DOI: 10.5061/dryad.tb2rbp09r
title: Data from: Anthropogenic noise exposure over development increases baseline auditory activity and decision-making time in adult crickets

description: Anthropogenic noise negatively affects some animals more so than others
 and often in different ways. Female crickets reared in traffic noise are
 reported to be faster or slower to locate male song than those reared in
 silence depending on species. No study has considered whether observed
 differences were due to hearing or decision-making. We reared female
 Teleogryllus oceanicus in traffic noise and silence, and adult females
 located male song broadcast amidst traffic noise or silence. We recorded
 activity of two auditory interneurons in a subset of individuals under
 identical acoustic conditions. Regardless of rearing treatment, crickets
 were slower to leave their shelter when presented with male song in
 silence than in traffic noise, while crickets reared in traffic noise were
 also slower to leave overall. Crickets reared in traffic noise also had
 higher baseline AN2 activity, but rearing condition did not affect hearing
 thresholds or auditory response to male song. Our results demonstrate
 behavioural and auditory effects of long-term exposure to anthropogenic
 noise. Further, they support the idea that silence itself is a potentially
 aversive acoustic condition.
description: <strong><em>Behaviour 
 </em></strong> To compare the effect of the
 two rearing treatments on female phonotaxis, we randomly assigned crickets
 at fourteen days after final moult to a playback condition of either a
 traffic noise or silence (speaker on but no sound) during male song
 playback. In the center of the experimental trial room (H: 2.3 m, W: 2.6
 m, L: 4.6m), we placed a circular arena made of cork (dia.: 1.22 m, 6.25
 mm thick) on the floor, the arena was marked with a circle of 1.2 m
 diameter, and the circle divided into 8 segments of equal arc (45 degrees;
 Fig. 1B). We randomly assigned two focal speakers (Ultrasonic Dynamic
 Speakers, Vifa, Avisoft, Berlin) to opposite sides of the arena for each
 trial, positioned on the dividing line between two segments. Each focal
 speaker was flanked by two paired speakers (Companion, Bose) 25 cm to
 either side of it along the circumference of the arena. All speakers faced
 the center of the arena. For each trial, we placed an
 adult female cricket under an egg carton cup in the center of the arena
 and then covered the cup with a darkened plastic container to constrain
 the cricket. The four flanking speakers played either a 3-minute repeating
 segment of traffic noise, selected for its low variability in amplitude
 (68-78 dB SPL at the center of arena), or a silent sound file. In both
 acoustic treatments, we randomly selected one of the focal speakers to
 play the modified calling song of <em>T. oceanicus</em> (70 dB
 SPL at center of arena). The crickets were given 60 seconds to acclimatise
 to their physical and acoustic environment before the plastic container
 was removed, marking the beginning of the trial. All trials were conducted
 at ~21°C. We recorded all trials at 15 frames per
 second using a camera (Analog CCTV Camera HD 1080P 4-in-1 Security Dome
 Camera) mounted 2.45 m overhead. Each trial lasted five minutes and was
 considered complete if the cricket reached the edge of the circle or
 failed to reach it by the five-minute mark. We first scored crickets based
 on whether they successfully reached the focal speaker playing the male
 song (i.e., exiting from one of the two segments on either side of the
 focal speaker). For crickets that exited the arena, we determined how long
 they took to (a) emerge from the egg carton shelter (start latency) and
 (b) to reach the edge of the arena (time to finish). We then calculated
 (c) search time, by subtracting (a) start latency from (b) time to finish.
 We weighed each cricket after the trial was finished to control for
 potential size-based variation in statistical analyses.
 <strong><em>Auditory
 activity</em></strong> We recorded neural
 responses to conspecific song under the same acoustic conditions used in
 the phonotaxis experiments. We first randomly selected 20 crickets used in
 the phonotaxis experiment, 10 of which had been reared under traffic noise
 and 10 under silence. To prepare crickets for neurophysiological
 recordings, we pinned them ventral side up to a block of modelling clay
 and removed the cuticle on the ventral surface between the head and
 thorax. This exposed the cervical connectives that contain the axons of
 the AN1 and AN2 auditory interneurons. We draped one of the cervical
 connectives over a hook-shaped tungsten recording electrode and placed a
 reference electrode in the abdomen. Electrodes were connected to a
 differential amplifier (model DP-301, Warner Instruments, USA) and the
 output was passed to a data acquisition board (UltrasoundGate 816H,
 Avisoft Bioacoustics) for digital recording on a computer.
 Neurophysiological recordings were made in one of the acoustic
 chambers. We used the same arenas used for phonotaxis trials, the only
 exceptions being a lack of shelter in the center and only one pair of
 flanking speakers surrounding one focal speaker. We also placed a
 microphone (CM16, Avisoft Bioacoustics ) one meter from the center of the
 arena opposite the set of speakers. The microphone was connected to a
 different channel of the same data acquisition board as the differential
 amplifier. We placed the cricket neural preparation in the center of this
 arena, with the ear ipsilateral to the recorded connective facing towards,
 and 0.6 m away from, the focal speaker. To measure AN2
 thresholds and AN1 activity across frequencies (i.e. produce audiograms),
 we played increasingly loud sound pulses from 2-30 kHz at 2 kHz intervals
 from the focal speaker. For each frequency, sound pulses increased in
 amplitude from 30-80 dB SPL in 2 dB steps. Sound pulses were 20 ms in
 duration (including 1 ms rise/fall times) and broadcast at 500 ms
 intervals to avoid neural adaptation. Audiograms were repeated twice, in
 silence and with traffic noise from the flanking speakers calibrated to be
 70 dB SPL at the cricket. To measure interneuron
 activity in response to cricket song under the conditions crickets
 experienced during the phonotaxis experiment, we played <em>T.
