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Title: Mechanisms of mechanosensation in Drosophila melanogaster proprioceptors
Author: Hunter, Iain
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2020
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Proprioception is the ability to detect position in space. It is necessary for normal motor control and could share molecular mechanisms with other senses, such as hearing. These mechanisms are poorly understood and clarifying them may reveal novel targets for treatment of muscle spasticity, seizure and hardness of hearing. This research uses Drosophila models to clarify the behavioural role and molecular properties of proprioceptors; the dbd neuron and the chordotonal neurons. I hypothesise that the dbd neuron is both a pain and stretch receptor that requires DmPiezo to respond to both physiological and nociceptive stimuli. In contrast, evidence suggests that chordotonal neurons sense could sound and stretch stimuli through different mechanisms, which depend on nan/ iav/ NompC and DmPiezo respectively. We employed optogenetics, crawling, nociceptive reflex (‘pinch’ response), GCaMP imaging and whole-cell patch-clamp electrophysiology to investigate the role and mechanisms of mechanosensation in the dbd neuron. Similarly, I used crawling, hearing and GCaMP experiments to assess the role and mechanisms of mechanosensation in the chordotonal neurons. I found the dbd neuron difficult to investigate; a ‘nociceptive’ phenotype originally attributed to dbd neuron stimulation disappeared when the related driver, Bd-Gal4, was expressed in the background of a mutant (amos1) that lacks the dbd neuron. Moreover, while electrophysiology gave results like those published previously, my data were limited by issues including low seal values (~40MΩ, significantly lower than the desired 1GΩ) that were exacerbated by stretch. Chordotonal (ch) neurons were easier to study. GCaMP imaging of the larval ventral nerve cord showed that ch neurons respond to both tonal (1024Hz) and muscle contraction stimulation (mean ΔF/ F0 (%) 11.47 ± 2.93 and 7.56 ± 4.38, respectively). I imaged the ch neurons (lch1-5, vch1 and vchAB) directly, and doing so revealed some interesting spatial and temporal differences in response to sound, which implies specific tuning of neurons within the chordotonal neuron population(s)(s). GCaMP imaging also showed that CG17669, a gene with a human orthologue (DNAAF3) associated with primary ciliary dyskinesia, is necessary for ch neuron response to 1024Hz and muscle contraction. In conclusion, the behavioural role and mechanisms of the dbd neuron remain unclear and require further investigation. However, it appears that while the ch neurons can detect stretch (and so act as proprioceptors), this function is not required for normal movement in larvae. The ch neurons appear to be a sense organ with a single mechanism of mechanosensation, that is optimised for detection of tonal stimuli in the hearing range. Finally, this research is the first to: (1) image the response of vch1 and vchAB ch neurons response to sound; (2) provide evidence that subsets of Drosophila ch neurons may be tuned to respond to specific amplitudes and/ or frequencies; (3) use real-time calcium imaging to demonstrate the effect of CG17669 mutation on the function of ch neurons.
Supervisor: Jarman, Andrew ; Bewick, Guy Sponsor: Biotechnology and Biological Sciences Research Council (BBSRC)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: proprioception ; muscle spindle ; mechanoelectrical transducer proteins ; dyneins ; primary ciliary dyskinesia ; Drosophila