Orcokinin and orcomyotropin were originally described as neuropeptides in crustaceans but have now been uncovered in many species of insects in which they are called orcokinin-A (OK-A) and orcokinin-B (OK-B), respectively. The two groups of mature peptides are products of alternatively spliced transcripts of the single copy gene orcokinin in insects. We investigated the expression patterns and the functions of OK-A and OK-B in the red flour beetle Tribolium castaneum. In situ hybridization and immunohistochemistry using isoform-specific probes and antibodies for each OK-A and OK-B suggests that both peptides are co-expressed in 5 to 7 pairs of brain cells and in the midgut enteroendocrine cells, which contrasts to expression patterns in other insects in which the two peptides are expressed in different cells. We developed a novel behavioral assay to assess the phenotypes of orcokinin RNA interference (RNAi) in T. castaneum. RNAi of ok-a and ok-b alone or in combination resulted in higher frequencies and longer durations of death feigning in response to mechanical stimulation in the adult assay. In the larval behavioral assays, we observed longer recovery times from knockout induced by water submergence in the insects treated with RNAi for ok-a and ok-b alone or in combination. We conclude that both OK-A and OK-B have “awakening” activities and are potentially involved in the control of circadian rhythms.

No abstract

Transient receptor potential channels (TRPs) are a family of cation channels involved in various sensory mechanisms in Drosophila melanogaster, including mechanosensing. Phylogenetic analysis of mechanosensory TRPs in seven members of the TRPV, TRPN and TRPA subfamilies reveals a unique TcTrpA5 in Tribolium castaneum that is likely lost in D. melanogaster. Because mechanosensors are implicated in various key physiology, we investigate the roles of eight candidate mechanosensory TRPs in survival, walking behaviour and tonic immobility in T. castaneum using the RNA interference technique. Double-stranded RNA treatment of nompC (dsNompC) and trpA5 (dsTrpA5) results in eclosion failure, causing 93% mortality of beetles subjected to each of these treatments. The beetles that survive the dsNompC treatment show defects during sclerotization of the elytra. Adult beetles treated with dsNanchung and dsInactive show defects in the folding of the joint between the femur and tibia segments of the hind legs, resulting in abnormal walking behaviour and reduced walking speed. Regarding tonic immobility induced by mechanical stimulations on the ventral surface, knockout responses (death feignings) are significantly extended with dsNanchung, dsInactive and dsWaterwitch treatments. These data suggest that both nompC and trpA5 are required for adult eclosion. The functions of nanchung and inactive are involved in the folding the hind leg segments. In addition, nanchung, inactive, and waterwitch are likely the receptors involved in recovery from tonic immobility.