The calcium dependency and tetrodotoxin sensitivity of extracellular dopamine levels were assessed by microdialysis in the neostriatum of 5-day-old rat pups and were compared with those obtained in adult rats. The removal of calcium from the dialysate reduced spontaneous levels of extracellular dopamine to 20% of normal in the 5-day-old pups and to 10% of normal in the adults. Calcium-free dialysate also decreased potassium-evoked dopamine release to approximately 20% of baseline in both ages. Furthermore, the addition of tetrodotoxin to the dialysate decreased spontaneous levels of extracellular dopamine to 10% of baseline in both ages. The effects of calcium removal and the addition of tetrodotoxin on extracellular levels of the dopamine metabolite 3,4-dihydroxyphenylacetic acid were less pronounced. The results of this study demonstrate that extracellular levels of dopamine sampled by microdialysis in rats as young as 5 days of age are both calcium dependent and tetrodotoxin sensitive; thus, they are derived from neuronal activity and not from injury caused by acute implantation of the probe. Other age-related differences support the hypothesis that dopamine release and turnover is greater in immature rats and may represent a form of compensation for incomplete dopamine nerve terminal ingrowth.