Accumulating evidence has emerged indicating that growth hormone-releasing factor (GRF) is present not only in the hypothalamus but in other tissues as well. Using highly specific and sensitive radioimmunoassays and immunoaffinity chromatography, we found low but clearly measurable GRF-like immunoreactivity (GRF-LI) levels in rat peripheral plasma. In order to verify the immunological findings, the peripheral plasma GRF-LI was characterized using fast protein liquid chromatography. The immunological peak was eluted at the position of synthetic rat GRF standard. These findings demonstrate that rat peripheral plasma GRF is immunologically and chromatographically indistinguishable from authentic rat GRF. Moreover, we performed studies with hypophysectomy to assess whether peripheral plasma GRF-LI changes in physiological status. At 4 weeks after hypophysectomy, there was a significant (p<0.05) increment in the rat plasma GRF-LI [12.4±0.5 (±SEM) pg/ml in hypophysectomized rats as opposed to 5.8±0.4 pg/ml in sham-operated control rars]. On the other hand, hypothalamic GRF-LI fell significantly as compared that of controls (36.1±1.0 vs. 78.3±3.0 pg/mg wet weight tissue). A similar pattern of changes in GRF-LI at 10 weeks after hypophysectomy was also revealed. The source of rat peripheral plasma GRF has not yet been elucidated. Our results, however, may suggest that GRF levels are modulated by negative feedback at the level of the hypothalamus by a pituitary factor, presumably growth hormone (GH) and that hypothalamic GRF release exceeds its synthesis in hypophysectomized rats.