Biology subjects

Sajadi, F.

Publications and source records attributed to Sajadi, F..

5 recordsLinked to original sources

bioRxiv2025

Dynamics of neuropeptide release and activity post-bloodmeal in the female mosquito, Aedes aegypti

Female Aedes aegypti secrete urine rapidly post-bloodmeal ingestion, with diuresis beginning immediately for removal of excess salts and water. This post-prandial diuresis includes a peak, post-peak, and late phase, involving the combined actions of multiple hormones, including diuretic and anti-diuretic factors. Calcitonin-like diuretic hormone 31 (DH31) and kinin peptides stimulate diuresis through actions on their cognate receptors localized in the Malpighian renal tubules (MTs). In contrast, the anti-diuretic neurohormone, CAPA, inhibits secretion by MTs stimulated by select diuretic hormones, including DH31. While DH31 and kinin are critical in achieving post-prandial diuresis, and CAPA functions as an important anti-diuretic hormone, the kinetics of their release and haemolymph levels remain unknown. Herein, using heterologously expressed A. aegypti DH31, CAPA, and kinin receptors, we investigated the titres of these hormones in the haemolymph of female mosquitoes at different time points after blood feeding. Haemolymph extracts from female mosquitoes contained levels of diuretic peptides, specifically kinin and DH31, that increased immediately post-bloodmeal, with levels peaking at 2 and 5 min, respectively, while DH31 levels remained elevated for 15 min. Comparatively, levels of CAPA peptides in the haemolymph steadily increase 15 min post-blood feeding, with levels peaking at 30 min. Synergistic actions were observed between DH31 and a kinin-like peptides on the MTs, providing a physiological context for the rapid release of these peptides into the female haemolymph. Altogether, these results demonstrate that DH31 and kinin are released immediately post-bloodmeal and, along with CAPA peptides, have a coordinative action on the MTs to maintain haemolymph homeostasis through regulation of primary urine secretion. Summary statementThis study characterizes neuropeptidergic regulators of the Malpighian renal tubules by quantifying their circulating titres and release post-blood feeding in the female mosquito, A. aegypti.

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bioRxiv2024

Anti-diuretic hormone ITP signals via a guanylate cyclase receptor to modulate systemic homeostasis in Drosophila

Insects have evolved a variety of neurohormones that enable them to maintain their nutrient and osmotic homeostasis. While the identities and functions of various insect metabolic and diuretic hormones have been well-established, the characterization of an anti-diuretic signaling system that is conserved across most insects is still lacking. To address this, here we characterized the ion transport peptide (ITP) signaling system in Drosophila. The Drosophila ITP gene encodes five transcript variants which generate three different peptide isoforms: ITP amidated (ITPa) and two ITP-like (ITPL1 and ITPL2) isoforms. Using a combination of anatomical mapping and single-cell transcriptome analyses, we comprehensively characterized the expression of all three ITP isoforms in the nervous system and peripheral tissues. Our analyses reveal wide-spread expression of ITP isoforms. Moreover, we show that ITPa-producing neurons are activated and release ITPa during dehydration. Further, recombinant Drosophila ITPa inhibits diuretic peptide-induced renal tubule secretion ex vivo, thus confirming its role as an anti-diuretic hormone. Using a phylogenetic-driven approach, an ex vivo secretion assay and a heterologous mammalian cell-based assay, we identified and functionally characterized Gyc76C, a membrane guanylate cyclase, as a bona fide Drosophila ITPa receptor. Thus, recombinant ITPa application leads to increased cGMP production in HEK293T cells expressing Drosophila Gyc76C. Moreover, knockdown of Gyc76C in renal tubules abolishes the inhibitory effect of ITPa on diuretic hormone stimulated secretion. Extensive anatomical mapping of Gyc76C reveals that it is highly expressed in larval and adult tissues associated with osmoregulation (renal tubules and rectum) and metabolic homeostasis (fat body). Consistent with this expression, knockdown of Gyc76C in renal tubules impacts tolerance to osmotic and ionic stresses, whereas knockdown specifically in the fat body impacts feeding, nutrient homeostasis and associated behaviors. We also complement receptor knockdown experiments with ITP knockdown and ITPa overexpression in ITPa-producing neurons. Interestingly, the ITPa-Gyc76C pathway examined here is reminiscent of the atrial natriuretic peptide signaling in mammals. Lastly, we utilized connectomics and single-cell transcriptomics to identify synaptic and paracrine pathways upstream and downstream of ITPa-expressing neurons. Our analysis identifies pathways via which ITP neurons integrate hygrosensory inputs and interact with other homeostatic hormonal pathways. Taken together, our systematic characterization of ITP signaling establishes a tractable system to decipher how a small set of neurons integrates diverse inputs to orchestrate systemic homeostasis in Drosophila.

