Involvement of the calcium-sensing receptor in calcium homeostasis in larval zebrafish exposed to low environmental calcium.

The involvement of the calcium-sensing receptor (CaSR) in Ca(2+) homeostasis was investigated in larval zebrafish, Danio rerio. The expression of CaSR mRNA was first observed at 3 h posfertilization (hpf) and increased with development until plateauing at ∼48 hpf. At 4 dpf, CaSR mRNA was increased in fish acclimated to low Ca(2+) water (25 μM vs. 250 μM in normal water). Using immunohistochemistry and confocal microscopy, we demonstrated that the CaSR is expressed in the olfactory epithelium, neuromasts, ionocytes on the yolk sac epithelium, and corpuscles of Stannius. Results of double immunohistochemistry and/or in situ hybridization indicated that the CaSR is localized to a subset of mitochondrion-rich ionocytes enriched with Na(+)/K(+)-ATPase and epithelial Ca(2+) channel (ecac). Translational knockdown of the CaSR prevented 4 dpf larvae from regulating whole body Ca(2+) levels when exposed to a low Ca(2+) environment. Further, the increases in ecac mRNA expression and Ca(2+) influx, normally associated with exposure to low-Ca(2+) water, were prevented by CaSR knockdown. These findings demonstrate that larval zebrafish lacking the CaSR lose their ability to regulate Ca(2+) when confronted with a low-Ca(2+) environment. Results from real-time PCR suggested that the mRNA expression of the hypocalcemic hormone stanniocalcin (stc-1) remained elevated in the CaSR morphants following acclimation to low-Ca(2+) water. Overall, the results suggest that the CaSR is critical for Ca(2+) homeostasis in larval zebrafish exposed to low environmental Ca(2+) levels, possibly owing to its modulation of stanniocalcin mRNA expression.