Double-labelled in situ Hybridization Reveals the Lack of Co-localization of mRNAs for the Circadian Neuropeptide PDF and FMRFamide in Brains of the Flies Musca domestica and Drosophila melanogaster

Abstract

Many lines of evidence have suggested that neuropeptides other than pigment-dispersing factor (PDF) are involved in regulating insect circadian rhythms, and FMRFamide-related peptides are additional candidates acting as such neuromodulators. Double-immunolabelling in insect brains with anti-crustacean beta-PDH and anti-FMRFamide antibodies had previously suggested that insect PDF and FMRFamide-like peptides may coexist in the same cells. However, it is critical for this kind of comparative investigations to use antibodies of proven specificity, to eliminate the possibility of both reciprocal cross-reactivity and the detection of unknown peptides. In the present study, we achieved the cDNA cloning of an fmrf mRNA from the housefly Musca domestica, for which co-localization of FMRFamide and PDF peptides was previously suggested. In order to examine the possible co-expression of this gene with the pdf gene, we carried out double-labelled in situ hybridization for simultaneous detection of both pdf and fmrf mRNAs in housefly, Musca brains. The results clearly indicated that they occur in distinctly different cells. This was also proven for the fruit fly Drosophila melanogaster by similar double-labelled in situ hybridization. The results thus revealed no reason to evoke the physiological release of FMRFamide and PDF peptides from the same neurons.

FMRFamide gene and peptide expression during central nervous system development of the cephalopod mollusk, Idiosepius notoides

“…However, characterizing neurons of the cephalopod brain by their transmitter phenotype and molecular fingerprinting might allow homologizing distinct neuronal populations or brain regions, a step that has already been undertaken for the simpler CNS of gastropods and other lophotrochozoans (e.g., McAllister et al 1983;Voronezhskaya and Elekes 2003;Denes et al 2007). Accordingly, the present work is a further step (i) toward an explanation of the physiological significance of the diversity and multiplicity of FaRPs in cephalopods, as it has already been undertaken for other metazoans (Matsushima et al 2007), and (ii) for inferences of neuronal homologies between the various molluscan clades.…”

Signalling through pigment dispersing hormone-like peptides in invertebrates

“…More recent findings contradict the presence of FMRFamide in PDF-neurons, at least for the fruit fly and the housefly. As these experiments were conducted in great detail, it is assumable that FMRF-neurons and PDF-neurons comprise separate, though closly associated neuron sets (Matsushima et al, 2007).…”