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UltraSoundGate

Manufactured by Avisoft
Sourced in Germany

The UltraSoundGate is a high-quality data acquisition system designed for recording and analyzing ultrasonic signals. It provides precise and reliable hardware for the capture and processing of acoustic data.

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11 protocols using UltraSoundGate

Adult 16p11.2 +/+ and B6 female mice in estrus were used as stimulus mice for adult male 16p11.2 deletion mice. Each female mouse was used as a partner for a maximum of three male subject mice. Each trial was run in a clean cage (36.9 × 15.6 × 13.2 cm3) with an 0.5 cm layer of bedding under dim red lighting in a sound-attenuating chamber. One female stimulus mouse and one male subject mouse were placed in the cage and behaviors were recorded with a video camera for 5 min. Interactions were scored from videos by human observers blind to genotype and treatment, using the Noldus Observer XT 8.0 event recording software. Social parameters scored in the male subjects included nose-to-nose sniff (sniffing snout region of partner), nose-to-anogenital sniff (sniffing anogenital region of partner), and follow (walking closely behind partner). Ultrasonic vocalizations were recorded during the 5 min session using an ultrasonic microphone (Avisoft UltraSoundGate) inside the sound attenuating chamber and subsequently scored using Avisoft SASLab Pro.
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We recorded ultrasonic vocalizations from three pairs of rats, which did not participate in the discrimination training or tests. As soon as each pair was placed in a sound-attenuated chamber (interior: 55 cm (L) × 30 cm (W) × 35 cm (H)), they started to emit PC with rough-and-tumble play2 (link) (Fig. 1a). Two out of three pairs continued the inter-male confrontation phase, and then the socially defeated male emitted DC5 (link) (Fig. 1a). Their vocalizations were recorded via an ultrasonic microphone (UltraSoundGate, Avisoft, Glienicke, Germany) placed in the chamber, and digitized by recording software (Recorder-USGH, Avisoft) at a 250-kHz sampling with 16-bit resolution. These vocalizations were recorded within 30 minutes.
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We used two types of microphones (Type 4158 N, Aco, Tokyo, Japan: CM16, Avisoft Bioacoustics, Berlin, Germany) and an A/D converter (Ultra-SoundGate, Avisoft Bioacoustics, SpectoLibellus 2D, Katou Acoustics Consultant Office, Kanagawa, Japan), which exhibit similar performance in recording USVs. None of the sets of microphones and converters caused any difference in USV analysis. The microphone was placed approximately 20 cm from the rats (see section "Recording of USVs during each condition).
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Experiments were conducted in a modular operant conditioning cage with a grid floor, pellet receptacle, and pellet dispenser (Behavioral Test Packages for Rat, Med Associates Inc., Fairfax, VT, USA) controlled by Med-PC® V software (version 5.08, Med Associates Inc.). Additionally, there was a USV condenser microphone CM16/CMPA (UltraSoundGate, Avisoft Bioacoustics, Glienicke/Nordbahn, Germany) placed in the glass ceiling of the cage and an ultrasonic speaker (Vifa, Avisoft Bioacoustics) placed in the corner of the cage, next to the pellet receptacle, connected to an UltraSoundGate Player 116 (Avisoft Bioacoustics). Both USV-playback and recording were performed using Avisoft Recorder USGH software (version 1.0.0.1, Avisoft Bioacoustics). The locomotor activity was recorded with a FLIR® camera (Teledyne FLIR LCC, Wilsonville, OR, USA) mounted behind or above the cage and controlled via Spinnaker® SDK software (version 1.15.0.63, Teledyne FLIR LCC, Wilsonville, OR, USA); home-cage activity was recorded with a Basler camera (acA1300-60gc, Basler AG, Ahrensburg, Germany) controlled via EthoVision XT software (version 10, Noldus, Wageningen, Netherlands). Non-flavored sucrose pellets weighing 45 mg (TestDiet®, St. Louis, MO, USA) were used as rewards in conditioning. The cage was cleaned and thoroughly wiped using 10% EtOH between animals.
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On the 18th day of the experiment, the test was carried out in a specially constructed cage (62 × 48 × 25 cm) made of transparent plastic (details in [50 (link)]). A perforated, transparent plexiglass divided the interior into two halves—one intended for the observer and the other for the demonstrator. Metal rods installed in the demonstrators’ section were connected to the current generator (MedAssociates), which allowed for the administration of mild electrical shocks. The rats could see, hear, and smell each other throughout the test. During the test, the animals were recorded with a digital camera and with 4 microphones (UltraSoundGate, Avisoft). The test session and the pre-exposure to shocks were conducted in separate rooms. To provide a different context, the lights were dimmed, and the cage was cleaned with a 1% acetone solution after testing each pair. Two minutes after inserting the rats into the appropriate chambers, the demonstrator was given a series of 10 foot shocks (1 s, 1.0 mA). In the ‘warned’ group, each electric shock was signaled by a 19 s, 1.75 kHz sound, equaling a total of 80 s between 2 shocks, whereas in the ‘unwarned’ group, there was no sound indication and the shocks were administered in a 1 min interval (Figure 1 and Figure 3).
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Heterospecific rough-and-tumble play was conducted as previously described [16 (link), 20 (link)]. Animals received three consecutive days of light-touch habituation, which does not induce ultrasonic vocalizations (USVs), before testing [20 (link)]. These animals did not receive sleep-EEG surgeries or testing. Briefly, heterospecific rough-and-tumble play stimulation was administered by the experimenter’s right hand. Rats received 3 minutes of play consisting of alternating 15-second blocks of play and 15 seconds of no stimulation. The experimenter was blind to the treatment condition of the animals. At the end of the 3-minute session, running speed (cm/second) for the animal to traverse a 57-centimeter arena and touch the experimenters’ hand to self-administer play was measured manually with a digital stopwatch. High-frequency USVs were recorded (see below; Avisoft UltraSoundGate, Germany) during the 6 × 15-second no-stimulation blocks and analyzed by sonogram (Avisoft SASlab Pro, Germany) in a blinded manner, as described previously [18 (link)]. Rats were dosed with NYX-2925 (1 mg/kg PO; Sai Advantium, India) in CMC or CMC vehicle 1 hour before testing. Previously it has been shown that positive modulation of the NMDAR with d-cycloserine has been shown to rescue sleep deprivation-induced deficits in learning [21 (link)].
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Prior to the commencement of SA recording sessions, a condenser microphone (CM16/CMPA, Avisoft) was suspended 2.5 cm above an arrangement of small holes in the top of the Plexiglas SA chamber. USVs were recorded at a 250 kHz sampling frequency (16 bits) using recording software (Ultrasound Gate, Avisoft, Glienicke/Nordbahn, Germany). Baseline USVs were recorded one week after surgery prior to the start of the first SA session over the same 6 h period as SA sessions were conducted. Subsequent USV recordings were obtained for 6 h during session 1 (T01), session 30 (T30), and the first day of abstinence (ABS; session 31) at the same time of day as a 6 h SA session. As characterization and scoring of USVs are time- and labor-intensive, this limited agenda was designed to capture affective calling during SA for the first time, SA for the 30th consecutive day, and the first time being deprived of the expected drug (18 h withdrawal).
Audio files were run through an automated detector, DeepSqueak [25 (link)], to isolate potential calls. These were then manually checked to distinguish between actual calls (which were accepted) and artifacts and background noise (which were rejected). The automatic detector outputs the specific frequency and exact timing of individual calls. Only manually accepted calls were used for analyses. Calls were designated as belonging to the 22 or 50 kHz ranges.
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One day after the conditioning, the rats were transferred into individual experimental cages, identical to home cages (plastic; 37 × 25 × 16 cm), and transported to the experimental room, where under white light, in the absence of the experimenter and other rats in the room, acoustic stimuli were presented through an ultrasonic speaker (Vifa, Avisoft Bioacoustics, Berlin, Germany), placed just above the shorter side of the cage, connected to an UltraSoundGate Player 116 (Avisoft Bioacoustics). USV emitted by the rat were recorded by a CM16/CMPA condenser microphone (UltraSoundGate, Avisoft Bioacoustics) placed 33 cm above the center of the cage floor, 20 cm away from the speaker. In this configuration, calls from the speaker were still visible in the recording (monitoring of playback), but they were distinctively weaker than USV emitted from the cage. Both playback and recording were performed using Avisoft Recorder USGH software (Avisoft Bioacoustics). The locomotor activity of the animal was recorded with a camera (acA1300-60gc, Basler AG, Ahrensburg, Germany) mounted above the cage and EthoVision XT software (version 10, Noldus, Wageningen, The Netherlands). Signals from radiotelemetric transmitters were collected by receivers located under the cage floor and then recorded by Ponemah software (version 6.32, Data Sciences International, St. Paul, MN, USA).
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9