 oceanicus</em> song from the focal speaker to the cricket neural
 preparations under two noise conditions: silence or traffic noise from the
 flanking speakers calibrated to be 70 dB SPL at the cricket. We first
 recorded neural activity in the background noise condition (silence or
 traffic noise) for one minute to establish a baseline response to the
 noise condition. After a minute, we began playback of the optimized
 <em>T. oceanicus</em> song from the focal speaker alongside
 the background noise condition for another minute.  Crickets were exposed
 to the two noise conditions in a random order.
description: # Data from: Anthropogenic noise exposure over development increases
 baseline auditory activity and decision-making time in adult crickets
 [https://doi.org/10.5061/dryad.tb2rbp09r](https://doi.org/10.5061/dryad.tb2rbp09r) Dataset consists of behavioural and neuro-physiological data collected over the 2021 and 2022, using the cricket Teleogryllus oceanicus.  ## Description of the data and file structure The data consists of several files.  **2021_2022_T.oceanicus_experiment**: This file contains the behavioural data. It is organized with each row being an individual cricket, each given a letter + number code that indicates which rearing treatment they were in and the order they were put into the rearing chamber. Missing data represents values that were not recorded for various reasons (i.e. died before testing, pilot runs of the experiment, video file corruption, equipment failure). Signal location represents which position on the circular arena the mating song was being played from (circular arena divided into 8 pie segments, with the speaker being placed on one of the numbered edges between two segments). Exit location lists the segment which the cricket exited the arena from, described using the two edges that border it. File names refer to the .txt files that contain the coordinates of the cricket from the video recording of it, outputted by Ctrax. File names were manually matched to the cricket entries by using the date and time in the file name itself.  **T_oceanicus_video_coord_2021.zip + T_oceanicus_video_coord_2022_exp_1.zip:** These folders contain the CSV files with the coordinates for every frame of each video, separated by which year the recordings were taken. These files were produced by Ctrax, a video processing software that inputs video files and returns coordinates ([https://ctrax.sourceforge.net/](https://ctrax.sourceforge.net/)). They were further processed by a data cleaning program for Matlab, available on the Ctrax website. Additional files are contained in these folders that are not used in the data set. Recordings were processed blind so several outtakes and scrapped trials were processed and still included in the folders.  **TO_song_data_20230410.csv:** The number of auditory neuron (AN2) spikes/firings per recording. Each row lists what cricket and noise were used and then what point in the recording the measurement was taken. Spikes were measured for 1 cricket song length at three points in the recording (baseline, song start and 30 seconds with song) to get a measure of background firing rate, how the neuron responds to song and acclimation to song. **TO_Audiograms_20230410.csv:** The volume at which the auditory neuron (AN2) first fires under the given condition (frequency of sound test sound pulse, type of background noise). Listed as threshold and measured in dB.SPL. Number of spikes at first response and the time gap between the sound arriving and spikes occuring (Latency) are also given. Each row represents a single frequency of sound that was presented to the cricket in a given recording. Missing data code: NA **TO_AN1_pulse_times_Audiogram_Erik.csv:** Time data of when each of the sound pulses occurred in each of the audiogram trials. Each row gives the time, frequency and sound pressure level of each sound pulse. Empty cells filled in as part of a later script. Infilling these cells will interfere with a script meant to analyze the file.  **TO_AN1_rms_measurements.csv:** Final root mean squared measurements of all the neural traces with the AN2 spikes removed. Organized identically to TO_AN1_pulse_times_Audiogram_Erik.csv. Additionally includes root mean squared values of the neural trace before and after the sound stimulus. **TO_AN1_rms_measurements_Hannah.csv:** A subset of TO_AN1_rms_measurements.csv that was calculated separately by H.M.H. and added to the main dataset afterward.  **Interspike durations.zip:** Folder containing .txt files with the location of all auditory neuron spikes detected in each file for each cricket. Output from SASlab Pro. ## Code/Software **T.ocean_exp_code.R:** R code for analyzing the behavioural data, including the video processing. Required packages are specified in the library list at the beginning of the script. Script includes code for both "Anthropogenic noise exposure over development increases baseline auditory activity and decision-making time in adult crickets" and a future study (listed as 2021+2022 and 2022 respectively). Contains code for loading the data, calculating certain variables such as speed, distance travelled and search time, linear models for calculating the effects, and plots.  **TO spike analysis.R:** R code for analyzing the neural data. Required packages are specified in the library list at the beginning of the script. Script includes code for both "Anthropogenic noise exposure over development increases baseline auditory activity and decision-making time in adult crickets" and a future study (Second study listed as "noise comparison" and/or "river noise paper"). Contains code for loading and assembling the data sets from the .csv and .txt files, doing repeated measure ANOVA's, and plots.  **TO AN1 amplitude measurements_Erik.R:** R code for calculating root mean square values of neural trace before and after the onset of a signal, using labelled spike counts from the audiograms and song stimulus tests. Takes TO_AN1_pulse_times_Audiogram_Erik.csv as input and produces TO_AN1_rms_measurements.csv.  **TO_AN1_amplitude_analysis_V5.R:** R code for comparing the root mean square values between conditions in TO_AN1_rms_measurements.csv.  Produces pairwise t-test comparisons for the audiograms and song stimulus experiments.  ## Version Changes: **11-Feb-2025:** Added TO AN1 amplitude measurements_Erik.R, TO_AN1_amplitude_analysis_V5.R, TO_AN1_pulse_times_Audiogram_Erik.csv, TO_AN1_rms_measurements.csv, and TO_AN1_rms_measurements_Hannah.csv to the dataset.