Time-Dependent Changes in Auditory Fear Generalization and Its Suppression by Extinction in Mice

Fear responses often generalize from threat cues to perceptually similar stimuli, and excessive generalization is implicated in anxiety-related disorders. Although contextual fear generalization can increase over time, whether auditory fear generalization gradients change after conditioning remains unclear. Here we used an auditory fear-conditioning paradigm in male C57BL/6J mice to examine time-dependent changes in tone generalization and whether CS-only extinction suppresses generalized fear. Mice received five tone-shock pairings at one of four shock intensities (0, 0.25, 0.4, or 1.0mA). Generalization was assessed using the conditioned stimulus (CS; 6kHz) and three lower-frequency generalization stimuli (GSs; GS1 3.8kHz, GS2 3.0kHz, GS3 2.4kHz). To avoid repeated generalization testing before extinction, generalization at 24h and 15 days after conditioning was assessed in independent cohorts. At 24h, the 0.4 and 1.0mA groups showed CS-biased gradients, with higher freezing to the CS than to the more distant GSs. At Day 15, freezing remained high but CS-GS discrimination was reduced; this was most evident in the 0.4mA group, which showed a significantly shallower subject-level slope than the corresponding 24-h group. Two sessions of CS-only extinction reduced freezing to both the CS and untrained GSs, consistent with suppression of generalized auditory fear. These findings suggest that auditory fear generalization can become less stimulus-specific over time and that extinction can suppress generalized fear responses.