Isolation-Induced Ultrasonic Vocalizations in Mouse Pups

To test isolation calls, mouse pups were removed from the nest, four at a time, and individually placed into sound attenuation chambers for recording. These chambers were constructed from small coolers (Coleman) that were coated inside with soundproof foam (Soundcoat). An ultrasonic microphone (UltraSoundGate, Avisoft Bioacoustics) was suspended above the pup. Recordings were conducted at P7 and P14 for a total of 15 minutes at each time point. In order to not affect calling patterns of any remaining pups in a given litter, only four (the total number of recording chambers) from each litter were recorded. After recording at P7, pups were tailed for genotyping, and tattooed to enable identification for re-recording at P14. The initial distance between the microphones and the pups was equivalent across chambers at P7 when pups are fairly immobile. At P14, pups are ambulatory so their distance from the microphone varied. Because of this, amplitude measurements were not included in the acoustic analysis. Pups were recorded within the same 2-hour time window each day (light: 14:00–16:00 hr) to avoid circadian effects. Temperature was maintained at 21–22 °C.
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For testing the segmentation performance of our procedure, we prepared datasets consisting of recorded sounds and manually detected onset/offset timing of syllables for three species in the rodent superfamily (Table 1). The manual segmentation for each species was performed by a different human expert. These experts segmented sound materials by visual inspection of a spectrogram, independently of any automatic segmentation system. They were not informed about any of the results of our procedure beforehand. Finally, we collected segmented data for each condition (species, strains, or contexts) as described below. For all species/strain/context conditions, ultrasonic sounds were recorded using a commercial condenser microphone and an A/D converter (Ultra-SoundGate, Avisoft Bioacoustics, Berlin, Germany; SpectoLibellus2D, Katou Acoustics Consultant Office, Kanagawa, Japan). All data were resampled at 250 kHz to have the same sampling rates before starting performance tests for consistency across datasets, though our procedure can be applied to data with much higher sampling rates. The whole dataset is available online (https://doi.org/10.5281/zenodo.3428024).
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