Characterization of the Adipokinetic Hormone/Corazonin-Related Peptide Signalling System in the Mosquito, Aedes Aegypti

The adipokinetic hormone/corazonin-related peptide (ACP) is an insect neuropeptide structurally intermediate between corazonin (CRZ) and adipokinetic (AKH) hormones, which all demonstrate homology to the vertebrate gonadotropin-releasing hormone (GnRH). AKH and CRZ are best known to function in energy mobilization and cardioacceleration, respectively, however, the function of the ACP signalling system remains unclear. Here the gene encoding the ACP receptor in Aedes aegypti has been identified. Functional deorphanization of AedaeACPR-I revealed a highly specific response for its native ligand, ACP. Analysis of spatial and developmental expression profiles reveals enrichment of ACP and ACPR in the central nervous system and post-eclosion, respectively. The cell-specific distribution of the ACP and ACPR within the central nervous system was examined revealing expression within distinct regions of the brain, thoracic ganglia, and abdominal ganglia. The findings of this thesis point to a role of ACP within the nervous system functioning either as a neuromodulator or neurotransmitter

Insight into mosquito GnRH-related neuropeptide receptor specificity revealed through analysis of naturally occurring and synthetic analogs of this neuropeptide family

Adipokinetic hormone (AKH), corazonin (CRZ), and the AKH/CRZ-related peptide (ACP) are neuropeptides considered homologous to the vertebrate gonadotropin-releasing hormone (GnRH). All three Aedes aegypti GnRH-related neuropeptide receptors have been characterized and functionally deorphanized. Individually they exhibit high specificity for their native ligands, prompting us to investigate the contribution of ligand structures in conferring receptor specificity for two of these receptors. Here, we designed a series of analogs based on the native ACP sequence and screened them using a heterologous system to identify critical residues required for ACP receptor (ACPR) activation. Analogs lacking the carboxy-terminal amidation, replacing aromatics, as well as truncated analogs were either completely inactive or had very low activities on ACPR. The polar threonine (position 3) and the blocked amino-terminal pyroglutamate are also critical, whereas ACP analogs with alanine substitutions at position 2 (valine), 5 (serine), 6 (arginine), and 7 (aspartate) were less detrimental including the substitution of charged residues. Replacing asparagine (position 9) with an alanine resulted in a 5-fold more active analog. A naturally-occurring ACP analog, with a conserved substitution in position two, was well tolerated yet displayed significantly reduced activity compared to the native mosquito ACP peptide. Chain length contributes to ligand selectivity in this system, since the endogenous octapeptide Aedae-AKH does not activate the ACPR whereas AKH decapeptides show low albeit significant activity. Similarly, we utilized this in vitro heterologous assay approach against an A. aegypti AKH receptor (AKHR-IA) testing carefully selected naturally-occurring AKH analogs from other insects to determine how substitutions of specific residues in the AKH ligand influence AKHR-IA activation. AKH analogs having single substitutions compared to Aedae-AKH revealed position 7 (either serine or asparagine) was well tolerated or had slightly improved activation whereas changes to position 6 (proline) compromised receptor activation by nearly 10-fold. Substitution of position 3 (threonine) or analogs with combinations of substitutions were quite detrimental with a significant decrease in AKHR-IA activation. Collectively, these results advance our understanding of how two GnRH-related systems in A. aegypti sharing the most recent evolutionary origin sustain independence of function and signaling despite their relatively high degree of ligand and receptor homology.York University Librarie

Transcriptomic analyses reveal neuronal specificity of Leigh syndrome associated genes

