Skeletal muscle atrophy is often debilitating. In women the decline in lean muscle mass accelerates at the onset of menopause due in part to lower estrogen (E2) levels. We evaluated the role of E2 to regulate skeletal muscle mass and maintain contractile function over 14 days of hindlimb unloading (HU) and 3 days of reloading (REC) in E2 replete (estrogen receptor‐α knock‐out, ERαKO) and E2 deficient (aromatase knockout, ARKO) mice, compared to wild type (WT) animals. Mice were separated into 3 groups: 1) CON: ambulatory controls, 2) HU: 14days of HU, 3) REC: 14 days of HU + 3 days REC. We observed differential effects on body weight (BW) after HU and REC. BW significantly decreased in ARKO mice (−10.9%) and ERαKO (−10.5%) after HU, while BW of WT mice was not different. Neither ARKO nor ERαKO recovered to pre‐HU BW values following REC. Expected declines in muscle mass of soleus (SOL), plantaris (PLAN), gastrocnemius (GAS), and tibialis anterior (TA) in WT mice after HU (Table 1) were observed. ARKO mice trended towards losing more muscle mass during HU and recovered slower than WT. Percent atrophy of muscle mass loss in HU & REC compared to CON. WT SOL PLAN GAS TA HU −38 −20 −15 −11 REC −18 −6 −6 −2 ARKO SOL PLAN GAS TA HU −43 −30 −23 −12 REC −34 −17 −19 −15 ERαKO SOL PLAN GAS TA HU −46 −9 −15 −4 REC −25 −7 −7 −8 The ERαKO muscle mass profile resembled that of WT mice, suggesting that the presence of E2 rather than the absence of ERα mediates estrogen effects during HU and REC. These data suggest that estrogen deficiency slows skeletal muscle mass recovery. Support: HD‐058834 & University of MO Life Sciences Fellowship.