Objective This study aimed to investigate the effects of sleep interruption (SI) cycles on emotional behavior in ICR mice, and to establish a mouse model of comorbid anxiety and depression induced by SI. Methods Seventy-two male ICR mice (4 ~ 5 weeks old) were divided randomly into a blank group and a model group. Mice in the model group were subjected to SI stress modeling for 1, 2, and 3 weeks, respectively. After modeling, emotional behaviors were evaluated using open-field, elevated plus maze, light-dark box, marble-burying, and forced-swimming tests. Serum corticosterone levels were detected by enzyme-linked immunosorbent assay. Results Mice in the model group buried significantly more marbles after 1 week of SI stress, compared with the blank group (P<0.05). After 2 weeks of stress, mice in the model group also showed a significant decrease in the number of crossings in the light-dark box (P<0.05) and a significant increase in the number of marbles buried (P<0.01) compared with the control group. After 3 weeks of stress, mice in the model group showed a significant increase in the number of marbles buried (P<0.05), a significant decrease in the number of crossings in the light dark box (P<0.05), and a significant increase in immobility time in the forced-swim test (P<0.01). Conclusions ICR mice exhibited significant anxiety-related behaviors after 2 weeks of SI modeling and significant anxiety- and depressive-related behavioral changes after 3 weeks. Three weeks of SI stress can be used to establish a model of comorbid anxiety and depression.