To investigate hypoxia effects on leucine‐enkephalin (L‐ENK) levels of median eminence (ME) of hypothalamus in rats, and the possible glucocorticoid involvement in modulation. Hypoxia was stimulated in a hypobaric chamber. L‐ENK levels were measured by specific radioimmunoassay during acute, and a comparison of L‐ENK alteration was taken for bilateral adrenalectomy (ADX) with or without a replacement of dexamethasone (DEX). Acute hypoxia 10.8% O2 for 30 min and 2 h markedly enhanced L‐ENK levels of ME from 23.99 ± 7.25 in control to 51.26 ± 16.96 (P <0.01), 53.29 ± 26.10 ng/mg protein (p <0.01), and acute hypoxia at 8.2% O2 significantly increased L‐ENK of ME to 36.76 ± 15.25 (p <0.05) and 32.09 ± 3.58 ng/mg protein (p <0.05). The increased L‐ENK was returned to normoxic level when hypoxia (10.8% O2) exposure lasted for 24 h. After ADX, 10.8% O2 hypoxia induced a sharp decline of L‐ENK in the ME, but this decline was completely reversed by treatment with DEX (500 μg/rat, i.p.). Acute hypoxia increases L‐ENK level of hypothalamic ME in rats, which may present a reduced and /or inhibited release of L‐ENK, acting through a fast negative feedback mechanism of acute hypoxia activated high circulating glucocorticoid level.