Identical cellular distribution of all abundant neuropeptides in the major abdominal neurohemal system of an insect (Periplaneta Americana)

Abstract

The median neurosecretory cells in abdominal ganglia of insects synthesize a number of putative hormones, which are abundant in the abdominal perisympathetic organs (PSOs). The peptide inventory of these prominent neurohemal release sites is best investigated in the American cockroach and strongly differs from that of head/thoracic neurohemal organs. In this study, we found a complete colocalization of all abundant neuropeptides in this hormonal system, including periviscerokinin-1 and -2, pyrokinin-5, YLSamide, VEAacid, and SKNacid. The first immunoreactive cells were detected on day 18 of embryonic development and already contained the complete set of peptides. By using antisera against the above-mentioned peptides, the development of this neurohormonal system could be studied and is described in detail. Subsequent electron microscopic immunogold stainings in PSO preparations revealed the costorage of PSO peptides in a single vesicle species. Surprisingly, all these peptides were found in axons containing clear vesicles, whereas all axons with dense core vesicles were totally devoid of immunoreactivity. Unlike the axons with dense core vesicles, immunostained axons ramify in the center of the PSO but exhibit only rare morphological signs of exocytosis. Instead, putative release sites of the clear vesicle-containing axons were detected peripherally to the PSOs, namely, on the hyperneural muscle.

Peptidomics of identified neurons demonstrates a highly differentiated expression pattern of FXPRLamides in the neuroendocrine system of an insect

“…Immunolabelings with the aPK-5 serum did not show any neurosecretory cells in the brain/ SEG; PK-5-immunoreactive interneurons have been described by Eckert et al (2002).…”

“…Some slices were used for normal ultrastructural observations. On the other grids, peptide immunogold reactions were performed as described by Eckert et al (2002). Finally, grids were rinsed in distilled water and counterstained with uranyl acetate and lead citrate.…”

Differential sorting and packaging of capa-gene related products in an insect

“…For double labeling, the same treatment was repeated on the other face of the grid. A detailed description of the procedure of triple labeling is given in Eckert et al (2002).…”

“…Antisera were produced, purified, and labeled as described by Eckert et al (2002). Briefly, anti-PVK-2, antiPea-PK-5, and anti-YLSamide sera, which cross-reacted with pyrokinins, were purified by affinity chromatography using CNBR-activated Sepharose 4B (Pharmacia Biotech, Freiburg, Germany) coupled to a Pea-PK-2 (SPPFAPRLa) thyroglobulin conjugate.…”

“…Postembedding immunoreactions were carried out as described in Eckert et al (2002). Briefly, after a short etching and deosmication, grids were thoroughly washed, pretreated with normal goat serum to cover aspecific binding sites, and transferred to drops with primary antibodies (anti-Pea-PVK-1, anti-Pea-PVK-2, anti-Pea-PK-5, anti-Pea-SKN-OH, anti-Pea-VEA-OH, anti-PeaYLSamide) diluted in TBS (Tris-buffered saline, pH 7.6, overnight reaction).…”