Figure 6.

Ca²⁺-dependent of PACAP- and Forskolin-induced PRL promoter activity in αT3-1 Cells. αT3-1 cells were transiently transfected with pPRL(−1156).LUC for 6 h by using lipofectamine. After 18 h recovery, the cells were incubated with respective drugs. The cells were treated for 24 hr with increasing doses of (A) Ca²⁺ ionophore A23187 or (B) L-type VSCC activator Bay K8644. Inhibiting extracellular Ca²⁺ entry on PRL mRNA expression in carp pituitary cells was examined. In this study, αT3-1 cells over-expressed pPRL(−1156).LUC were treated with (C) oPACAP₃₈ (10 nM, 24 hrs) or (D) Forskolin (100 nM, 24 hrs) in the Ca²⁺-free medium (with different doses of EGTA). Further, αT3-1 cells over-expressed pPRL(−1156).LUC were treated with (E) oPACAP₃₈ (10 nM, 24 hrs) or (F) Forskolin (100 nM, 24 hrs) in the presence or absence of L-type VSCC blocker nifedipine (10 μM). After drug treatment, cell lysate was prepared for dual-luciferase measurement. Data presented were expressed as percentage of control by conversing the ratio of firefly and renilla luciferase in the same sample. Data presented are expressed as mean ± SEM (n = 4) and different letters denote a significant difference at p < 0.05 (ANOVA followed by Fisher’s LSD Test).