The relationship between concentrations of vasopressin in plasma and cerebrospinal fluid (CSF) was examined under basal and hypoxic conditions in unanesthetized and unrestrained adult female sheep. Under basal conditions mean (± SE) concentration of vasopressin (4.1 ± 0.3 pg/ml) in CSF was significantly (p< 0.001) higher than the concentration in arterial plasma (2.4 + 0.2 pg/ml). There was a significant positive correlation between CSF and plasma vasopressin concentrations (r = 0.66, p < 0.01). The introduction of severe hypoxia by exposure of the sheep to an inspired gas mixture of 5% O₂ in N₂, a known stimulus to systemic release of vasopressin, was associated with an increase in the concentration of vasopressin in plasma to 339 ± 65 pg/ml and CSF to 19 ± 3.9 pg/ml. This increase of vasopressin in CSF was delayed in time with respect to the increases in the plasma and of a lesser magnitude. With moderate hypoxia (10% O₂ in N₂ exposure) no increase in CSF or plasma vasopressin concentration was observed, suggesting a threshold response. Intravenous infusion of vasopressin to achieve plasma values comparable to those seen with hypoxia was not accompanied by a significant increase in CSF vasopressin concentration. Thus, severe hypoxia is a potent stimulus for release of vasopressin into both CSF and plasma. Furthermore, data suggest that vasopressin may be released into CSF via a separate route from that released into plasma.