title: 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.
authors.affiliation: Laboratory of Structure-Function Biochemistry, Department of Chemistry, Faculty and Graduate School of Sciences, Kyushu University, Fukuoka 812-8581, Japan.
authors.forename: Ayami
authors.initials: A
authors.lastname: Matsushima
authors.forename: Katsuhiro
authors.initials: K
authors.lastname: Takano
authors.forename: Taichi
authors.initials: T
authors.lastname: Yoshida
authors.forename: Yukimasa
authors.initials: Y
authors.lastname: Takeda
authors.forename: Satoru
authors.initials: S
authors.lastname: Yokotani
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Shimohigashi
authors.forename: Miki
authors.initials: M
authors.lastname: Shimohigashi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000378:metabolism*; D002940:Circadian Rhythm; D029721:Drosophila Proteins / Q000494:pharmacology*; D004331:Drosophila melanogaster; D019835:FMRFamide / Q000494:pharmacology*; D005786:Gene Expression Regulation; D006793:Houseflies; D017403:In Situ Hybridization / Q000379:methods*; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D029721:Drosophila Proteins; D009479:Neuropeptides; D012333:RNA, Messenger; C113883:pdf protein, Drosophila; D019835:FMRFamide