Mice underwent cued and contextual fear conditioning, as previously described [24 (link)]. Briefly, on conditioning day, mice were allowed to explore the conditioned context, which consisted of a Plexiglas chamber and metal floor grid (model H10-11M; Coulbourn Instruments, Whitehall, PA, USA). After 3 min, 15 s of white noise (80 dB) was presented co-terminating with a 2 s, 0.75 mA foot shock. This noise-shock pairing was repeated twice for a total of 3 shocks with a 30 s interval between shocks. Twenty-four hours later, mice were re-exposed to the conditioning chamber for 5 min and freezing responses were measured using FreezeView Software (Coulbourn Instruments) to test contextual long-term memory. Four hours later, mice were placed in a novel context consisting of a 15 cm open-topped plastic cylinder with bedding on the floor for 3 min followed by re-exposure to the white noise for 3 min, to test hippocampal-independent memory. All data were video recorded using FreezeFrame Video-Based Conditioned Fear System and analyzed by FreezeView Software (Coulbourn Instruments). CFC was performed in all male mice before female mice.
Freezeview software
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FreezeView Software is a tool for the analysis and visualization of freeze-drying and lyophilization data. It provides users with the ability to view, analyze, and interpret the data collected during these processes.
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14 protocols using freezeview software
1
Cued and Contextual Fear Conditioning in Mice
2
Cued and Contextual Fear Conditioning in Mice
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Mice underwent cued and contextual fear conditioning, as previously described [24 (link)]. Briefly, on conditioning day, mice were allowed to explore the conditioned context, which consisted of a Plexiglas chamber and metal floor grid (model H10-11M; Coulbourn Instruments, Whitehall, PA, USA). After 3 min, 15 s of white noise (80 dB) was presented co-terminating with a 2 s, 0.75 mA foot shock. This noise-shock pairing was repeated twice for a total of 3 shocks with a 30 s interval between shocks. Twenty-four hours later, mice were re-exposed to the conditioning chamber for 5 min and freezing responses were measured using FreezeView Software (Coulbourn Instruments) to test contextual long-term memory. Four hours later, mice were placed in a novel context consisting of a 15 cm open-topped plastic cylinder with bedding on the floor for 3 min followed by re-exposure to the white noise for 3 min, to test hippocampal-independent memory. All data were video recorded using FreezeFrame Video-Based Conditioned Fear System and analyzed by FreezeView Software (Coulbourn Instruments). CFC was performed in all male mice before female mice.
3
Cued and Contextual Fear Conditioning in Mice
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Mice underwent cued and contextual fear conditioning, as previously described (Matousek et al., 2012) (link). Brie y, on conditioning day, mice were allowed to explore the conditioned context, which consisted of a Plexiglas chamber and metal oor grid (model H10-11M; Coulbourn Instruments, Whitehall, PA, USA). After 3 min, 15 sec of white noise (80 dB) was presented co-terminating with a 2 s, 0.75 mA foot shock. This noise-shock pairing was repeated twice for a total of 3 shocks with a 30 sec interval between shocks. Twentyfour hours later, mice were re-exposed to the conditioning chamber for 5 min and freezing responses were measured using FreezeView Software (Coulbourn Instruments) to test contextual long-term memory. Four hours later, mice were placed in a novel context consisting of a 15 cm open-topped plastic cylinder with bedding on the oor for 3 min followed by re-exposure to the white noise for 3 min, to test hippocampal-independent memory. All data were video recorded using FreezeFrame Video-Based Conditioned Fear System and analyzed by FreezeView Software (Coulbourn Instruments). CFC was performed in all male mice before female mice.
4
Contextual Fear Conditioning in Mice
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Immediately after the NOR task was finished, mice underwent cued and contextual fear conditioning, as previously described (Matousek et al., 2010 (link)). Briefly, on conditioning day, mice were allowed to explore the conditioning context, which consisted of a Plexiglas chamber and metal floor grid. After 3 min, 15 s of white noise was presented co-terminating with a 2 s, 0.75 mA foot shock. This noise-shock pairing was repeated twice for a total of 3 shocks with a 30 s interval between shocks. Twenty-four hours later, mice were re-exposed to the conditioning chamber for 5 min each to test long-term contextual memory. Four hours later, mice were placed in a novel context consisting of a 15 cm open-topped plastic cylinder with bedding on the floor for 3 min followed by re-exposure to the white noise for 3 min, to test hippocampal-independent memory. All data were video recorded and analyzed by FreezeView Software (Coulbourn Instruments).
5
Fear Conditioning in Male AS and Wild-Type Mice
6
Cue-dependent Fear Extinction Assay
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Cue-dependent fear extinction was tested 24 h after fear conditioning and extinction retention occurred 24 h after fear expression. For extinction, mice were placed in a novel context with a different olfactory cue, lighting and flooring and exposed to 15 or 30 tones (30 s, 6 kHz, 65–70 db) with an inter-trial-interval of 60 s. Freezing was measured using Freeze View software (Coulbourn Instruments Inc., Whitehall, PA).
7
Contextual Fear Conditioning in Mice
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Contextual fear conditioning tests were conducted in the same manner as previously described [36] (link). Briefly, mice were first habituated for 30 minutes in the behavioral suite and then placed in fear conditioning chambers. A training protocol was administered using Freeze Frame software (Coulbourn Instruments), which consists of two instances of a low tone paired with a mild electric foot shock (0.7 mA) in a 3-minute time frame. The amount of time that the mice were immobile, designated as “freezing” in this protocol was recorded by the software. For short-term recall, mice were placed back in the same chamber without a tone or foot shock, to measure contextual fear conditioning 2 hours after testing. For long-term recall, mice were again placed in the same chamber 24 hours after testing. Both outputs were measured using Freeze View software (Coulbourn Instruments) after individually adjusting a baseline freezing threshold for each mouse that was tested.
8
Fear Conditioning in Transgenic Mice
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AS mice and their WT littermates were randomly assigned to either control or p18 siRNA groups and blinded to the examiner. Four weeks after AAV injection, mice were placed in the fear-conditioning chamber (H10-11M-TC, Coulbourn Instruments). The conditioning chamber was cleaned with 10% ethanol to provide a background odor. A ventilation fan provided a background noise at ∼55 dB. After a 2 min exploration period, three tone-footshock pairings separated by 1 min intervals were delivered. The 85 dB 2 kHz tone lasted 30 s and co-terminated with a footshock of 0.75 mA and 2 s. Mice remained in the training chamber for another 30 s before being returned to home cages. Context test was performed 1 day after training in the original conditioning chamber with 5 min recording. On day 3, animals were subjected to cue/tone test in a modified chamber with different texture and color, odor, background noise, and lighting. After 5 min recording, mice were exposed to a tone (85 dB, 2 kHz) for 1 min. Mouse behavior was recorded with the Freezeframe software and data were analyzed using the Freezeview software (Coulbourn Instruments). Motionless bouts lasting more than 1 s were considered as freezing. The percent of time animal froze was calculated and group means with SEM were analyzed.
9
Auditory Fear Conditioning in Mice
10
Cued Fear Conditioning and Extinction
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