Elsevier

Peptides

Volume 22, Issue 2, February 2001, Pages 199-208
Peptides

Myoinhibitory neuropeptides in the American cockroach☆

☆ Taken from a paper presented at the Winter Neuropeptide Conference 2000, Invertebrate Division, Hua Hin, Thailand, January 10–15, 2000.
https://doi.org/10.1016/S0196-9781(00)00383-1Get rights and content

Abstract

A large number of myostimulatory neuropeptides from neurohaemal organs of the American cockroach have been described since 1989. These peptides, isolated from the retrocerebral complex and abdominal perisympathetic organs, are thought to be released as hormones. To study the coordinated action of these neuropeptides in the regulation of visceral muscle activity, it might be necessary to include myoinhibitors as well, however, not a single myoinhibitory neuropeptide of the American cockroach has been described so far. To fill this gap, we describe the isolation of LMS (leucomyosuppressin) and Pea-MIP (myoinhibitory peptide) from neurohaemal organs of the American cockroach. LMS was very effective in inhibiting phasic activity of all visceral muscles tested. It was found in the corpora cardiaca of different species of cockroaches, as well as in related insect groups, including mantids and termites. Pea-MIP which is strongly accumulated in the corpora cardiaca was not detected with a muscle bioassay system but when searching for tryptophane-containing peptides using a diode-array detector. This peptide caused only a moderate inhibition in visceral muscle assays. The distribution of Pea-MIP in neurohaemal organs and cells supplying these organs with Pea-MIP immunoreactive material, is described. Additionally to LMS and Pea-MIP, a member of the allatostatin peptide family, known to exhibit inhibitory properties in other insects, was tested in visceral muscle assays. This allatostatin was highly effective in inhibiting spontaneous activity of the foregut, but not of other tested visceral muscles of the American cockroach.

Introduction

The neuropeptide pattern in neurohaemal organs of the American cockroach, Periplaneta americana, was intensively investigated in the last few years. Hereby, we concentrated on myotropic neuropeptides of the retrocerebral complex and abdominal perisympathetic organs which are the major neurohaemal release sites of the brain/subesophageal ganglion and ventral nerve cord, respectively. Two general conclusions can be drawn from these studies. Firstly, members of all myostimulatory neuropeptide families which were initially isolated from head extracts of Leucophaea maderae [12] are also present in considerable amounts in the corpora cardiaca/allata complex (CC/CA) of the American cockroach. This fact supports the idea that these peptides will be released into the circulation. Secondly, the abdominal perisympathetic organs (PSOs) were found to contain a neuropeptide pattern totally different from that of the retrocerebral complex. Altogether, not less than five different neuropeptide families with myostimulatory properties were identified from neurohaemal organs of the American cockroach, namely kinins, sulfakinins, pyrokinins, corazonin, and periviscerokinins [27].

A detailed knowledge of the structures and the distribution of myotropins in neurohaemal organs of P. americana makes it possible to investigate fluctuations in the amount of these peptides during development and certain physiological events. Muscle activity, however, can also be influenced by myoinhibitory neurohormones. For an understanding of the coordinated action of myotropic neurohormones it is therefore necessary to include myostimulatory peptides, as well as myoinhibitors. No myoinhibitory peptides have been reported from the American cockroach thus far. To fill this gap, we isolated and identified myoinhibitors from neurohaemal organs of P. americana. These putative hormones include leucomyosuppressin (LMS), initially isolated from L. maderae [11] and a novel member of myoinhibitory peptides (Pea-MIP). The efficacy of the different myoinhibitors of the American cockroach in visceral muscle assays is compared. During these experiments we also tested allatostatins which were known to inhibit spontaneous activity of the foregut of Leucophaea maderae [6].

Section snippets

Animals

Cockroaches were reared under a 12-h light, 12-h dark photoperiod at a constant temperature of 28°C. Only adult males four to six weeks after their last moult were used throughout the experiments. They were fed rat food pellets and had free access to water.

Bioassays

The assays on the isolated visceral muscles were performed as previously described [20]. The insect saline (pH 7.25) contained NaCl (8.19 g/liter), KCl (0.37 g/liter), CaCl2 (0.56 g/liter), MgCl2 × 6H2O (0.2 g/liter), glucose (0.9 g/liter)

Isolation and structural elucidation of leucomyosuppressin

An extract of 800 CC/CA was chromatographed on an Alphasil C18 column with TFA as organic modifier. The resulting fractions between 10 and 50 min were collected manually and then tested for myotropic activity on different visceral muscles, namely the hyperneural muscle, heart, and hindgut. The purification of peptides with myostimulatory properties has been described elsewhere (see 27). Only one fraction (Fig. 1 ) clearly inhibited the activity of isolated heart and hindgut. This fraction was

Discussion

The only myoinhibitor, which was highly effective in all visceral muscle assays used during this study, was LMS. Action, biologically active core sequence, and distribution of this peptide was extensively investigated in blaberoid cockroaches [1], [3], [5], [9], [16], [17], [18]. LMS seems to be conservative not only in its function as myoinhibitor in insects but also in its sequence which is obviously highly conserved in cockroaches and related insect orders. These properties resemble those of

Acknowledgements

We thank Mrs. Virginia Johnson (Protein Technologies Laboratory, Texas A & M University, College Station) for excellent sequence analysis; Mrs. Renate Winkler, Mrs. Erika Krause and Mrs. Ch. Raue (Institut für Allgemeine Zoologie und Tierphysiologie Jena, Germany) for perfect technical assistance. This study was supported with a post-doc fellowship of the German Academy of Science Leopoldina (LPD 1997, to RP) and grants from Deutsche Forschungsgemeinschaft (Predel 595/1–1,2; Eckert 122/4–1).

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