Immunostaining for allatotropin and allatostatin-A and -C in the mosquitoes Aedes aegypti and Anopheles albimanus (original) (raw)
Related papers
Peptides, 2012
Allatotropin is an insect neuropeptide with pleiotropic actions on a variety of different tissues. In the present work we describe the identification, cloning and functional and molecular characterization of an Aedes aegypti allatotropin receptor (AeATr) and provide a detailed quantitative study of the expression of the AeATr gene in the adult mosquito. Analysis of the tissue distribution of AeATr mRNA in adult female revealed high transcript levels in the nervous system (brain, abdominal, thoracic and ventral ganglia), corpora allata-corpora cardiaca complex and ovary. The receptor is also expressed in heart, hindgut and male testis and accessory glands. Separation of the corpora allata (CA) and corpora cardiaca followed by analysis of gene expression in the isolated glands revealed expression of the AeATr primarily in the CA. In the female CA, the AeATr mRNA levels were low in the early pupae, started increasing 6 hours before adult eclosion and reached a maximum 24 hours after female emergence. Blood feeding resulted in a decrease in transcript levels. The pattern of changes of AeATr mRNA resembles the changes in JH biosynthesis. Fluorometric Imaging Plate Reader recordings of calcium transients in HEK293 cells expressing the AeATr showed a selective response to A. aegypti allatotropin stimulation in the low nanomolar concentration range. Our studies suggest that the AeATr play a role in the regulation of JH synthesis in mosquitoes.
Journal of Biological Chemistry, 2006
Aedes aegypti PISCF-allatostatin or allatostatin-C (Ae-AS-C) was isolated using a combination of high performance liquid chromatography and enzyme-linked immunosorbent assay (ELISA). The matrix-assisted laser desorption/ionization time-of-flight (TOF) mass spectrum of positive ELISA fractions revealed a molecular mass of 1919.0 Da, in agreement with the sequence qIRYRQCYFNPISCF, with bridged cysteines. This sequence was confirmed by matrix-assisted laser desorption/ionization tandem TOF/TOF mass spectrometry analysis. The corresponding Ae-AS-C cDNA was amplified by PCR, and the sequence of the peptide was confirmed.
Regulatory Peptides, 2004
We investigated the role of head factors and allatostatins (ASs) on the regulation of juvenile hormone (JH) synthesis in female adult mosquito. The biosynthetic activity of the Aedes aegypti corpora allata (CA) in vitro was inhibited by factors present in the head. Disconnecting the CA from the brain resulted in a significant increase in the rate of JH biosynthesis. Inhibition was not dependent on intact nervous connections; co-incubation of CA with brains or brain extracts resulted in a significant decrease of JH biosynthesis. This inhibitory effect of brain extracts was reversible and heat stable; extracts lost the inhibitory activity after proteinase K digestion suggesting a peptidic structure. In a first attempt to elucidate the nature of this inhibitory factor, we tested in our CA in vitro system the effect of members of two families of allatostatins already described in mosquitoes. Anopheles gambiae PISCF-allatostatin (homolog to Manduca PISCF-allatostatin) significantly inhibited JH synthesis, while Ae. aegypti YXFGL-amide-allatostatins (homologs to cockroach YXFGL-amide-allatostatins) did not affect JH synthesis. These results represent the first description of an allatostatic effect of PISCF-allatostatins outside the Lepidoptera. D
Peptides, 2003
Using a polyclonal antiserum to Dippu-allatostatin 7 (Dippu-AST 7; formerly AST 1) of the cockroach Diploptera punctata, we have demonstrated the presence of AST-like immunoreactivity (ALI) in cells and processes throughout the nervous system, gut, and peripheral tissues of unfed fifth instar and adult Rhodnius prolixus. ALI in apparent neurosecretory cells of the brain, suboesophageal ganglion, and mesothoracic ganglionic mass, as well as in midgut endocrine cells, suggests that Rhodnius allatostatins may act as neurohormones/hormones. The presence of ALI in possible interneurons and areas of neuropile throughout the CNS also suggests roles as neuromodulators and/or neurotransmitters. Dippu-AST 7 inhibits spontaneous and leucokinin 1 (LK 1)-induced contractions of the Rhodnius hindgut in a dose-dependent manner. The low concentrations capable of inhibiting both spontaneous (10 −12 M) and LK 1-induced contractions (10 −10 to 10 −9 M) suggest that ASTs may be acting as neurohormones/hormones on the hindgut. We have also shown that Dippu-AST 7 influences the muscle activity of the Rhodnius dorsal vessel at concentrations as low as 10 −11 M.
