Figure 5.

Functional role of cAMP/PKA pathway in PACAP stimulation of 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. (A) αT3-1 cells over-expressed pPRL(−1156).LUC were treated for 24 hrs with increasing doses of cpt-cAMP (1–100 μM). (B) αT3-1 cells over-expressed pPRL(−1156).LUC were treated for 24 hrs with increasing doses of Forskolin (10–1000 nM). Effects of cAMP/PKA inhibitors on PACAP-induced PRL mRNA expression were then investigated. αT3-1 cells over-expressed pPRL(−1156).LUC were challenged with oPACAP₃₈ (10 nM, 24 hr) in the presence or absence of (C) AC inhibitor MDL12330A (10 μM) or (D) PKA blocker H89 (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).