Fear conditioning

Fear conditioning and memory retrieval were performed as described in (Cummings and Clem 2020). Briefly, mice underwent fear conditioning consisting of six pairings of an auditory tone (2 kHz, 90 dB, 20 s) with a co-terminating foot shock (0.7 mA, 2 s). The test of fear memory retrieval consisted of four tone presentations in a different context (Context B), and three laser light presentations (473 nm; 20 Hz, 10 ms pulses; 5–8 mW for 20 s). Two of the tone presentations were paired with laser light in an alternating fashion. For optogenetic stimulation during fear conditioning, we administered laser light (473 nm; 20 Hz, 10 ms pulses; 5–8 mW for 20 s) overlapping with all CS-US pairings during conditioning to assess whether optogenetic manipulations disrupt fear learning. To measure retention of fear learning, we examined the response to 2 CS presentations in the absence of laser light stimulation. Freezing and fictive eating was manually scored by a trained observer blind to experimental conditions from videos recorded using Video Freeze^® software (Med Associates, Fairfax, VT, USA).