Impact of Circulating Estrogen and Mild Systemic Dehydration on Ultrasonic Vocalizations in Female Rats

Purpose: Hormonal and hydration states are known to individually influence laryngeal physiology, yet their distinct and combined effects on vocal function remain poorly understood. This study investigated the interaction of reduced circulating estrogen, estradiol replacement, and mild systemic dehydration (DEHY) on ultrasonic vocalization (USV) in adult female rats. Method: Forty-eight Long–Evans rats were assigned to six groups ( n = 8 per group) that differed in hormonal state (control, ovariectomized + estradiol pellet OVX + Estradiol, OVX + placebo pellet) and hydration status (euhydration EUHY, DEHY). USVs were recorded during a food-reward elicitation paradigm before hormonal (pellet) replacement, after hormonal replacement, and after 5 days of EUHY or DEHY. Dependent measures included USV number, duration, frequency range, and maximum power. Physiological measures (body weight, water intake, kidney renin expression, uterine weight) supported experimental manipulations. Results: OVX animals produced shorter duration and lower power USVs compared with controls, although total number and frequency range were largely preserved. Even after hormonal replacement, OVX + Estradiol animals emitted fewer USVs with shorter duration, narrower frequency range, and reduced power as compared with control animals. DEHY narrowed USV frequency range across hormonal groups and interacted with hormonal status to modulate USV number and duration. Conclusions: Both circulating estrogen and systemic hydration status selectively influenced USV profiles. Although statistically significant, USV changes were generally of small magnitude, warranting caution in interpretation. These findings highlight the nuanced and context-dependent nature of endocrine and hydration influences on vocalizations and establish a preclinical framework for investigating hormone–hydration interactions in laryngeal function.