Influence of the aquatic environment and 1α,25(OH)2 vitamin D3 on calcium influx in the intestine of adult zebrafish

Abstract

We investigated the effects of environment calcium challenge and 1α,25(OH)₂ vitamin D₃ (1,25-D₃) on ⁴⁵Ca²⁺ influx in the intestine of zebrafish (ZF). In vitro⁴⁵Ca²⁺ influx was analyzed using intestines from fed and fasted fish. ZF were held in water containing Ca²⁺ (0.02, 0.7, 2.0 mM) to analyze the ex vivo⁴⁵Ca²⁺ influx in the intestine and for histology. Intestines from fish held in water with Ca²⁺ were incubated ex vivo to characterize ion channels, receptors, ATPases and ion exchangers that orchestrate ⁴⁵Ca²⁺ influx. For in vitro studies, intestines were incubated with antagonists/agonist or inhibitors to study the mechanism of 1,25-D₃ on ⁴⁵Ca²⁺ influx. Fasted ZF reached a plateau for ⁴⁵Ca²⁺ influx at 30 min. In vivo fish at high Ca²⁺ stimulated ex vivo⁴⁵Ca²⁺ influx and increased the height of intestinal villi in low calcium. In the normal calcium, ⁴⁵Ca²⁺ influx was maintained by the reverse-mode Na⁺/Ca²⁺ (NCX) activation, Na⁺/K⁺-ATPase pump and sarco/endoplasmic reticulum calcium ATPase (SERCA) pump. However, Ca²⁺ hyperosmolarity is supported by L-type voltage-dependent calcium channels (L-VDCC), transient receptor potential vanilloid subfamily 1 (TRPV1) and Na⁺/K⁺-ATPase activity. The calcium challenge causes morphological alteration and changes the ion type-channels involved in the intestine to maintain hyperosmolarity. 1,25-D₃ stimulates Ca²⁺ influx in normal osmolarity coordinated by L-VDCC activation and SERCA inhibition to keeps high intracellular calcium in intestine. Our data showed that the adult ZF regulates the calcium challenge (per se osmolarity), independently of the hormonal regulation to maintain the calcium balance through the intestine to support ionic adaptation.

Keywords: Calcium; Danio rerio; Homeostasis; Intestine; Osmolarity; Vitamin D.