Previous data have suggested an age-related increase in renal tubular secretion of digoxin in infants and children receiving long-term digoxin therapy. This phenomenon could be the result of a maturational process or secondary to chronic substrate stimulation. To investigate this question, two groups of 2-week-old paired littermate rats received intraperitoneal injections of either digoxin or an equal volume of normal saline (control) on alternate days until sacrificed at 4, 6, and 8 wk of age. An additional group of 12-wk-old-rats were studied as controls.125I-labeled digoxin uptake was measured in renal cortical slices as the cPM/mg wet tissue slice/medium ratio (S/M). Both digoxin-treated and control rats demonstrated significant age-related increments in digoxin uptake. S/M ratios at 4, 6, 8, and 12 wk in the control group were 1.34 × 0.06, 1.39 × 0.14, 1.62 × 0.18 and 1.93 × 0.23, respectively (mean × S.D.) (r= 0.81;P< 0.001). S/M ratios in the digoxin-treated animals at 4, 6, and 8 wk were 1.28 × 0.16, 1.33 × 0.09, and 1.52 × 0.23, respectively (r= 0.50;P< 0.025), but did not differ significantly at each age from those in the control group.125I uptake was significantly reduced by both dinitrophenol and sodium azide, as well as by a 100% nitrogen atmosphere.