The phenotypes of calbindin-D9k (CaBP-9k) and -28k (CaBP-28k) single knockout (KO) mice are similar to wild-type (WT) mice due to the compensatory action of other Calcium transport proteins. In this study, we generated CaBP-9k/CaBP-28k double knockout (DKO) mice in order to investigate the importance of CaBP-9k and CaBP-28k in active calcium processing. Under normal dietary conditions, DKO mice did not exhibit any changes in phenotype or the expression of active calcium transport genes as compared to WT or CaBP-28k KO mice. Under calcium-deficient dietary conditions, the phenotype and expression of calcium transport genes in CaBP-28k KO mice were similar to WT, whereas in DKO mice, serum calcium levels and bone length were decreased. The intestinal and renal expression of transient receptor potential vanilloid member 6 (TRPV6) mRNA was significantly decreased in DKO mice fed a calcium-deficient diet as compared to CaBP-28k KO or WT mice, and DKO mice died after 4 weeks on a calcium-deficient diet. Body weight, bone mineral density (BMD) and bone length were significantly reduced in all mice fed a Calcium and 1,25-(OH)(2)D-3-deficient diet, as compared to a normal diet, and none of the mice survived more than 4 weeks. These results indicate chat deletion of CaBP-28k alone does not affect body calcium homeostasis, but that deletion of CaBP-9k and CaBP-28k has a significant effect oil calcium processing under calcium-deficient conditions, confirming the importance of dietary calcium and 1,25-(OH)(2)D-3 during growth and development. (C) 2008 Elsevier Inc. All rights reserved.