Infants are exposed to variable copper (Cu) intake; Cu in breast milk is low, while infant formulas vary in Cu content, as well as the water used for their preparation. Little is known about the regulation of Cu absorption during infancy. The objectives of this study were to determine effects of Cu supplementation on Cu absorption and tissue distribution and the expression of Cu transporters in an infant rat model. Suckling rat pups were orally dosed with 0, 10, or 25 µg Cu /day. Intestine and liver were collected at d 10 and 20 and Cu concentration, Ctr1, Atp7A, Atp7B, and MT mRNA and protein levels were measured. ⁶⁷Cu absorption was measured at d 10 and 20. Total ⁶⁷Cu absorption decreased and intestinal ⁶⁷Cu retention increased with increased Cu intake. At day 10, intestine Cu concentration, MT mRNA and Ctr1 protein levels increased with supplementation but no changes in Atp7A or Atp7B levels were observed. At day 20, intestine Cu concentration was unaffected by Cu supplementation but Ctr1 protein and Atp7A mRNA and protein levels were higher than in controls. In liver, Cu level reflected Cu intake at d 10 and d 20. There was a significant increase in Ctr1, Atp7B, and MT mRNA expression in liver at both ages with Cu supplementation. In conclusion, the ability of suckling rat pups to tolerate varying amounts of dietary Cu may be due to changes in Cu transporters, facilitated by transcriptional and post-translational mechanisms. Despite these adaptive changes, Cu supplementation resulted in elevated alanine aminotransferase (ALT) levels, suggesting a risk of Cu toxicity with supplementation during infancy.