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bioRxiv2024

Molecular characterization, localization, and physiological roles of ITP and ITP-L in the mosquito, Aedes aegypti

The insect ion transport peptide (ITP) and its alternatively spliced variant, ITP-like peptide (ITP-L), belong to the crustacean hyperglycemic hormone family of peptides and are widely conserved among insect species. While limited, studies have characterized the ITP/ITP-L signaling system within insects, and putative functions including regulation of ion and fluid transport, ovarian maturation, and thirst/excretion have been proposed. Herein, we aimed to molecularly investigate Itp and Itp-l expression profiles in the mosquito Aedes aegypti, examine peptide immunolocalization and distribution within the adult central nervous system, and elucidate physiological roles for these neuropeptides. Transcript expression profiles of both AedaeItp and AedaeItp-l revealed distinct enrichment patterns in adults, with AedaeItp expressed in the brain and AedaeItp-l expression predominantly within the abdominal ganglia. Immunohistochemical analysis within the central nervous system revealed expression of AedaeITP peptide in a number of cells in the brain and in the terminal ganglion. Comparatively, AedaeITP-L peptide was localized solely within the pre-terminal abdominal ganglia of the central nervous system. Interestingly, prolonged desiccation stress caused upregulation of AedaeItp and AedaeItp-l levels in adult mosquitoes, suggesting possible functional roles in water conservation and feeding-related activities. RNAi-mediated knockdown of AedaeItp caused an increase in urine excretion, while knockdown of both AedaeItp and AedaeItp-l reduced blood feeding and egg-laying in females as well as hindered egg viability, suggesting roles in reproductive physiology and behaviour. Altogether, this study identifies AedaeITP and AedaeITP-L as key pleiotropic hormones, regulating various critical physiological processes in the disease vector, A. aegypti.

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bioRxiv2022

The V-type H+-ATPase is targeted in anti-diuretic hormone control of the Malpighian (renal) tubules in Aedes aegypti

