These tiny graphene oxide flakes fight PTSD in rats

April 3, 2021
A method to prevent PTSD in rats might have greater implications. (Unsplash/Alexandr Gusev)

A method to prevent PTSD in rats might have greater implications. (Unsplash/Alexandr Gusev)

Researchers have used minuscule flakes of graphene oxide to disrupt a harmful form of plasticity in the amygdala region of the brain, overcoming post-traumatic stress disorder in an animal model of the mental health condition.

"They are able to affect locally synapses in a very precise manner," Laura Ballerini, a physiologist at the International School for Advanced Studies (SISSA) and an author of the study, published March 4 in Biomaterials, told The Academic Times. "This blockage will show by electrophysiology — but is it good enough to block a behavior?"

Ballerini and her colleagues previously found that graphene oxide can interfere with the excitatory neurotransmitter glutamate. Unlike inhibitory neurotransmitters, which suppress neuronal firing, excitatory neurotransmitters such as glutamate make neurons likelier to fire — and glutamate-related overexcitement in the brain has been linked to mental illness. The researchers hypothesized that graphene oxide would interfere with unwanted hyperactivation of glutamate in the amygdala, a region of the brain that is believed to store the aversive memories that characterize post-traumatic stress disorder. To test this, they used a rat model of PTSD.

The researchers familiarized a group of rats with what they called an "odor avoidance box," made up of an open area and a small hidey-hole. Then, they exposed some rats to a collar that had been worn by a cat and exposed the others to an unworn collar. The scientists reexposed the rats to the collars two days and then six days later. At both of those times, they assessed the rats' anxiety-related behavior by tracking how often they spent in the open area versus the hidey-hole. Rats that had been exposed to the worn collar spent a lot more time defensively scanning the environment from inside that hole.

"It's a very well-established model," Ballerini explained. "I find it very elegant."

Next, the scientists injected graphene oxide flakes into the amygdalas of some of the rats. To assess whether the amygdala really mattered, they delivered the graphene oxide to a different brain region, the perirhinal cortex, in some rats. They also included control rats that had been injected with a saline solution.

Two days after surgery, they reexposed the rats to the collars. Strikingly, the rats whose amygdalas had been targeted with graphene oxide showed considerably less anxiety than the rats injected with saline or injected in a different brain region.

"They behaved like control animals," Ballerini said. "So we stepped back to in vitro again, because the hypothesis there was that we blocked the long-term potentiation — the plasticity of the synapse — which, in this case, is assumed to be responsible for the pathological plasticity of the behavior. We used acute slices of the amygdala to demonstrate that there is specificity on the glutamatergic synapses of the [graphene oxide] activity." These in vitro experiments confirmed their behavioral study.

The graphene flakes in those synapses could be loosely compared to an insulator in an electrical circuit. "It is isolating certain synapses from working in a very specific manner for a transient time, which is long enough to affect long-term consequences," Ballerini said. "To me, it's interesting that the interaction of the graphene flakes has to be a physical one."

Scientists do not yet know why the graphene flakes affect synapses in the rat brain in this way, according to Ballerini. "We have some ideas, of course," she said. "Why is it affecting the glutamatergic [synapse] and not the inhibitory one? Maybe it's due to the hyperactivity that's present at the glutamatergic one, or maybe it's the size and the profile of the synapse per se."

Ballerini noted that the graphene she used is effectively nontoxic, at least when delivered in the short-term, controlled fashion used in biomedical testing. "The material is dangerous and might be noxious and toxic, but in the medical applications, it's a completely different setting," she said.

Ballerini also distinguished the pathological plasticity she targeted with graphene oxide from neuroplasticity more generally, which is generally seen as positive. This "bad plasticity" could be compared to an autoimmune response, in which the body's indispensable protective force, the immune system, goes awry and starts attacking the body itself.

"Plasticity is a good thing," she explained. "We think this is the molecular mechanism for memory storage, for learning and for development. But then there is more recent evidence that there might be pathological behavior or brain pathologies related to bad plasticity, which is exerted by exactly the same molecular mechanisms. You might look at any process as a good physiological one, but when it gets into a pathology, it needs to be interrupted if you want to interrupt the pathology."

Ballerini sees her work in the wider context of precision medicine, which aims to target drugs and other therapies to specific kinds of patients. "Beyond the fact that you might use the flakes as a drug delivery vector, or a vector which goes directly on a certain location, the concept per se is the one which I'm interested in," she said.

Yet Ballerini is cautious when commenting on her work's value to PTSD in people. "In general, the strategy of selectively affecting one synapse and obtaining the reverse of a disease has an attraction in translational applications," she said. "I'm looking for the mechanisms. Once the mechanisms are elucidated, this might be taken over by more clinical groups, to test whether this can be translated to applications."

The article, "Graphene oxide prevents lateral amygdala dysfunctional synaptic plasticity and reverts long lasting anxiety behavior in rats," published March 4 in Biomaterials, was authored by Audrey Franceschi Biagioni, Giada Cellot, Elisa Pati, Raffaele Casani, and Laura Ballerini, International School for Advanced Studies (SISSA); Neus Lozano and Belén Ballesteros, Catalan Institute of Nanoscience and Nanotechnology; Norberto Cysne Coimbra,  Ribeirão Preto Medical School of the University of São Paulo (FMRP-USP); and Kostas Kostarelos, Catalan Institute of Nanoscience and Nanotechnology and the University of Manchester.

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