Early studies performed mainly in vitro using rat small intestine (SI) demonstrated that the duodenum and jejunum are responsible for the bulk of intestinal phosphate (Pi) absorption ( Walling, 1977). The protein responsible for this process has been identified as NaPi IIb, a sodium-dependent transporter expressed at the enterocyte brush border membrane (Murer et al., 2001). More recent studies have demonstrated that this protein is expressed in the ileum, as well as the proximal SI of mice (Personal communication). Based on these discrepancies, we aimed to examine the involvement of distinct SI regions on the absorption of Pi in vivo in rats and mice. Male Sprague-Dawley rats (250 g) were anaesthetized with pentobarbitone sodium (50 mg/kg i.p.). Segments of duodenum, jejunum or ileum (7-10 cm long) were selected and flushed with warm 0.9% saline, followed by air. Uptake buffer (500μl) containing 100μM Pi labeled with ³²P was instilled into the lumen and the segment was tied off. Blood was collected via cardiac puncture after 10 minutes and Pi absorption calculated from the ³²P activity in plasma and expressed as a percentage of that in the uptake buffer. At the end of experiments animals were humanely killed. Results are given as a mean percentage of Pi absorbed into 1ml of plasma by 5cm of intestine ± SEM (n=6). The same procedure was performed using male C57 BL/6 mice with the following minor modifications. Segments of proximal SI (7-10cm) comprised the duodenum and jejunum, whilst a distinct region of ileum was selected. For mice 200μl of uptake buffer was used to avoid intestinal distention. Pi transfer from lumen to blood by the rat SI was highest in the duodenum (0.089 ± 0.025%) followed by the jejunum (0.021 ± 0.005%), with relatively little absorption occurring in the ileum (0.0068 ± 0.001%). In contrast, in the mouse, SI uptake was significantly higher in the ileum than in the duodenum/jejunum segment (0.290 ± 0.036 vs. 0.143 ± 0.027%, P〉0.01 using an unpaired t test). The mouse small intestine has the ability to absorb phosphate in all segments examined, with the highest rate of absorption occurring in the ileum. In contrast the ileum of the rat has little ability to absorb this anion. These findings highlight a distinct difference in the handling of phosphate by the rat and mouse SI.