Faculty of Science

Permanent URI for this communityhttps://hdl.handle.net/10155/386

The Faculty of Science (FSCI) provides students with the tools needed to adapt to future developments in the scientific path of their choice. Areas of study include applied & industrial mathematics, chemistry, integrative neuroscience, forensic science, computer science and physics.

Browse

Browsing Faculty of Science by Subject "Acetylcholine"

Filter results by typing the first few letters
Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Characterization of the ACC-1 Family of cys-loop ligand gated ion channels from the parasitic nematode Haemonchus contortus.
    (2020-12-01) Habibi, Sarah A.; Forrester, Sean
    Nematode cys-loop ligand-gated ion channels (LGICs) have been extensively studied for decades because of their role in current anthelmintic action and potential as targets for future drugs. Many families of cys loop receptors have not yet been pharmacologically characterized in parasitic nematodes, and thus provides an opportunity for further exploration into their role in the nervous system of these pathogens, and relevance for anthelmintic action. The ACC-1 family of receptors is a group of inhibitory acetylcholinegated chloride channels that are unique to invertebrate species. Specifically, the ACCs have been identified in both free-living (Caenorhabditis elegans) and parasitic (Haemonchus contortus) nematodes. However, their pharmacological properties in H. contortus have yet to be explored. My PhD thesis focused on identifying and characterizing the role of the ACC-1 family of receptors in H. contortus, using molecular cloning, pharmacological characterization and an investigation of the ligand-binding pockets using site directed mutagenesis and computational protein modelling. I conducted an extensive pharmacological characterization of the following receptors, ACC-2, ACC-1/ACC-2, ACC-1/LGC-46, LGC-46, LGC-39/ACC-1, and LGC-40/ACC-1, and generated homology models for each with various ligands bound. The ACC receptors from H. contortus are sensitive to various cholinergic ligands, nicotinic acetylcholine receptor anthelmintics, classical antagonists, and minimally sensitive to nicotine. Although one particular subunit, ACC-1, does not form a functional channel alone, it does associate with other members of the ACC family, ACC-2 and LGC-46, as well the previously uncharacterized cholinergic receptors, LGC-39 and 40, to form unique heteromeric channels. In addition, it was found that a single point mutation of a phenylalanine residue to a tyrosine in Loop C of the ACC binding pocket results in a hypersensitive receptor. Finally, sequence analysis of the ACCs revealed that the characteristic tryptophan residue, which contributes to π-cationic stabilizing interactions with ligands in the binding pocket, is located in Loop C of these receptors, whereas this residue is commonly found in Loop B of mammalian nAChRs. Together, this dissertation provides the first characterization of these inhibitory ACh receptors in parasitic nematodes and further sheds light into cholinergic neurotransmission in H. contortus.
  • Thumbnail Image
    Item
    Identification and characterization novel of cystine-loop ligand- gated chloride channels from Dirofilaria immitis: pharmacological analysis and novel compound screening
    (2023-12-01) Varley, Sierra; Forrester, Sean
    Dirofilaria immitis, otherwise known as heartworm, is a parasite that infects the hearts of dogs and causes serious health consequences. While preventative treatments are available drug resistance is developing at an alarming rate. Cystine-loop ligand-gated ion channels are important receptors in nematode neurobiology, and as such are promising drug targets. The UNC-49 (GABA-gated chloride channel) and ACC (acetylcholine-gated chloride channel) family of receptors have been characterized as potential drug targets in other nematodes. However, these receptors have yet to be identified or characterized in D. immitis. This thesis investigates the cloning and pharmacological characterization of 5 novel receptor subunits from D. immitis: UNC-49B and UNC-49C, ACC-1, LGC-46 and LGC-47. Additionally, novel derivatives of the antiparasitic drug levamisole were tested on ACC receptors to determine if any modifications enhanced levamisole action. D. immitis UNC-49B assembled as a homomeric channel and exhibited an EC50 of 5mM for GABA. UNC-49B also formed a functional channel with UNC-49C, which exhibited a decrease in GABA sensitivity. Additionally, the D. immitis UNC-49 receptors were significantly more sensitive to the open channel blocker picrotoxin compared to the same receptors from the sheep parasite Haemonchus contortus. Moreover, D. immitis UNC-49C, unlike other UNC-49C subunits, did not cause a decrease in picrotoxin sensitivity when assembled with UNC-49B. D. immitis ACC-1, LGC-46, and LGC-47 were unable to form functional channels through heterologous expression in Xenopus laevis oocytes. Levamisole derivatives were therefore tested on H. contortus ACC-2, which identified at least one showing a higher sensitivity compared to levamisole. Overall, this study characterized 2 novel UNC-49 receptors, identified 3 potential members of the ACC family in D. immitis and tested 8 novel derivatives of levamisole. This study lays foundation for the identification of more ligand-gated ion channels and will serve as a starting point for future researchers looking at new drug potential targets in D. immitis.
  • Thumbnail Image
    Item
    Isolation and partial characterization of three members of the GGR-1 family in the parasitic nematode Haemonchus contortus
    (2019-08-01) Nazareth, Kristen; Forrester, Sean
    Haemonchus contortus is a blood sucking parasitic nematode that infects ruminant animals. These parasites are of significant economic importance as they are estimated to cost the agricultural industry billions of dollars each year in management. Reliance and overuse of drugs has resulted in resistance, and there is an urgent need for novel anti-parasitic drugs. Cys-loop ligand gated ion channels have been recognized as important invertebrate specific drug targets. However, specific targets must be identified and characterized prior to drug development. This thesis describes the identification and partial characterization of three ion channel subunits from the GGR-1 family – Hco-lgc-39, Hco-lgc-40, and Hco-ggr-2. The phylogenetic analysis conducted in this study revealed that these genes are widely conserved across several different parasitic nematodes. Two of these genes, lgc-39 and ggr-2, have not been previously characterized in another organism. Although lgc-40 has been previously investigated in C. elegans, the channel found in H. contortus was found to be similarly sensitive to choline and 18-fold more sensitive to acetylcholine. This increased sensitivity to acetylcholine could potentially indicate a different function in H. contortus. Pharmaceutical characterization of Hco-LGC-39, revealed that this protein subunit forms a homomeric channel that is responsive to acetylcholine, and to a higher degree, methacholine. A ligand for the Hco-GGR-2 homomeric channel has not yet been identified. Additionally, homology modelling of Hco-LGC-39 and Hco-LGC-40 provided further insight into receptor ligand interactions, and helped to identify several residues that may be important for ligand binding. This partial characterization of unique receptor subunits provides the beginning step for the production of novel anthelminthic drugs to combat the increasingly problematic parasite, H. contortus.