Prothoracicotropic hormone of Rhodnius prolixus: partial characterization and rhythmic release of neuropeptides related to Bombyx PTTH and bombyxin

Summary

The brain-retrocerebral complex (br-complex) of Rhodnius prolixus was found both to contain and release neuropeptides related to Bombyx mori PTTH and bombyxin. A > 10 kDa peptide fraction obtained from extracts and incubation media of br-complex exhibited high steroidogenic activity on Rhodnius prothoracic glands and reacted with a Bombyx PTTH antibody on dot blots. The release from the Br-complex of this immunoreactive peptide fraction showed a daily rhythm: high release during the night and little on no release during the day. On Western blots, a single 68 kDa peptide in the > 10 kDa peptide fraction was recognized by the Bombyx PTTH antibody and was also released rhythmically during a day. This peptide was reduced to a doublet of about 17 kDa that retained immunoreactivity. Double immunoprecipitation of the > 10 kDa peptide fraction from brain media using the Bombyx PTTH antibody and agarose-bound secondary antibody removed the steroidogenic activity in this fraction; it also removed the 68 kDa peptide on Western blots. A bombyxin antiserum recognized a 3–5 kDa peptide in a <10 kDa peptide fraction; this peptide fraction was also released with a daily rhythm but possessed weak steroidogenic activity. The natural PTTH of Rhodnius, therefore, appears to be a 68 kDa peptide, possibly composed of several 17 kDa subunits, that is related to Bombyx PTTH.     

Haemolymph ecdysteroid titres at metamorphosis in the fruit fly Ceratitis capitata: multiple peaks not apparent in whole body extracts

Abstract. The titre profile of haemolymph ecdysteroids in the fruit fly Ceratitis capitata was constructed from determinations of haemolymph taken from single animals and compared with ecdysteroid levels in whole body extracts. Larvae were sampled throughout the period from the late third instar larva until eclosion and were synchronized using a conspicuous behavioural marker ('leaping') that occurs at the end of the wandering period. Extracts of the whole body exhibit two peaks of ecdysteroids, one associated with pupariation ('pupariation peak' of other authors) and the other with pharate adult development. However, the 'pupariation peak' corresponded with three temporally distinct peaks in the titre of haemolymph ecdysteroids. The first peak occurs at the time of head retraction (lh before formation of the white puparium) and may represent the initial response to release of prothoracicotropic hormone. The second peak occurs 2–3 h later at the onset of tanning and may be responsible for induction of dopa-decarboxylase. These peaks are brief ( c . 2h each), whereas the third is much longer (16 h) and occupies most of the 'pupariation peak'. The first two peaks have not been reported previously in any dipteran. Their existence illustrates that extremely brief pulses of ecdysteroids occur in vivo , apparently with profound developmental consequences. The fact that brief pulses in the haemolymph titre are not apparent in whole body extracts emphasizes that results obtained using the latter method must be interpreted with caution.