic circular pattern of green and white spots in the center of the video is a visual timestamp system projected onto the plate that is used for synchronizing the timing of video analysis, as described in [19]. (MP4)</p

Supplementary text detailing the additional control experiments to show that blue light alone cannot restore the mechanosensory evoked reversals.

Supplementary text detailing the additional control experiments to show that blue light alone cannot restore the mechanosensory evoked reversals.</p

Example of a worm reversing in response to optogenetic stimulation of its gentle-touch mechanosensory neurons delivered during forward locomotion.

Recording is from [19]. Animals express Chrimson in gentle-touch mechanosensory neurons (strain name: AML67). Stimulus was delivered in open loop. Green dot denotes the animal’s head. Green line denotes its centerline. Yellow line shows the trajectory of a point midway along the animal’s centerline over the past 10 s. Red indicates area illuminated by red light. (MP4)</p