Shared behavioural impairments in visual perception and place avoidance across different autism models are driven by periaqueductal grey hypoexcitability in Setd5 haploinsufficient mice. Authors Laura E. Burnett1, Peter Koppensteiner1, Olga Symonova1, Tomás Masson1, Tomas Vega-Zuniga1, Ximena Contreras1, Thomas Rülicke2, Ryuichi Shigemoto1, Gaia Novarino1, Maximilian Joesch1*. Affiliations 1 Institute of Science and Technology Austria; Am Campus 1, 3400 Klosterneuburg, Austria. 2 Department of Biomedical Sciences and Ludwig Boltzmann Institute for Hematology and Oncology, University of Veterinary Medicine; Veterinärplatz 1, 1210 Vienna, Austria. *Corresponding author. Email: maxjosch@ist.ac.at Compressed zip files include: * Behaviour * DTX_cannula_loom_response * setd5 * Looming_Escape_Response * cul3 * ptchd1 * setd5 * different_contrasts * setd5_emx1 * setd5_males * Optogenetic_stimulation * Setd5 * AAV9_ChR2 * AAV9_GCaMP_controls * PatchClamp * cul3 * ptchd1 * setd5 * periaqueductal_grey * a-dendrotoxin * superior_colliculus * SiliconProbe * cul3 * ptchd1 * setd5 * WesternBlot Behaviour Raw behavioural Data will be provided on request due to file sizes. The data uploaded are the MATLAB (.mat) files containing the preprocessed data collated across experimental cohorts. The data files included in these folders are explicitly referred to in the accompanying excel files (D1_data.xlsx, S1_data.xlsx, etc.), together with the appropriate MATLAB code found in the ‘Burnett_et_al_2024’ GitHub repository (https://zenodo.org/doi/10.5281/zenodo.11130587), for each figure panel. The data are organised into folders in the following structure: Within the ‘Behaviour’ folder, in both the ‘DTX_cannula_loom_response’ and ‘Looming_Escape_Response’ folders there are files that contain ‘ALL_XYLOOM’ files and ‘xy_analysis’ in the name that contain processed information of the speed of the animal during looming escape response (LER) trials. Both types of file contain MATLAB tables where each row corresponds to an individual experiment, shown in the ‘Exp’ column of the table. There should be 3 experiments per animal per day (‘01_Loom’, ‘02_Loom’, etc.’). Both tables should contain columns containing information about the ‘Date’, the ‘Animal’ number and the genotype (‘Geno’) of the animal. ‘ALL_XYLOOM…’ files additionally contain the columns ‘Speed’, ‘MaxSp’ and ‘T2M’. * ‘Speed’ contains a cell array of size [1, 780]. This is the speed the animal is moving in every frame from 3s before the loom stimulus begins to 10s after. The video was recorded at 60fps (60*(10+3) = 780) and the speed is in the units cm s-1. * The column ‘MaxSp’ contains a single float value containing the maximum speed reached by the animal between the start of the stimulus and 10s after the start of the stimulus. * The column ‘T2M’ contains the time to the maximum speed in seconds after the start of the stimulus. ‘xy_analysis…’ files contain the additional columns: * ‘Mean_5s’ and ‘Mean_10s’: The mean speed in the 5s or 10s before the start of the looming stimulus, respectively. * ‘DShelt_Start’: The distance of the animal from the edge of the shelter at the start of the looming stimulus in cm. * ‘ReturnToShelter’: Binary value for if the mouse returns to the shelter within 7s of the start of the stimulus. * ‘SpeedToShelter’: the average speed of the mouse (in cm s-1) from the start of the stimulus until either the mouse re-enters the shelter or the end of the 10s after the start of the stimulus. * ‘MaxSpEscape’: the maximum speed in cm s-1 reached by the mouse between the start of the looming stimulus and until either the mouse re-enters the shelter or the end of the 10s after the start of the stimulus. * ‘TimeToMaxSp’: the time to reach this maximum speed value in seconds. There are also files that contain ‘EXITS’ and ‘exit_analysis’ in the name, that contain processed information about the shelter exits of the animals during the experiments. Similarly to ‘xy_analysis’, ‘exit_analysis’ files contain tables where each row corresponds to an individual experiment, again shown in the ‘Exp’ column. ‘exit_analysis…’ files contain the additional columns: * ‘Date’: the date of the experiment (YYMMDD). * ‘data’: for a-DTX experiments this column is ‘1’ for experiments with