Figure 6.

Trpv5/6 has different roles in regulating IGF-PI3K-Akt signaling under normal and low [Ca²⁺]. (a–d) Effects of cadmium (Cd²⁺), ruthenium red (RR), lanthanum (La³⁺), and gadolinium (Gd³⁺) on pAkt. Larvae (72 hpf) were transferred to 0.2 mM or 0.001 mM [Ca²⁺] water containing the indicated concentrations of the drugs. After 8 h, they were stained for pAkt and scored. The total number of larvae analyzed from three independent experiments is shown on the top of each column. (e) Blockage of the Trpv5/6 channel abolishes the low [Ca²⁺]-induced increase in NaR cell density. Larvae (72 hpf) were transferred to 0.2 mM or 0.001 mM [Ca²⁺] water containing the indicated concentrations of Cd²⁺. At 120 hpf, they were sampled and analyzed by in situ hybridization for igfbp5a. (f) The low [Ca²⁺]-induced Akt signaling is not mediated by intracellular Ca²⁺/CaM signaling. Larvae (72 hpf) were transferred to 0.001 mM [Ca²⁺] water containing BAPTA-AM (100 μM), W7 (50 μM), or Calmidazolium (Calm, 1 μM). After 8 h, they were fixed, stained for pAkt, and scored as described earlier. (g) KCl treatment activates Akt signaling in an IGF1R and PI3K-dependent manner. Larvae (72 hpf) were transferred to 0.2 mM or 0.001 mM [Ca²⁺] water containing KCl (100 mM), sorbitol (200 mM), NMDG-Cl (100 mM), BMS (0.3 μM), and Wortmannin (0.06 μM), alone or in combination. After 2 h, they were stained for pAkt. (h) KCl reverses the effect of Trpv5/6 blockage. Larvae (72 hpf) were transferred to 0.2 mM or 0.001 mM [Ca²⁺] water containing KCl (100 mM), RR (5 μM), and Cd²⁺ (1 μM), alone or in combination. After 2 h, they were stained for pAkt