在昆虫中,促肾上腺皮质激素释放因子/利尿激素(CRF/DHs)负责调节一系列生理和行为过程,如喂养,利尿,和繁殖。Rhopr-CRF/DH在常红的摄食和利尿中起关键作用,一种吸血的昆虫和人类查加斯病的媒介。这里,我们扩展了我们对这种神经激素在成年雌性R.prolixus生殖中的作用的理解。双标记免疫组织化学显示了CRF样和糖蛋白激素(GPA2/GPB5)亚基GPB5样免疫反应性的共定位染色,它们在成年女性R.prolixus中的相同神经分泌细胞(NSCs)中及其神经位点,这表明这些肽可以共同调节生理过程。qPCR分析显示,Rhopr-CRF/DH受体2(Rhopr-CRF/DH-R2)的转录物在成年雌性R.prolixus的生殖组织和脂肪体(FB)中表达,其表达在血餐后(PBM)增加,引发利尿和生殖的刺激。利用RNA干扰,Rhopr-CRF/DH-R2的转录表达被敲低,通过检查主要的蛋黄蛋白来监测产蛋量,卵黄蛋白原(Vg),产卵的数量和质量,和它们的孵化率。将dsCRFR2注射到成年雌性体内可降低Rhopr-CRF/DH-R2转录物的表达,加速卵子发生,增加产卵的数量,并降低雌性R.prolixus的孵化率。Rhopr-CRF/DH-R2的下调导致RhoprVg1在脂肪体和卵巢中的转录表达增加,并增加Vg受体的转录水平,RhoprVgR,在卵巢里。还观察到脂肪体和血淋巴中Vg含量的显着增加。将分离的组织与Rhopr-CRF/DH一起孵育会导致脂肪体中RhoprVg1和卵巢中RhoprVg1的转录本表达显着降低。此外,Rhopr-CRF/DH降低蜕皮类固醇生物合成酶的转录物表达并降低含有分离卵巢的培养基中的蜕皮类固醇滴度。这些结果表明CRF信号通路参与生殖,Rhopr-CRF/DH在成年雌性R.prolixus中充当性腺抑制激素,如前面所示的共定位糖蛋白,GPA2/GPB5。
Within insects, corticotropin-releasing factor/diuretic hormones (CRF/DHs) are responsible for the modulation of a range of physiological and behavioural processes such as feeding, diuresis, and reproduction. Rhopr-CRF/DH plays a key role in feeding and diuresis in Rhodnius prolixus, a blood-gorging insect and a vector for human Chagas disease. Here, we extend our understanding on the role of this neurohormone in reproduction in adult female R. prolixus. Double-label immunohistochemistry displays co-localized staining of CRF-like and the glycoprotein hormone (GPA2/GPB5) subunit GPB5-like immunoreactivity in the same neurosecretory cells (NSCs) in the mesothoracic ganglionic mass (MTGM) and in their neurohemal sites in adult female R. prolixus, suggesting these peptides could work together to regulate physiological processes. qPCR analysis reveals that the transcript for Rhopr-CRF/DH receptor 2 (Rhopr-CRF/DH-R2) is expressed in reproductive tissues and fat body (FB) in adult female R. prolixus, and its expression increases post blood meal (PBM), a stimulus that triggers diuresis and reproduction. Using RNA interference, transcript expression of Rhopr-CRF/DH-R2 was knocked down, and egg production monitored by examining the major yolk protein, vitellogenin (Vg), the number and quality of eggs laid, and their hatching ratio. Injection of dsCRFR2 into adult females reduces Rhopr-CRF/DH-R2 transcript expression, accelerates oogenesis, increases the number of eggs produced, and reduces hatching rate in female R. prolixus. Downregulation of Rhopr-CRF/DH-R2 leads to an increase in the transcript expression of RhoprVg1 in the fat body and ovaries, and increases the transcript level for the Vg receptor, RhoprVgR, in the ovaries. A significant increase in Vg content in the fat body and in the hemolymph is also observed. Incubation of isolated tissues with Rhopr-CRF/DH leads to a significant decrease in transcript expression of RhoprVg1 in the fat body and RhoprVg1 in the ovaries. In addition, Rhopr-CRF/DH reduces transcript expression of the ecdysteroid biosynthetic enzymes and reduces ecdysteroid titer in the culture medium containing isolated ovaries. These results suggest the involvement of the CRF-signaling pathway in reproduction, and that Rhopr-CRF/DH acts as a gonad-inhibiting hormone in the adult female R. prolixus, as previously shown for the colocalized glycoprotein, GPA2/GPB5.