Spreading behavior of hemocytes (= insect blood cells) is essential for cellular immune responses against various microbial pathogens. It is activated by prostaglandin E₂ (PGE₂) via its membrane receptor associated with secondary messenger, cAMP, in insects. This study observed an increase of calcium ion (Ca²⁺) level after an acute increase of cAMP induced by PGE₂ treatment and clarified the intracellular signals underlying the hemocyte-spreading behavior. Inhibition of Ca²⁺ flux significantly impaired the hemocyte-spreading and subsequent cellular immune response, phagocytosis. The up-regulation of intracellular Ca²⁺ in response to PGE₂ was dependent on cAMP because RNA interference (RNAi) of PGE₂ receptor expression or inhibiting adenylate cyclase prevented Ca²⁺ mobilization. The up-regulation of Ca²⁺ was induced by inositol triphosphate (IP₃) via its specific IP₃ receptor. Furthermore, inhibition of ryanodine receptor impaired Ca²⁺ mobilization, suggesting Ca²⁺-induced Ca²⁺ release. However, the effective spreading behavior of hemocytes was dependent on both secondary messengers. Ca²⁺ signal stimulated by cAMP was required for activating small G proteins because RNAi treatments of small G proteins such as Rac1, RhoA, and Cdc42 failed to stimulate hemocyte-spreading. In contrast, aquaporin was activated by cAMP. Its activity was necessary for changing cell volume during hemocyte-spreading. These results indicate that PGE₂ mediates hemocyte-spreading via cAMP signal to activate aquaporin and via Ca²⁺ signal to activate actin cytoskeletal rearrangement.

血细胞（=昆虫血细胞）的传播行为对于针对各种微生物病原体的细胞免疫反应至关重要。它是由前列腺素 E ₂ (PGE ₂ ) 通过其与昆虫中的次级信使 cAMP 相关的膜受体激活的。本研究观察到PGE ₂处理诱导的 cAMP 急剧增加后钙离子 (Ca ²⁺ ) 水平的增加，并阐明了血细胞扩散行为背后的细胞内信号。^{Ca 2+}通量的抑制显着损害了血细胞扩散和随后的细胞免疫反应、吞噬作用。^{细胞内 Ca 2+}响应 PGE ₂的上调依赖于 cAMP，因为 PGE ₂受体表达的 RNA 干扰 (RNAi) 或抑制腺苷酸环化酶阻止了 Ca ²⁺动员。三磷酸肌醇（IP ₃）通过其特异性IP ₃受体诱导Ca ²⁺的上调。此外，兰尼碱受体的抑制损害了Ca ²⁺的动员，表明Ca ²⁺诱导的Ca ²⁺释放。然而，血细胞的有效传播行为取决于两个次要信使。钙²⁺cAMP 刺激的信号是激活小 G 蛋白所必需的，因为对小 G 蛋白（如 Rac1、RhoA 和 Cdc42）的 RNAi 处理未能刺激血细胞扩散。相反，水通道蛋白被 cAMP 激活。它的活性对于在血细胞扩散过程中改变细胞体积是必要的。这些结果表明，PGE ₂通过 cAMP 信号介导血细胞扩散以激活水通道蛋白，并通过 Ca ²⁺信号以激活肌动蛋白细胞骨架重排。