Calcitonin-like diuretic hormone influences reproduction in adult female Rhodnius prolixus

Calcitonin-like diuretic hormone (CT/DH) is a pleiotropic neuropeptide in insects that plays a role in regulating diuresis in Rhodnius prolixus, a blood-feeding insect and a vector of Chagas' disease. Here, we examine the role of this neuropeptide in another feeding-dependent event, that of reproduction in female R. prolixus. CT/DH-like immunoreactivity is present in the female reproductive system, including cells of the tropharium and pedicel of the ovariole, and nerve processes over the oviducts and calyx; tissues involved in egg maturation, ovulation, and egg laying. qPCR analysis reveals that the transcripts for the Rhopr-CT/DH receptors, Rhopr-CT/DH-R-1B/C and Rhopr-CT/DH-R-2B are present in reproductive tissues and fat body, and appear to be regulated post blood meal, a stimulus that triggers diuresis and reproduction. Knockdown of the two receptors using RNA interference reduces the number of eggs produced and their hatching rate. Downregulation of Rhopr-CT/DH-Rs does not alter protein levels, including vitellogenin, in the fat body or hemolymph and does not change transcript expression of RhoprVg1 in the fat body. Knockdown does lead to a decrease in transcript expression of RhoprVg1 and the Vg receptor, RhoprVgR, in the ovaries. In vitro analyses show that incubation of tissues with Rhopr-CT/DH leads to a notable but not significant increase in transcript expression of RhoprVg1 and a significant increase in RhoprVgR in the ovaries but has no effect on fat body RhoprVg1 or vitellogenin release. Rhopr-CT/DH causes a dose-dependent increase in phasic contraction of the oviducts and increases the amplitude and frequency of intrinsic rhythmic contractions. Collectively, these results suggest the involvement of Rhopr-CT/DH in coordinating aspects of reproduction in female R. prolixus.