saline and ‘2’ for experiments with a-DTX. * ‘Animal’: for animal ID number. * ‘X’ : the ‘x’ coordinate position of the animal during every frame of the recorded behavioural video. * ‘Y’: the ‘y’ coordinate position of the animal during every frame of the recorded behavioural video. * ‘Speed’ : the speed of the animal during every frame of the recorded behavioural video in cms-1. * ‘DShelt’: the distance of the animal from the edge of the shelter during every frame of the recorded behavioural video in cm. * ‘NumOut’: the number of shelter exits during the experiment. * ‘NumOutTrig’: the number of shelter exits during the experiment where the mouse passes the trigger threshold. * ‘MeanD’: the mean distance covered per shelter exit across all shelter exits in that experiment (cm). * ‘MaxD’: the maximum distance covered for any shelter exit in that experiment (cm). * ‘InterBoutInterval’: the mean interval (s) between shelter exits for that experiment. * ‘ExitDur’: the mean exit duration across all exits during that experiment (s). * ‘LoomRows’: the frame of the behavioural video when the mouse first leaves the shelter, per exit. * ‘Geno’: the genotype of the animal. 1 = wild-type, 2 = mutant. ‘EXITS’ files contain tables where each row corresponds to an individual exit, not each experiment. These files contain the columns: * ‘Animal’: for the animal ID number. * ‘Date’: the date of the experiment (YYMMDD). * ‘data’: for a-DTX experiments this column is ‘1’ for experiments with saline and ‘2’ for experiments with a-DTX. * ‘Exp’: the experiment number. * ‘ExitNum’: the exit number in chronological ascending order. * ‘LoomTrig’: boolean value, 1 = loom stimulus triggered during exit, 0 = no loom stimulus triggered during exit. * ‘FrameOut’: frame of the behavioural video that the mouse left the shelter. * ‘FrameIn’: frame of the behavioural video that the mouse re-entered the shelter. * ‘FramesIBI’: the total number of frames in the inter-bout interval (IBI). * ‘IBI’: the total time of the interbout interval between this exit and the previous exit (s). * ‘ExitDur’: the total time (s) that the mouse was outside of the shelter. * ‘MaxDistance’: the maximum euclidean distance between the mouse and the centre of the shelter during the exit (cm). * ‘OutSpeed’: the speed of the mouse (cms-1) when leaving the shelter. * ‘Geno’: genotype of the mouse. 1 = wild type, 2 = mutant. For the a-dendrotoxin (a-DTX) cannula experiments, there are additional files (MaxSp_DATA_DTX_Setd5.mat and ReactionTime_DATA_DTX_Setd5.mat) that contain the per animal data values for these two metrics sorted into separate arrays for wild-type and mutant animals with saline (‘valsWT1’, ‘valsHET1’) and after a-DTX application (‘valsWT2’, ‘valsHET2’). Within the ‘Looming_Escape_Response’ folder there are also ‘activity_analysis’ files. These files contain the tables, ‘animal_activity_table’ which contains: * the averaged values across experiments per animal for the speed travelled during the experiments (‘Speed’, cm s-1) * the maximum speed reached during the experiment (‘MaxSpeed’, cm s-1) * the percentage of the experiment that the animal was travelling at <2 cm s-1 (‘PerSlow’, %) * the percentage of the experiment that the animal was outside of the shelter (‘PerOutBox’, %) * the percentage of the experiment that the animal was in the centre of the arena (‘PerCentre’, %) * the percentage of the experiment that the animal was at the edge of the box (‘PerEdge’, %) * the genotype (‘Geno’). There are also tables containing statistical values either by pooling across all experiments per genotype (‘stats_firing_table’) or by pooling across values averaged per animal (‘AA_stats_firing_table’), and ‘ACTIVITY_ARRAY’ files which contain ‘ALL_ACTIVITY_ARRAY’ tables which have one row per experiment and columns containing the values for the metrics described in the ‘animal_activity_table’ above. Files that were used to assess the exploratory behaviour of the animals in the ‘preloom’ period before having been exposed to a single looming stimulus contain the word ‘preloom’ in their name. For assessing the change in the speed of the animals upon visual stimulation with the looming stimulus, there are files with ‘spat_spim’ in the title. These contain arrays with the