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Immune responses and intracellular signaling pathways were examined after hemolymph of Litopenaeus vannamei being incubated in Crustacean hyperglycemic hormone (CHH), dopamine (DA) and DA antagonist (Y). The results showed that the effect CHH and CHH + DA + Y on viability of hemocytes were no significant changes compared to the control group. However, in DA, DA + Y and CHH + DA groups, the viability of hemocytes decreased significantly. The phagocytic activity and the antibacterial activity of CHH group were increased significantly within 12h. Whereas the CHH + DA, DA were significantly lower than the control. PO in haemolymph was up-regulated after CHH and DA incubation. The proPO has the opposite change in all groups. In addition, DA + Y, CHH + DA + Y has a similar trend with the DA and CHH respectively. Furthermore, a significant increase of cAMP, CaM and cGMP were found in treatment groups except for the CaM concentration of the CHH group and the cGMP concentration of DA group. There is no significant change observed in the CHH group about CaM concentration. Whereas the cGMP of DA group decreased within 12h. The results suggest that DA could depress the immune responses by cAMP-, CaM-pathways. However, the CHH is on the contrary, which transduced the signals from cAMP, cGMP to PKA, PKC and PKG to enhance the immune response parameters.DA could depress the immune responses by cAMP-, CaM-pathways in vitro.CHH could enhance the immune response parameters in vitro.CHH transduce the signals from cAMP, cGMP to PKA, PKC and PKG.DA and CHH simultaneous cause immune response through PKA, PKC and PKG pathways.