The Na⁺/H⁺ exchanger isoform 3 is required for active paracellular and transcellular Ca²⁺ transport across murine cecum.

Intestinal calcium (Ca²⁺) absorption occurs via paracellular and transcellular pathways. Although the transcellular route has been extensively studied, mechanisms mediating paracellular absorption are largely unexplored. Unlike passive diffusion, secondarily active paracellular Ca²⁺ uptake occurs against an electrochemical gradient with water flux providing the driving force. Water movement is dictated by concentration differences that are largely determined by Na⁺ fluxes. Consequently, we hypothesized that Na⁺ absorption mediates Ca²⁺ flux. NHE3 is central to intestinal Na⁺ absorption. NHE3 knockout mice (NHE3-/-) display impaired intestinal Na⁺, water, and Ca²⁺ absorption. However, the mechanism mediating this latter abnormality is not clear. To investigate this, we used Ussing chambers to measure net Ca²⁺ absorption across different segments of wild-type mouse intestine. The cecum was the only segment with net Ca²⁺ absorption. Quantitative RT-PCR measurements revealed cecal expression of all genes implicated in intestinal Ca²⁺ absorption, including NHE3. We therefore employed this segment for further studies. Inhibition of NHE3 with 100 μM 5-(N-ethyl-N-isopropyl) amiloride decreased luminal-to-serosal and increased serosal-to-luminal Ca²⁺ flux. NHE3-/- mice had a >60% decrease in luminal-to-serosal Ca²⁺ flux. Ussing chambers experiments under altered voltage clamps (-25, 0, +25 mV) showed decreased transcellular and secondarily active paracellular Ca²⁺ absorption in NHE3-/- mice relative to wild-type animals. Consistent with this, cecal Trpv6 expression was diminished in NHE3-/- mice. Together these results implicate NHE3 in intestinal Ca(2+) absorption and support the theory that this is, at least partially, due to the role of NHE3 in Na⁺ and water absorption.