a rotation project in John Tuthill’s Lab
project overview
brains evolved to produce movements. the only way that we can interact with the external environment is through movements. this highlights the importance of studying motor control.
brains produce movements, on a broad scale, through motor neuron activity controlling which muscles are on or off. for different movements, we need different combinations of muscles and different combinations of motor neurons activated. and for all of these behaviors, we need to rapidly and continuously adjust our body movements to produce smooth and stabile posture. proprioception is the sense of self movement and body position (tuthill, azim 2018). there are specific proprioceptors that are activated when a limb moves in a specific direction, activating motor neurons for the respective muscle, but also send indirect inhibitory projects to antagonistic muscles that tend to work against that movement. This type of connection is a simplified description of a phenomenon like how reflexes work. for any movement at any given time, there are thousands of proprioceptors used for thousands of neurons.
flies are great models to study proprioception and motor control due to the incredible genetic access we have to neurons in the brain and ventral nerve cord, the connectomic access to the entire nervous system, and simple yet dynamic behaviors. One proprioceptor involved in a phenomenon like reflexes are the hair plates. hair plates are small bundles of 6-10 hairs. each hair innervates a single mechanosensory neuron when deflected during leg bending. the hair plates help convey joint limits and drive reflexive movements.
connectomic analysis of hair plate neurons reveals at a synapse resultion the circuits used to coordinate and accomplish diverse motor outputs. one of the hairplates, coxa hairplate 8, connects directly to coxa posterior movement motor neurons. However, it also has a ton of indirect inhibitory connections to other motor pools (pratt et al. 2024). One of these groups of inhibitory neurons are a group of cells that come from the 13a hemilineage. these neurons receive a ton of sensory input and input from other motor neurons. 13a neurons were shown to help coordinate leg movements (Syed et al 2024).
for my rotation project, i wanted to patch the specific set of 13a neurons downstream of coxa hairplate 8 neurons. To do this, I expressed GFP in those neurons, mounted a living fly, removed the cuticle to expose the T1 segment of the ventral nerve cord, then guided an electrode to those cells to hopefully seal and then break into the cell to record its electrophysiological properties. In some of my preliminary recordings I found that one of the 13a neurons was a non spiking neuron, but could not tell whether leg movements changed with current injection or produce spiking if moved.
Understanding the electrophysiological properties of these neurons gets us closer to understanding how hairplates and more broadly how premotor circuits help coordinate activation of different motor neurons for reflexive behaviors. using ephys is important because the timescale for these fast behaviors, like the time between each step, is less than 30 ms which is much faster than what we can accurately capture with other techniques like calcium imaging.
Now that I have the technique down, I will continue this work to produce a full dataset of the 13a neurons.
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timeline
an example timeline of how a lab notebook may look:
2024-05-30 - patched from spiking cell
2024-05-29 - gave practice rotation talk
… lots of work with some successful patching but incomplete note taking :(
2024-05-03 - tried new mounting technique, unsuccessful patching
2024-04-30 - patched a cell and tried to make a recording
2024-04-26 - first successful patch!
2024-04-25 - patching practice
2024-04-24 - patching practice
2024-04-23 - patching practice
2024-04-19 - continued practicing patching
2024-04-18 - almost patched a cell
2024-04-16 - changed objectives, tried patching with John
2024-04-15 - took pictures of both GFP drivers lines
2024-04-12 - tried to fix contrast on scope, cleaned cells
2024-04-10 - found 13>GFP under scope!
2024-04-09 - background reading of lab papers, learned fix electrode programs, met with brandon about connectomic analysis
2024-04-05 - first 13A>GFP neurons emerged, could see them under the scope
2024-04-04 - reading, journal club, more practice
2024-04-03 - practiced mounting dissecting
2024-04-02 - practiced cleaning with ellen
2024-03-29 - worked on mounting / dissecting
2024-03-28 - shadowed john almost patched a cell
2024-03-27 - background reading
2024-03-26 - axon guide reading
2024-03-25 - background reading