Like other insects, secretion by mosquito Malpighian tubules (MTs) is driven by the V-type H+-ATPase (VA) localized in the apical membrane of principal cells. In Aedes aegypti, the anti-diuretic neurohormone CAPA inhibits secretion by MTs stimulated by select diuretic hormones; however, the cellular effectors of this inhibitory signaling cascade remain unclear. Herein, we demonstrate that the VA inhibitor bafilomycin selectively inhibits serotonin (5HT)- and calcitonin-related diuretic hormone (DH31)-stimulated secretion. VA activity increases in DH31-treated MTs, whereas CAPA abolishes this increase through a NOS/cGMP/PKG signaling pathway. A critical feature of VA activation involves the reversible association of the cytosolic (V1) and membrane (Vo) complexes. Indeed, higher V1 protein abundance was found in membrane fractions of DH31-treated MTs whereas CAPA significantly decreased V1 abundance in membrane fractions while increasing it in cytosolic fractions. V1 immunolocalization was observed strictly in the apical membrane of DH31 treated MTs whereas immunoreactivity was dispersed following CAPA treatment. VA complexes colocalized apically in female MTs shortly after a blood-meal consistent with the peak and post-peak phases of diuresis. Comparatively, V1 immunoreactivity in MTs was more dispersed and did not colocalize with the Vo complex in the apical membrane at 3 hours post blood-meal, representing a timepoint after the late phase of diuresis has concluded. Therefore, CAPA inhibition of MTs involves reducing VA activity and promotes complex dissociation hindering secretion. Collectively, these findings reveal a key target in hormone-mediated inhibition of MTs countering diuresis that provides a deeper understanding of this critical physiological process necessary for hydromineral balance. Significance StatementThe V-type H+ ATPase (VA), or proton pump, provides the driving force for transepithelial ion and fluid secretion in insect Malpighian tubules (MTs). While studies have shown diuretic stimulation activates various signaling pathways, including cAMP and downstream effectors promoting increased VA activity, our understanding of anti-diuretic signaling and its potential regulation of the VA remains rudimentary. Herein, we show that CAPA neuropeptide acts through the NOS/cGMP/PKG pathway to inhibit DH31-stimulated VA activity, supporting the notion that the anti-diuretic regulation is achieved through dissociation of the VA complexes. These results demonstrate a critical role of VA inhibition and trafficking necessary for anti-diuretic signaling and advances our understanding of the complex neuroendocrine control of the MTs in this important human disease-vector mosquito.

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bioRxiv2019

An anti-diuretic hormone receptor in the human disease vector, Aedes aegypti: identification, expression analysis and functional deorphanization

Insect CAPA neuropeptides, which are homologs of mammalian neuromedin U, have been described in various insect species and are known to influence ion and water balance by regulating the activity of the Malpighian renal tubules (MTs). A number of diuretic hormones have been shown to increase primary fluid and ion secretion by the insect MTs and, in the adult female mosquito, a calcitonin-related peptide (DH31) also known as mosquito natriuretic peptide, increases sodium secretion at the expense of potassium to remove the excess salt load acquired upon blood-feeding. An endogenous mosquito anti-diuretic hormone was recently described, having inhibitory activity against select diuretic factors and being particularly potent against DH31-stimulated diuresis. In the present study, we have functionally deorphanized, both in vitro and in vivo, a mosquito anti-diuretic hormone receptor (AedaeADHr). Expression analysis by quantitative PCR indicates the receptor is highly enriched in the MTs, and fluorescent in situ hybridization confirms expression within principal cells. Characterization using a heterologous system demonstrated the receptor was highly sensitive to mosquito CAPA peptides. In adult females, AedaeADHr transcript knockdown using RNAi led to the abolishment of CAPA-peptide induced anti-diuretic control of DH31-stimulated MTs. The neuropeptidergic ligand is produced within a pair of neurosecretory cells in each of the six abdominal ganglia, whose axonal projections innervate the abdominal neurohaemal organs (known as the perivisceral organs), where these neurohormones are released into the open circulatory system of the insect. Furthermore, pharmacological inhibition of PKG/NOS signalling abolished the anti-diuretic activity of AedaeCAPA-1, which collectively confirms the role of cGMP/PKG/NOS in this anti-diuretic signalling pathway.\n\nSignificanceInsects are by far the most successful and abundant group of organisms on earth. As a result of their small size, insects have a relatively large surface area to volume ratio, raising the potential for rapid gain or loss of water, ions and other molecules including toxins - a phenomenon that applies to insects living in both aquatic and terrestrial environments. In common with many other organisms, hormones are key regulators of the excretory system in insects, and numerous factors control the clearance of excess water and ions (i.e. diuretics) or retention of these elements (i.e. anti-diuretics). Here we characterized an endogenous anti-diuretic hormone receptor in the human disease vector, Aedes aegypti, demonstrating its expression is highly enriched in the Malpighian renal tubules and is necessary for eliciting anti-diuretic control of this key component of the mosquito excretory system.

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