It has recently been reported that both physical and psychological stress elevate plasma inter-leukin (IL)-6 levels independently of endotoxemia, tissue damage, or inflammation. However, the mechanism of plasma IL-6 elevation in these models is poorly understood. In the present study, plasma IL-6 levels were measured using the IL-6-dependent murine hybridoma subclone B9 cell line, which is commonly used by other investigators. We first demonstrated that an immobilization (IM) stress, a typical physicopsychological stress, increased plasma IL-6 levels. Then the contribution of the hypothalamic-pituitary-adrenal (HPA) axis and the central and peripheral catecholaminergic systems in IM-induced plasma IL-6 elevation were examined because these mechanisms play important roles in host defense against stress. Blood samples were collected through an indwelling jugular venous catheter before, during, and after IM; the number of samples taken serially from each animal was 12-13. Blood cells were resuspended in a saline solution and injected into the animals through the same catheter after each blood collection in order to prevent loss of blood volume. After initiation of restraint, plasma IL-6 levels significantly increased at 60 min and peaked at 90 min in the animals immobilized for either 30 or 120 min. The peak levels of IM-induced plasma IL-6 in the animals immobilized for 120 min (1,905 ± 414 U/ml) were significantly higher than those in the animals subjected to 30 min IM (837 ± 95 U/ml; p < 0.05). In the hypophysectomized (Hypox) or adrenalectomized (ADX) rats, the peak levels of IM-induced (60 min) plasma IL-6 were considerably higher than in sham-operated rats (Hypox: 33,281 ± 4,983 U/ml at 150 min; ADX: 52,020 ± 19,231 U/ml at 120 min). In addition, in order to determine the possible involvement of endogenous catecholamines in IM-induced plasma IL-6 elevation, 6-hydroxydopamine (6-OHDA) was injected into the lateral cerebral ventricle (i.cv., 100 µg/rat) or jugular vein (i.v., 100 mg/kg) of the rats, and the animals were exposed to IM stress 1 week (i.cv.) or 3 days (i.v.) after injection. Both i.cv. and i.v. injections of 6-OHDA markedly attenuated the plasma IL-6 response to IM as compared to the respective vehicle-injected group, whereas the plasma adrenocorticotropic hormone response to IM stress was reduced only by pretreatment with an i.cv. injection of 6-OHDA. These data suggest that (1) neither the pituitary nor the adrenal gland is a major source of increased plasma IL-6 induced by IM stress; (2) the HPA axis exhibits a suppressive regulatory role in the IM-induced IL-6 response; (3) both the central and peripheral catecholamines play critical roles in causing IL-6 elevation induced by IM stress, and (4) the involvement of peripheral catecholamines in elevating plasma IL-6 by IM stress is independent of HPA axis activation. Moreover, to determine whether splenocytes are the source of IM-induced plasma IL-6, splenectomized (Splex) rats were examined for their plasma IL-6 response to IM stress. The peak IL-6 levels after IM in Splex animals were significantly higher than those in sham-operated animals (p < 0.05). The immune-competent cells in the spleen, as well as the pituitary and adrenal glands do not seem to be a source of the increased levels of plasma IL-6 induced by IM st