per animal data points for the average speed across loom trials either ‘at’ the looming stimulus, or immediately after the looming stimulus onset (‘im’). Trials were split based on whether the animal responded to the looming stimulus within the first presentation of a looming stimulus (‘valsWT1at’, ‘valsWT1im’, ‘valsHET1at’, ‘valsHET1im’) or after the first presentation of a looming stimulus (‘valsWT2at’, ‘valsWT2im’, ‘valsHET2at’, ‘valsHET2im’). For the Setd5 animals there are also tables containing information related to the relative position of the animal with respect to the shelter and the arena during the loom presentations. This can be found in the ‘Setd5_Position_Control_Data.mat’ and contains a table ‘FULL_XY_TABLE’ that contains columns with useful data related to the position of the animal during the experiment. Useful columns include: * ‘X’: x position of the animal per frame. * ‘Y’: y position of the animal per frame. * ‘DIST_C’: distance of the animal (cm) from the centre of the arena per frame. * ‘DIST_S’: distance of the animal (cm) from the shelter per frame. * ‘MIN_DIST’: the minimum distance (cm) from the position where the mouse was at the stimulus’ onset and the shelter. * ‘TRUE_DIST’: the true distance travelled by the animal (cm) from the position at stimulus onset until it re-entered the shelter. * ‘DIST_RATIO’: the ratio between ‘MIN_DIST’ and ‘TRUE_DIST’. * ‘ANG’: the angle between the head of the mouse and the shelter at stimulus onset. * ‘RETURN’: boolean value, 1 = returned to the shelter within 7s of stimulus onset. * ‘SpeedATLoom’: instantaneous speed of the animal at the loom’s onset (cm s-1). For Setd5 animals there are also files containing data acquired during the ‘reward’ trials, these have ‘reward’ in their title but are in the same format as the regular loom experiment files. For some experiment types, multiple tables/arrays have been grouped together and saved as one file, for example ‘Setd5_Emx1_Looming_Escape_Data.mat’. This file contains: * ‘all_xy_analysis’: the ‘xy_analysis’ format table mentioned earlier for all trials. * ‘ALL_XYLOOM_TABLE’: the ‘XY_LOOM’ format table mentioned earlier for all trials. * ‘xy_return’: contains the data from ‘all_xy_analysis’ but only for the trials where the mouse performs an escape response. * ‘ALL_XYLOOM’: contains the data from ‘ALL_XYLOOM_TABLE’ but only for the trials where the mouse performs an escape response. * ‘animal_xy_table’: contains the data from ‘xy_return’ averaged across trials per animal. * ‘l2m_array’: each row is an animal (ID number is the number in column 1). Subsequent columns contain the proportion of trials where the mouse escaped within the first, between looms 1-2, between looms 2-3, between looms 3-4, between looms 4-5, after the 5th loom, or not at all. The last column contains the genotype of the animal (1 = wild type, 2 = mutant). For the optogenetics experiments, the files contain a table (‘xy_opto_infoA’) and a matrix (‘xy_optoA’). ‘xy_opto_infoA’ is similar to ‘xy_analysis’ tables in that each row corresponds to one experimental trial. Other columns of interest in ‘xy_opto_infoA’ are: * ‘NumPulses’: the number of light pulses in the optogenetic stimulus. * ‘T_pulse’: the time (ms) of each light pulse in the optogenetic stimulus. * ‘FreqPulse’: the frequency of the light pulses (Hz) in the optogenetic stimulus. * ‘EC’: the value of the ‘electric current’ used. * ‘SpAt’: the speed of the animal (cm s-1) at light onset. * ‘SpImmed’: the speed of the animal (cm s-1) in the period following light onset. ‘xy_optoA’ is a matrix where the rows correspond to the rows in ‘xy_opto_infoA’, but contains the instantaneous speed of the mouse (cm s-1) per frame in each column. The optogenetic stimulation occurs at column 600 and each row array contains the speed data of the animal 10s before the onset of the light stimulation to 10s after. - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - For the PatchClamp data: ‘Spike_Table’ files contain tables where each row corresponds to a single spike and contains columns of interest that include: * ‘MaxAmp’: the maximum amplitude of the spike (mV). * ‘V’: array of recorded voltage (mV) with the peak of the spike occurring at value 1000. * ‘dvdt’: array of recorded change in