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Simple Summary: Psychological stress triggers an imbalance of neural and hormonal systems, leading to pathological conditions in many organ systems, especially the urinary bladder. Water avoidance stress (WAS) exposure for 10 days is a well-known rodent model that induces chronic psychological stress and impairs urinary bladder structure and function. However, the impact of WAS exposure in a different period apart from 10 days on anxiety-related behavior and urinary bladder structure and function remains unknown. Here, we investigated the effect of 1 day (acute), 10 days (chronic), and 28 consecutive days (prolonged) of WAS exposure and determined changes in anxiety-related behaviors, voiding patterns, mast cell numbers in bladder tissues, and bladder contractile properties that reflect pathological conditions from the stress exposure. Our study provides useful information on using WAS in an acute period (1 day) and prolonged period (28 days) on changes in anxiety-related behaviors and urinary bladder functions. The findings from this study are also essential for biomedical researchers to appropriately utilize the period of WAS exposure to investigate the effect of psychological stress-induced changes in the urinary bladders or other internal organs in a shorter or longer period than 10 days of WAS induction in a mouse model. Repeated water avoidance stress (WAS) for 10 days is a common rodent model to mimic the effect of chronic psychological stress on urinary bladder dysfunction. However, it remains obscure whether changes in the stress exposure period impact urinary bladder impairment differently. Therefore, this study aimed to investigate the effect of 1 (acute), 10 (chronic), and 28 (prolonged) days of WAS on anxiety-related behavior, voiding pattern, urinary bladder mast cells, and bladder contractility in C57BL/6J male mice. Mice exposed to 1 and 10 days of WAS showed decreased unsupported rearing. A decreased total void area after 1 and 10 days of the WAS was observed, which was reversed in the 28-day-WAS group. There was an increased number of degranulated mast cells in the bladder of the 10-day-WAS group. The 1-day WAS exposure enhanced tonic contractile response to a muscarinic agonist, carbachol, which was reversed by 5-HT3 receptor antagonist pre-incubation. Interestingly, the 28-day WAS group showed a similar tonic contractile response to the control group. Our findings provide more insightful information about using 1-day WAS as an acute psychological stress model, and stress exposure longer than 10 days did not produce anxiety-like behavior and urinary bladder impairment. [ABSTRACT FROM AUTHOR]