Allatostatins: Diversity in Structure and Function of an Insect Neuropeptide Family
Annals of The New York Academy of Sciences, 1997
The juvenile hormones (JHs) are a unique group of sesquiterpenoids, identified definitively only in insects, that are responsible for the maintenance of juvenile characteristics. A reduction in JH titer is generally believed to be required for metamorphosis to the adult form. Reproductive functions in most adult female insect species are also regulated by JH, and oocyte growth and maturation, including vitellogenesis, show an absolute dependency on JH-in the absence of the hormone, oocyte growth is arrested.' JH titer is regulated in part by the rate of biosynthesis within the endocrine glands known as the corpora allata (CA), which can he considered analogous to the adenohypophysis of vertebrates. Based on the now classic experiments involving severance of nerve tracts originating in the brain and innervating the CA, Scharrer demonstrated that CA function was under close-range neural control. Scharrer suggested that signals in the hemolymph might also act to regulate the CA.2 Consistent with Scharrer's original observations, the regulation of JH biosynthesis by CA is now known to be stimulated or inhibited by two groups of peptides, allatotropins and allatostatins (ASTs), respectively. This review will examine the ASTs which occur in multiple forms, are pleiotropic in function, and are widely distributed in both neural and nonneural tissue. The structural and functional features of the ASTs appear to parallel the vertebrate somatostatins and may provide one of the best examples of the parallel evolution of peptides.* MOLECULAR ISOLATION AND CHARACTERIZATION OF THE COCKROACH ALLATOSTATIN PRECURSOR The initial characterization of the first complete AST coding region was accomplished by polymerase chain reaction (PCR) amplifications3 of specific sequence from cDNA derived from mKNA isolated from the brains of virgin females of the cockroach Diploptera p~n c t a t a .~ Initially, a pair of highly degenerate primers was used to produce an internal DNA consensus sequence representing the peptide sequence of Dip-AST2
Neuropeptide F and its expression in the yellow fever mosquito, Aedes aegypti
Peptides, 2002
A neuropeptide F (NPF) was isolated from an extract of adult Aedes aegypti mosquitoes based on its immunoreactivity in a radioimmunoassay for Drosophila NPF. After sequencing the peptide, cDNAs encoding the NPF were identified from head and midgut. These cDNAs encode a prepropeptide containing a 36 amino acid peptide with an amidated carboxyl terminus, and its sequence shows it to be a member of the neuropeptide F/Y superfamily. Immunocytochemistry and Northern blots confirmed that both the brain and midgut of females are likely sources of NPF, found at its highest hemolymph titer before and 24 h after a blood meal.
Archives of Insect Biochemistry and Physiology, 1998
Immunoreactivity against peptides of the allatostatin family having a typical YXFGL-NH 2 C-terminus has been localized in different areas of the central nervous system, stomatogastric nervous system and gut of the cockroach Blattella germanica. In the protocerebrum, the most characteristic immunoreactive perikarya are situated in the lateral and median neurosecretory cell groups. Immunoreactive median neurosecretory cells send their axons around the circumesophageal connectives to form arborizations in the anterior neuropil of the tritocerebrum. A group of cells in the lateral aspect of the tritocerebrum project to the antennal lobes in the deutocerebrum, where immunoreactive arborizations can be seen in the periphery of individual glomeruli. Nerve terminals were shown in the corpora allata. These terminals come from perikarya situated in the lateral neurosecretory cells in the pars lateralis and in the subesophageal ganglion. Immunoreactive axons from median neurosecretory cells and from cells positioned in the anteriormost part of the tritocerebrum enter together in the stomatogastric nervous system and innervate foregut and midgut, especially the crop and the valve between the crop and the midgut. The hindgut is innervated by neurons whose perikarya are located in the last abdominal ganglion. Besides immunoreactivity in neurons, allatostatin-immunoreactive material is present in endocrine cells distributed within the whole midgut epithelium. Possible functions for these peptides according to their localization are discussed. Arch. Insect Biochem. Physiol. 37:269-282, 1998.
Cell and Tissue Research, 1995
Methanolic brain extracts of Locusta migratoria inhibit in vitro juvenile hormone biosynthesis in both the locust L. migratoria and the cockroach Diploptera punctata. A polyclonal antibody against allatostatin-5 (AST-5) (dipstatin-2) of this cockroach was used to immunolocalize allatostatin-5-1ike peptides in the central nervous system of the locusts Schistocerca gregaria and L. migratoria and of the fleshfly Neobellieria bullata. In both locust species, immunoreactivity was found in many cells and axons of the brain-retrocerebral complex, the thoracic and the abdominal ganglia. Strongly immunoreactive cells were stained in the pars lateralis of the brain with axons (NCC II and NCA I) extending to and arborizing in the corpus cardiacum and the corpora allata. Although many neurosecretory cells of the pars intercerebralis project into the corpus cardiacum, only 12 of them were immunoreactive and the nervi corporis cardiaci I (NCC I) and fibers in the nervi corporis allati II (NCA II) connecting the corpora allata to the suboesophageal ganglion remained unstained. S. gregaria and L. migratoria seem to have an allatostatin-like neuropeptide present in axons of the NCC II and the NCA I leading to the corpus cardiacum and the corpora allata. All these data suggest that in locusts allatostatinlike neuropeptides might be involved in controlling the production of juvenile hormone by the corpora allata and, perhaps, some aspects of the functioning of the corpus cardiacum as well. However, when tested in a L. migratoria in-vitro juvenile hormone-biosynthesis assay, allatostatin-5 did not yield an inhibitory or stimulatory effect. There is abundant AST-5 immunoreactivity in cell bodies of the fleshfly N. bullata, but none in the CA-CC complexes. Apparently, factors that are immunologically related to AST-5 do occur in locusts and fleshflies but, the active portion of the peptide required to inhibit: JH biosynthesis in locusts is probably different from that of AST-5.