voltage over time centred on the peak of the spike at value 1000. * ‘APThresh’: action potential threshold (mV) * ‘RiseT’: rise time (ms) * ‘whp’: the width at half the peak (ms) * ‘DecayT’: decay time (ms) * ‘MinAmp’: the afterhyperpolarisation amplitude (mV) ‘SPIKE_DATA’ files contain tables where each row is a cell and it contains the columns ‘ST1’ to ‘ST23’ for current step 1 to current step 23. Current step 1 is 0pA and then each subsequent step increases by 10pA. The values recorded are the number of spikes per step. ‘Patch_variables’ files contain tables where every row is a cell and it contains columns with useful information about the intrinsic properties of the cells, including: * ‘Rin’: input resistance * ‘T’: tau * ‘Cm’: Membrane capacitance * ‘RMP’: resting membrane potential * ‘SpFr’: average spike frequency * ‘SpAmp’: average spike amplitude * ‘SpRise’: average spike rise time * ‘SpDecay’: average spike decay time In these tables, the column ‘data’ refers to the genotype of the animal (1=WT, 2 = HET). - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - For the SiliconProbe data there are files that contain the following tables: ‘Loom_Table’ contains one row for each ‘unit’ and contains the following columns: * ‘Date’: date of the recording (YYMMDD) * ‘Ani’: animal ID number * ‘Exp’: experiment number * ‘Geno’: genotype (1=wild type, 2 = mutant) * ‘ProbeDepth’: the depth (um) of the tip of the probe when recording. The probe was zeroed at the brain surface. * ‘Total spikes’: the total number of spikes recorded during this stimulus * ‘Depth’: depth of the unit (um) * ‘L25’: activity over the presentation of the 5 x 5 loom stimuli (Hz) * ‘L5REP’: average activity during the presentation of the 5 loom stimuli. Averaged across the 5 repetitions. (Hz) * ‘L1ONLY’: average activity across a single loom stimulus, average across the 5 repetitions of only the first loom presented (Hz). * ‘MeanBefore’: average baseline spiking before the start of the stimulus (Hz). * ‘MaxL1’: the maximum firing (Hz) during the presentation of one loom stimulus * ‘T2Max’: the time to reach the maximum firing (ms) ‘Flash_Table’ contains one row for each ‘unit’ and contains many of the same columns as ‘Loom_Table’ but additionally contains: * ‘OFO’: activity (Hz) across on on-off-on light flash stimulus. * ‘SPT’: the spikes per time over the course of the flash stimulus. * ‘SPAV’: the firing to the flash stimulus averaged over the repetitions. * ‘MaxON’: maximum firing (Hz) during the ‘ON’ part of the stimulus. * ‘MaxOFF’: maximum firing (Hz) during the ‘OFF’ part of the stimulus. ‘Loom_Variables’ the rows correspond to the rows in ‘Loom_Table’ files but the columns contain: * ‘Avb4’: the average activity in the time before the stimulus (Hz). * ‘m1’-’m5’: the maximum firing during the first to the 5th loom * ‘maxAV’: the average of the maximum firing across the five loom stimuli. * ‘av1’-’av5’: the average firing across one loom stimulus for the first to the 5th loom. * ‘avAV’: the average of the average firing across a single loom presentation across the 5 loom stimuli. For ‘kmeans’ data there are matrices called ‘data’ that contain the spiking (Hz) to the flash stimulus (columns 1 - 120) for every unit recorded (row), column 121 contains the genotype of the animal (1 = wild type, 2 = mutant) and column 122 contains the k-means cluster that the unit belongs to. For Setd5 animals there is a table ‘FLASH_PUPIL_SPIKES’ that has every experiment run as a row and contains columns such as ‘Pupil_1REP’ that has the averaged pupil diameter size averaged across the 10 flash repetitions and ‘MOVING’ which contains a series of 0s and 1s for every frame of the corresponding behavioural video (1 for when the mouse is moving on the ball and 0 for when it is stationary). - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - For the Western Blot data there is a single MATLAB file, ‘220812_WesterBlot_Kv1-1_Rsults_Ctx_SC_PAG.mat’ that contains the variable ‘data’ that is a matrix of the Kv1.1 expression level data where column 1 corresponds to cortical data, column 2 to superior colliculus data and column 3 to periaqueductal grey data. The first three rows are from the 3 wild type replicates and the bottom three rows to the Setd5+/- replicates.