Learning to identify and appropriately react to stimuli that predict threat is crucial for survival. Pavlovian fear conditioning is widely used to investigate the neuronal mechanisms of threat learning. In this study, we used fibre photometry calcium imaging in freely moving male and female mice to determine activity dynamics in nucleus accumbens (NAc), medial prefrontal cortex (mPFC), and ventral hippocampus (vHip) during a Pavlovian fear conditioning task in which one auditory cue was paired with foot shock (CS+) and another remained unreinforced (CS-). Both cues elicited increased activity across regions, with larger increases to CS+ than CS- as animals learned to discriminate between threat and non-threat cues. While the CS+ increase was transient in mPFC and NAc, it was sustained across the duration of the cue in vHip. Further, the auditory properties of the cues profoundly influenced behavioural learning and the dynamics of neuronal encoding. Behavioural discrimination between CS+ and CS- emerged earlier and was more robust when CS+ was a clicker and CS- was white noise than vice versa. Surprisingly, the opposite relationship was observed in neural encoding: discrimination between CS+ and CS- emerged earlier, was more robust, and endured under extinction conditions when CS+ was white noise and CS- was a clicker. These findings have broad implications for the interpretation of correlated changes in neuronal activity and behaviour, and highlight the importance of cue parameters in conditioning tasks.