In situ hybridization analysis of leucomyosuppressin mRNA expression in the cockroach,Diploptera punctata

Abstract

In the cockroach Diploptera punctata, sequencing of the cDNA for the insect myoinhibitory neuropeptide, leucomyosuppressin (LMS), has demonstrated that LMS is the only Phe-Met-Arg-Phe-amide (NH2) (FMRFamide)-related peptide to be encoded by this gene (Donly et al. [1996] Insect Biochem. Mol. Biol. 26:627-637). However, in the present study, high performance liquid chromatography analysis of brain extracts showed six discrete FMRFamide-like immunoreactive fractions, one of which co-eluted with LMS. This study compared the distribution of FMRFamide-related peptides visualized by immunohistochemistry with LMS mRNA expression demonstrated by in situ hybridization in D. punctata. Immunohistochemistry with a polyclonal antiserum generated against FMRFamide, but which recognizes extended RFamide peptides, demonstrated numerous RFamide-like immunoreactive cells and processes in both nervous and nonnervous tissues. RFamide-like immunoreactivity was found in cells and processes of the brain and optic lobes, the stomatogastric nervous system, including the frontal and ingluvial ganglia, and the suboesophageal ganglion. Immunoreactivity was also present in all ganglia of the ventral nerve cord and in the alimentary canal. Within the alimentary canal, positively stained processes were found in the crop, midgut, and hindgut, and immunoreactive endocrinelike cells were located in the midgut. In situ hybridization with a digoxigenin-labeled RNA probe spanning the entire LMS coding region showed cell bodies containing LMS mRNA in all ganglia studied, other than the ingluvial ganglion. Expression was most abundant in the brain and optic lobes and in the frontal and suboesophageal ganglia. LMS mRNA was also apparent, although less intensely, in all other ganglia of the ventral nerve cord. Within the alimentary canal, LMS mRNA-positive cells were only visible in the anterior portion of the midgut, in the endocrinelike cells. The appearance of LMS mRNA in the central nervous system, stomatogastric nervous system, and midgut suggests that LMS may play a central role in Diploptera and may be associated with feeding and digestion.

Examination of the role of FMRFamide-related peptides in the circadian clock of the cockroach Leucophaea maderae

“…In addition to possible roles as neuromodulators, a neurohormonal role of FaRPs in L. maderae is suggested from the presence of immunoreactive somata in the pars intercerebralis and stained fibers projecting toward the retrocerebral complex. This assumption is in accordance with reports of FaRPs in neurosecretory cells and the retrocerebral complex in other insects (Meola et al 1991;Donly et al 1996;East et al 1997;Fuse et al 1998;Nässel 2000Nässel , 2002. Furthermore, FaRPs have been detected in the hemolymph from where they can reach all organs and tissues of physiological interest (Elia et al 1993(Elia et al , 1995.…”

“…In Diploptera punctata, for example, the FMRFamide antiserum detects, in addition to five high-performance liquid-chromatographic fractions, the LMS-containing fraction. Therefore, the anti-FMRFamide antiserum probably recognizes most FaRPs in L. maderae (Fuse et al 1998). This assumption is confirmed by a comparison of anti-DMS immunoreactivity with antiFMRFamide immunoreactivity obtained with two different anti-FMRFamide antibodies (Petri et al 1995;Söhler et al 2007).…”

“…Several FaRPs have been identified in various cockroaches. They include a single myosuppressin (L. maderae: Holman et al 1986; Diploptera punctata: Donly et al 1996;Fuse et al 1998; Periplaneta americana: Predel et al 2001), two sulfakinins (L. maderae ; P. americana : Veenstra 1989), a single sNPF (head peptide, P. americana: Veenstra and Lambrou 1995), and 24 forms of the extended FMRFamide gene (P. americana: Predel et al 2004).…”

Regulatory peptides in fruit fly midgut

“…Sulfakinins are evolutionarily related to cholecystokinin and, like this peptide, induce satiety in several insect species (e.g., Wei et al 2000;Meyering-Vos and Müller 2007), suggesting their possibile expression in midgut endocrine cells. The absence of myosuppressin in Drosophila midgut seems peculiar as, in other insect species, myosuppressins have been found to be expressed in midgut endocrine cells (Fuse et al 1998;Kingan et al 1997;Lee et al 2002;Lu et al 2002). However, neither our antisera nor our transgenic flies have detected the expression of these genes in Drosophila midgut, and we have thus confirmed their absence from this tissue.…”

An unusual myosuppressin from the blood-feeding bugRhodnius prolixus

“…Recently, Rhopr-MS expression was also observed in the posterior midgut of R. prolixus but not in the crop (anterior midgut) or hindgut . In situ hybridization in M. sexta and D. punctata reveals myosuppressin expression in endocrine cells of the posterior midgut (Fusé et al, 1998;Lu et al, 2002).…”