Sulfakinins (SKs) are neuropeptides that have been shown to function as feeding satiety factors in insects. We confirmed the presence of two SKs (Rhopr-SK-1 and Rhopr-SK-2) in Rhodnius prolixus. Reverse transcriptase quantitative PCR (RT-qPCR) demonstrated that the Rhopr-SK transcript is mainly expressed in the central nervous system (CNS) in unfed fifth-instar R. prolixus. Fluorescent in situ hybridization (FISH) showed transcript expression only in neurons in the brain. Immunohistochemical staining of SK-like peptides was observed in the same neurons in the brain and in processes extending throughout the CNS, as well as over the posterior midgut and anterior hindgut. Rhopr-SK-1 induced contractions of the hin... More
Sulfakinins (SKs) are neuropeptides that have been shown to function as feeding satiety factors in insects. We confirmed the presence of two SKs (Rhopr-SK-1 and Rhopr-SK-2) in Rhodnius prolixus. Reverse transcriptase quantitative PCR (RT-qPCR) demonstrated that the Rhopr-SK transcript is mainly expressed in the central nervous system (CNS) in unfed fifth-instar R. prolixus. Fluorescent in situ hybridization (FISH) showed transcript expression only in neurons in the brain. Immunohistochemical staining of SK-like peptides was observed in the same neurons in the brain and in processes extending throughout the CNS, as well as over the posterior midgut and anterior hindgut. Rhopr-SK-1 induced contractions of the hindgut in a dose-dependent manner, but had no effect on heartbeat frequency. Injection of Rhopr-SK-1 significantly decreased the overall size of the blood meal consumed, suggesting SK’s role as a satiety factor in R. prolixus. Two seven-transmembrane Rhopr-SK G protein-coupled receptors (GPCRs), Rhopr-SKR-1 and Rhopr-SKR-2, were cloned and characterized. RT-qPCR of the two Rhopr-SKR transcripts revealed that the target tissues for Rhopr-SK-1 and Rhopr-SK-2 are located in the CNS, heart, gut, salivary glands, Malpighian tubules, fat body, as well as, male and female reproductive systems. Rhopr-SK-1 inhibits contractions of the oviduct and ejaculatory duct in adult R. prolixus, whilst having no effect on contractions of the bursa. A III functional receptor assay that utilized Human Embryonic Kidney (HEK)-293 cells stably expressing a modified cyclic nucleotide-gated (CNG) channel (HEK293/CNG) characterized the two Rhopr-SK receptors. Modified HEK293/CNG assays showed that the PLC/IP3 pathway, and not the cAMP pathway, is solely initiated upon the activation of either Rhopr-SK receptor. Nonsulfated Rhopr-SK-1 was not able to activate either Rhopr-SK receptor. Both Rhopr-SKRs were exclusively activated via Rhopr-SK-1 and Rhopr-SK-2. The importance of the SK signaling pathway in the modulation of feeding was demonstrated by the knockdown of the transcripts for Rhopr-SKs and Rhopr-SKRs, which led to an increase in the overall intake of blood during feeding.