Leigh syndrome is a rare inherited, complex, neurometabolic disorder with genetic and clinical heterogeneity. Features present in affected patients range from classical stepwise developmental regression to ataxia, seizures, tremor, and occasionally psychiatric manifestations. Currently, more than 100 monogenic causes of Leigh syndrome have been identified, yet, the pathophysiology remains unknown. Here, we sought to determine the cellular specificity within the brain of all genes currently associated with Leigh syndrome. Further, we aimed to investigate potential genetic commonalities between Leigh syndrome and other disorders with overlapping clinical features. Enrichment of our target genes within the brain was evaluated with co-expression (CoExp) network analyses constructed using existing UK Brain Expression Consortium data. To determine the cellular specificity of the Leigh associated genes, we employed expression weighted cell type enrichment (EWCE) analysis of single cell RNA-Seq data. Finally, CoExp network modules demonstrating enrichment of Leigh syndrome associated genes were then utilised for synaptic gene ontology analysis and heritability analysis. CoExp network analyses revealed that Leigh syndrome associated genes exhibit the highest levels of expression in brain regions most affected on MRI in affected patients. EWCE revealed significant enrichment of target genes in hippocampal and somatosensory pyramidal neurons and interneurons of the brain. Analysis of CoExp modules enriched with our target genes revealed preferential association with pre-synaptic structures. Heritability studies suggested some common enrichment between Leigh syndrome and Parkinson disease and epilepsy. Our findings suggest a primary mitochondrial dysfunction as the underlying basis of Leigh syndrome with associated genes primarily expressed in neuronal cells

CAPA neuropeptides and their receptor form an anti-diuretic hormone signalling system in the human disease vector, Aedes aegypti

Insect CAPA neuropeptides are homologs of mammalian neuromedin U and are known to influence ion and water balance by regulating the activity of the Malpighian ‘renal’ tubules (MTs). Several diuretic hormones are known to increase primary fluid and ion secretion by insect MTs and, in adult female mosquitoes, a calcitonin-related peptide (DH31) called mosquito natriuretic peptide, increases sodium secretion to compensate for the excess salt load acquired during blood-feeding. An endogenous mosquito anti-diuretic hormone was recently described, having potent inhibitory activity against select diuretic hormones, including DH31. Herein, we functionally deorphanized, both in vitro and in vivo, a mosquito anti-diuretic hormone receptor (AedaeADHr) with expression analysis indicating highest enrichment in the MTs where it is localized within principal cells. Characterization using a heterologous in vitro system demonstrated the receptor was highly sensitive to mosquito CAPA neuropeptides while in vivo, AedaeADHr knockdown abolished CAPA-induced anti-diuretic control of DH31-stimulated MTs. CAPA neuropeptides are produced within a pair of neurosecretory cells in each of the abdominal ganglia, whose axonal projections innervate the abdominal neurohaemal organs, where these neurohormones are released into circulation. Lastly, pharmacological inhibition of nitric oxide synthase (NOS) and protein kinase G (PKG) signaling eliminated anti-diuretic activity of CAPA, highlighting the role of the second messenger cGMP and NOS/PKG in this anti-diuretic signaling pathway.York University Librarie

Disability training for health workers: A global evidence synthesis.

Background Health worker training on disability is a recognized component of achieving high standards of health for people with disabilities, given that health worker’s lack of knowledge, stigma, and negative attitudes towards people with disabilities act as barriers to high quality health care. Objective To understand the published literature on training health workers about disability. Methods We searched five databases for relevant peer-reviewed articles published between January 2012 and January 2021. Studies that focused on training health care workers to improve knowledge, confidence, self-efficacy, and competence to support people with physical, sensory, or intellectual impairments were included. Data about the details of the intervention (setting, participants, format, impact assessments, etc.) and its effects were extracted. Results There is an array of highly local tools to train health workers across stages of their training and careers (preservice, in-service, and continuing professional development). Studies involving people with disabilities in the training, community placements, simulations, or interactive sessions were found to be most effective in improving knowledge, confidence, competency, and self-efficacy. Conclusions As part of initiatives to build inclusive health systems and improve health outcomes for people with disabilities, health workers around the world need to receive appropriate and evidence-based training that combines multiple methods and involves people with disabilities. To monitor progress on the impact of training, there should also be a standardized measure of impact on core outcomes