The aim of our study was to evaluate whether adrenals are involved in the secretion of endogenous digitalis-like factor(s) with polarity similar to (or less than) that of ouabain (EDLF-1), and whether acute plasma volume expansion is a physiological releasing stimulus for this factor(s) in humans. For this purpose, we measured the concentration of this substance(s) by a human placenta radioreceptor assay (RRA) and by the Du-Pont-NEN ouabain-EIA (immunoreactive ouabain, I-Oua) in plasma C18-extracts of eight normotensives and six patients with bilateral adrenalectomy before and after acute salt loading (2 lt 0.9% NaCl/2 h). The study was repeated after 2 weeks of increased sodium intake (200 mEq/day). Under basal conditions, EDLF-1 by RRA and I-Oua were similar in adrenalectomized patients and in controls and were not significantly modified by saline infusion. After 15 days of high sodium intake, basal plasma EDLF-1 and I-Oua were not significantly different from prediet levels, both in adrenalectomized patients and controls and were likewise unaffected by saline loading. Saline infusion, by contrast, significantly (P < or = .05) suppressed hematocrit and PRA and increased ANP both in controls and in patients, either before or after prolonged high dietary sodium. Plasma aldosterone (ALD) was similarly reduced (P < .001) in controls and, as expected, was undetectable in adrenalectomized patients. Our data indicate that in adrenalectomized patients circulating levels of EDLF-1 and I-Oua are similar to those of controls and that, in both groups, acute saline loading before and after sodium repletion does not influence circulating levels of these compounds. These findings suggest that, at least in humans, adrenals are not the main source of endogenous digitalis-like factor(s) with polarity similar to (or less than) that of ouabain, and that plasma volume expansion may be not a sufficient stimulus for the release of this factor(s).