Study Explores How Modulating V2a Neurons Could Activate Muscles to Improve Breathing

Patients with neuromuscular disease or spinal cord injury face a myriad of medical challenges. One of the most distressing is difficulty breathing, as certain neuronal circuits deteriorate. Steven Crone, PhD, a neurodevelopmental biologist at Cincinnati Children’s, has devoted much of his research to understanding those circuits and exploring ways to modulate them. Recently that has included exploring the relationship between V2a neurons and accessory respiratory muscles (ARMs). These muscles play a role in increasing ventilation during exercise and when neuromuscular disease and spinal cord injury are present.

Crone, who works in the Division of Pediatric Neurosurgery, discusses his lab’s recent discoveries about this circuitry and how these findings could help scientists develop therapies that target neurons to enhance ARMs. “V2a Neurons Constrain Extradiaphragmatic Respiratory Muscle Activity at Rest,” was published in eNeuro (July/August 2019).

Tell us about your recent study.

Previously, my lab had shown that when we increased the excitability of V2a neurons in adult mice, ARMs activated. We decided to explore that further with a control study and discovered that inhibiting the excitability of V2a neurons in adult mice also led to ARM activation. This was an unexpected and novel finding, so we changed course to investigate further.

Our studies used different types of “designer receptors exclusively activated by designer drugs” (DREADD) to manipulate the firing activity of V2a neurons in adult and neonatal transgenic mice. In the more recent study, we injected a chemical into two lines of mice. In one mouse line, the chemical increased the excitability of V2a neurons, and in the other, the chemical inhibited V2a neurons. Then, we measured the activity of ARMs and the diaphragm using electromyography (EMG) recordings.

Did you face technical challenges?

Yes. One was that we wanted to measure ARM and diaphragm activity in mice that were awake and active because anesthesia could potentially interfere with ARM activation. One of our researchers devised a method to safely implant radio transmitter electrodes into mice to perform EMG recordings of the diaphragm. Once in place, the transmitter delivered data to a receiver, and we used a plethysmography chamber to measure pressure changes while also measuring muscle activity.

What were the key findings of this study?

When we silenced V2a neurons in adult mice, ARMs activated and breathing patterns increased slightly. But the mice did not experience apnea or irregular breathing. That is an encouraging finding because it supports the feasibility of targeting V2a-containing circuits to control ARMs. This could improve breathing in individuals with neuromuscular disease or spinal cord injury, without risk of impairing respiratory rhythm generation.

When we silenced V2a neurons in neonatal mice, the effect was different. Breathing slowed and became irregular. This distinction provides more evidence that neuronal function can change over time, and it demonstrates the value of doing experiments on subjects at different stages of development.

What’s next for your lab?

We recently began a new R01 study to identify the different types of V2a neurons and test which ones are activating and which ones are silencing the pathway. Longer-term, we hope to identify molecules in V2a neurons to use as drug targets to improve breathing.

(Published August 2023)

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