Date of Award
1-1-2024
Document Type
Dissertation
Degree Name
Ph.D. in Biological Science
First Advisor
Gregg W. Roman
Second Advisor
Alberto G. Del Arco
Third Advisor
Joshua Bloomekatz
School
University of Mississippi
Relational Format
dissertation/thesis
Abstract
Dunc13 is a Drosophila active zone protein homologous to the mammalian Munc13that interacts with RIM and Rab3 as a heterotrimer. Alcohol binds to the C1 domain of Dunc13/Munc13 and inhibits its ability to bind to Diacylglycerol, inhibiting neuronal signaling. The genetic reduction of Dunc13 activity in the Dunc13P84200/+ heterozygotes induces a state in flies that mimics alcohol tolerance, evidenced by increased self-administration, and behavioral, physiological, and molecular resistance. This includes changes in the expression of key synaptic proteins, such as decreased expression of Dunc13 and Syb mRNA, and increased expression of Rim and Rab3 4hrs after ethanol exposure. Dunc13 has 2 major isoforms, Dunc13A, and Dunc13B that are localized differently with voltage-gated calcium channels (VGCCs)during vesicle priming. In Drosophila, Dunc13A is closely coupled with VGCCs recruited by Brp, interacts with RIM, Rab3, and Rbp, and promotes phasic signaling. Dunc13B is loosely coupled with VGCCs recruited by Syd-1, interacts with Liprin-α, and promotes tonic signaling. Genetic reduction of Dunc13A and Brp increased ethanol sedation sensitivity whereas Dunc13Band Syd-1reduction increased resistance to ethanol sedation. These results suggest that Dunc13 isoforms differentially regulate ethanol sedation sensitivity. Furthermore, Rim plays a crucial role in recruiting synaptic vesicles (SVs) closer to the proximity of the active zone while tethering VGCCs with Rbp interaction. Both Rim andRab3 are important in the docking and priming of synaptic vesicles. Genetically reducing Rim, Rab3, and Rbp increased ethanol sedation sensitivity whereas, overexpressing Rim and Rab3increased resistance to ethanol mimicking tolerance. These results suggest a bi-directional modulation of ethanol sedation behavior through presynaptic vesicle fusion proteins. Interestingly, Liprin-α interacts with Dunc13B, and Liprin-α mutants showed increased sensitivity to ethanol sedation contrary to Dunc13 mutants, suggesting a broader role of Liprin-α at the presynaptic terminal than currently known. Finally, our genetic interaction studies using Dunc13P84200, Dunc13A-Null (EMS7.5), RIMEX73, and LiprinEX15mutants suggest at least two possible concurrent models that describe presynaptic vesicle fusion protein-mediated modulation of ethanol sedation sensitivity in flies. The first model involves behavior modulation via different signaling of Dunc13 isoforms. The second model is the regulation of the presynaptic vesicle pool size.
Recommended Citation
Shrestha, Gaurav, "Dunc13 isoforms, RIM, and Rab3 Differentially Modulate Ethanol Sedation Sensitivity in Drosophila melanogaster." (2024). Electronic Theses and Dissertations. 2978.
https://egrove.olemiss.edu/etd/2978