The presence of a circadian clock in the retinal pigment epithelium (RPE) was discovered recently. However, little is known about mechanisms or processes regulated by the RPE clock. We cultured ARPE-19 monolayers in a transwell culture system, and we found rhythmic mRNA expression of the sodium-potassium-chloride co-transporter SLC12A2. We localized the corresponding protein product, NKCC1, on the apical membrane of ARPE-19 cells. We found that concentrations of sodium, potassium, and chloride oscillated in apical supernatants. The ion concentration gradients between supernatants strongly correlated with SLC12A2 mRNA expression. Our results suggest that the circadian clock regulates ion transport by the RPE via NKCC1 expression.