In the kidney, approximately 80% of the filtered phosphate (Pi) is reabsorbed along the proximal tubule. Changes in renal Pi reabsorption are associated with modulation of the sodium-dependent Pi cotransporter type IIa (NaPi-IIa) and type IIc (NaPi-IIc) protein abundance in the brush-border membrane (BBM) of proximal tubule cells. NaPi-IIa is mainly regulated by dietary Pi intake and parathyroid hormone (PTH). The purpose of the present study was to elucidate the effect of alteration in dietary magnesium (Mg2+) intake on renal Pi handling. Urinary Pi excretion and renal expression of NaPi-IIa and NaPi-IIc were analyzed in rats fed a normal (0.2%) or high-Mg2+ (2.5%) diet. A high-Mg2+ diet resulted in decreased renal Pi excretion and increased protein expression of NaPi-IIa and NaPi-IIc. Serum FGF-23 (fibroblast growth factor 23) levels were unchanged under a high-Mg2+ diet. Serum PTH levels were slightly decreased under a high-Mg2+ diet. To examine whether the observed changes in renal Pi reabsorption are PTH dependent, expression of NaPi-IIa and NaPi-IIc was also analyzed in parathyroidectomized (PTX) rats fed a normal or high-Mg2+ diet. In PTX rats, Mg2+ had no significant effect on renal Pi excretion or NaPi-IIa protein expression. Mg2+ increased NaPi-IIc protein expression in PTX rats. This experiment shows for the first time on the molecular level how Mg2+ stimulates renal Pi reabsorption. Under a high-Mg2+ diet, NaPi-IIa expression is dependent on PTH levels, whereas NaPi-IIc expression seems to be independent of PTH levels.