A recent study published in the journal Neuron provides evidence that a specific group of adrenaline-producing brain cells acts as a major driver of fear and anxiety behaviors. Activation of these cells produces immediate and long-lasting anxiety, and dampening their activity reduces stress responses. These findings suggest that targeting these specific neurons could offer a more precise way to treat anxiety disorders in the future.
Anxiety disorders affect over 300 million people worldwide and significantly impact daily life. Current treatments, including various medications and therapies, can cause adverse side effects like insomnia, jitteriness, or cognitive impairment. Relapse is also common when patients stop taking their prescribed drugs. This ongoing challenge highlights a need for more specific and localized therapeutic targets within the brain.
The physical symptoms of stress, such as elevated heart rate and rapid breathing, are coordinated by a region in the lower brainstem. Within this region, two populations of neurons sit very close together and share similar genetic profiles. A1 cells produce norepinephrine, and C1 cells produce epinephrine, a chemical messenger also known as adrenaline.
Because these two cell types are heavily intermingled, previous scientific tools lacked the specificity to study them independently. Prior studies generally grouped them together, operating under the assumption that they primarily managed physical functions like blood pressure rather than complex emotions. A research team based at St. Jude Children’s Research Hospital, led by Lindsay A. Schwarz alongside Carlos Fernández-Peña, sought to isolate the exact behavioral functions of these individual cell groups. They wanted to know if C1 neurons directly modulate psychological anxiety rather than just physical stress responses.
To separate C1 cells from A1 cells, the scientists developed a specialized line of genetically modified mice. They combined these mice with engineered viral vectors to create biological logic gates, allowing them to selectively target only the C1 neurons. The researchers used adult male and female mice ranging from 6 to 20 weeks old for their behavioral experiments.
In their first test, they exposed the animals to a 30-minute physical restraint stressor. By examining RNA markers in the brain tissue afterward, they found that this stressful event preferentially activated the C1 neurons over the A1 neurons. RNA is a molecule that helps carry out genetic instructions, and its presence indicates that a cell has recently been active.
Next, the authors used optogenetics, a technique that uses focused light to control genetically modified cells. They implanted tiny optic fibers into the brains of the mice to shine light directly onto the C1 neurons. During a five-minute test in an open field arena measuring 40 by 40 by 20 centimeters, the light stimulation caused the mice to spend significantly more time hiding in the corners.
The scientists also tested the mice on an elevated zero maze, which is a circular platform 100 centimeters in diameter and elevated 61 centimeters off the ground. The maze features alternating open and enclosed sections. Mice naturally fear the exposed areas and prefer the walled sections.
When the researchers activated the C1 neurons with light, the mice spent much less time in the open areas, which indicates a heightened state of anxiety. Interestingly, when the researchers stimulated both A1 and C1 neurons simultaneously in a separate test, the mice simply stopped moving entirely. This temporary behavioral arrest was never observed when targeting C1 cells alone.
This distinction provides evidence that the two neighboring cell types serve very different functions. The scientists then traced the physical connections of the C1 neurons to see where they sent their signals. They discovered dense connections traveling to the ventrolateral periaqueductal gray. This midbrain region is well known for processing threats and triggering defensive behaviors like freezing in place.
To test this specific pathway, the researchers moved their optic fibers to shine light solely on the C1 nerve endings located inside the midbrain. Activating this isolated connection produced a severe anxiety response on the elevated zero maze. The authors also found that a brief activation of this pathway led to persistent anxiety.
They stimulated the C1 connections while the mice were in their home cages, then left them completely undisturbed for seven days. When tested on the elevated zero maze a full week later, these mice still displayed elevated anxiety behaviors compared to control groups. The researchers also tested a real-time place preference model, where mice avoided a specific side of a chamber if it triggered the light stimulation, showing that the animals found the C1 activation highly unpleasant.
To observe natural brain activity, the researchers utilized fiber photometry, a method that measures calcium fluctuations to track real-time cell firing. As the mice explored the elevated zero maze, the C1 neurons rapidly spiked in activity exactly when the animals stepped into the scary, open areas. This natural brainstem activity directly excited the downstream neurons in the midbrain.
The midbrain neurons stayed active for several seconds after the mice entered the open zones. This sustained firing suggests that the circuit constantly calculates environmental threats. To see if blocking this circuit could prevent anxiety, the scientists used designer drugs to temporarily turn off the C1 neurons.
They administered a specialized chemical compound called DCZ at a dose of 500 micrograms per kilogram of body weight 15 minutes before testing. In one experiment, the scientists exposed the mice to a looming fear test, where a dark, expanding circle is projected from above to mimic a flying predator. Mice normally freeze in terror at this sight. Inhibiting the C1 neurons significantly reduced both the number of freezing events and their total duration.
The researchers also tested a fear conditioning model, where a specific tone was paired with a mild, one-second electric foot shock. Days later, playing the tone alone caused the mice to freeze. Mice with suppressed C1 activity showed a much faster reduction in this learned freezing behavior over time.
In a final behavioral test, the scientists combined a 30-minute physical restraint period with the elevated zero maze. Normally, prior restraint causes mice to hide almost entirely in the closed sections of the maze. Turning off the C1 neurons right before the restraint period completely eliminated this stress-induced anxiety on the maze.
Using a modified rabies virus as a biological tracker, the team mapped all the upstream brain areas that send instructions to the C1 cells. They found that C1 neurons receive input from regions that regulate pain, breathing, sleep, and basic internal body states. This network arrangement tends to position the C1 cells as a central hub, gathering physiological data to inform emotional responses.
A potential limitation of the study is the exclusive use of animal models. Human brains are far more complex, and human anxiety is heavily influenced by learned psychological factors rather than just innate predator avoidance. Functional connections between the brainstem and the midbrain do exist in humans, meaning this circuit likely plays a similar role in people, but human studies are needed to confirm this.
Another limitation is that the researchers pooled male and female mice together for their analysis. They did not independently analyze the data to look for sex-based differences. Future studies will need to investigate whether this anxiety circuit operates differently depending on biological sex.
The short lifespan of the designer drug used to inhibit the cells also restricted the testing windows. When the researchers tried to add a delay period between the stressful event and the behavioral test, the drug wore off, and the anxiety returned. Longer-lasting inhibition methods will be necessary to fully understand the long-term benefits of turning off these neurons.
Future research should explore how C1 neurons behave under different internal states. During one test involving fasted mice in a novel environment, stimulating the C1 cells actually encouraged the animals to eat faster. This unexpected result suggests the circuit might adapt its function based on extreme hunger or other pressing biological needs. The team hopes to map exactly how interoceptive signals, such as changes in breathing or heart rate, feed into the C1 neurons to trigger panic.
The study, “Autonomic C1 neurons promote anxiety via activation of vlPAG,” was authored by Carlos Fernández-Peña, Rachel L. Pace, Lourds M. Fernando, Heather Sheppard, Brittany G. Pittman, and Lindsay A. Schwarz.