Covid virus increases brain cell energy expenditure to multiply

Recent memory disorders and mental confusion are among the most common neurological sequelae of Covid-19. And experiments with hamsters conducted at the University of São Paulo (USP) can help to understand how these symptoms arise and perhaps even indicate a way to fight them.

The research was conducted with live animals and also with astrocytes isolated from the central nervous system of rodents and cultivated in vitro. The results suggest that Sars-CoV-2 infection accelerates the metabolism of these nerve cells and increases the consumption of molecules used in energy generation, such as glucose and the amino acid glutamine.

The big problem is that glutamine is also important for the synthesis of glutamate –the main neurotransmitter involved in communication between neurons–, which apparently is impaired. In animals, the presence of the virus and changes in the level of proteins related to energy metabolism were observed in the hippocampus (a region of the brain essential for memory consolidation and learning) and in the cortex (also important for memory, cognition and the language).

“Apparently, Sars-CoV-2 overactivates the metabolism of astrocytes in order to obtain more energy to replicate their genetic material and produce new viral particles. So much so that when we use a drug to block glutaminolysis [a produção de energia a partir de glutamina], viral replication in cultured cells was reduced by about a third”, says Jean Pierre Peron, professor at the Institute of Biomedical Sciences (ICB-USP), researcher at the Scientific Platform Pasteur-USP (SPPU) and coordinator of the investigation.

The project had the collaboration of groups from the State University of Campinas (Unicamp) and the Research Center for Inflammatory Diseases (CRID) of the Faculty of Medicine of Ribeirão Preto (FMRP-USP). It received support from FAPESP through seven projects (20/06145-4, 20/07251-2, 17/27131-9, 15/15626-8, 20/04579-7, 20/04746-0 and 15/25364- 0). Preliminary results were published in the bioRxiv repository, in an article still without peer review.

Previous evidence

Astrocytes are the most abundant cells in the central nervous system and among their various functions is to support the functioning of neurons by providing nutrients such as glucose and glutamine. They also regulate the concentration of neurotransmitters and other substances with the potential to interfere with neuronal function, such as potassium. Furthermore, they integrate the blood-brain barrier, which protects the brain against pathogens and toxins.

Still in 2020, the group coordinated by Thiago Cunha at FMRP-USP analyzed the brain tissue of people who died of Covid-19 and confirmed the presence of Sars-CoV-2 inside astrocytes.

At Unicamp, Daniel Martins-de-Souza’s team demonstrated that the new coronavirus is capable of infecting and replicating in human astrocytes derived from induced pluripotent stem cells (IPS), a method that consists of reprogramming adult skin cells or other easily accessible tissues.

In vitro tests carried out at the time indicated that the infection induced alterations in biochemical pathways related to energy metabolism. This finding has now been reinforced with experiments done at the SPPU.

“This entire set of data suggests that the involvement of the central nervous system in patients infected with Sars-CoV-2 passes through astrocytes and that energy metabolism plays an important role in this process”, says Martins-de-Souza to the FAPESP Agency.

recent results

After infecting hamster astrocytes with Sars-CoV-2, the researchers observed that the cells started to produce inflammatory molecules (cytokines) and noticed a change in the expression of proteins related to carbon metabolism (glucose). When analyzing the metabolites present in the cell culture, they noticed that some substances were well reduced compared to the control (uninfected astrocytes).

“We saw that there was a smaller amount of glutamine and other molecules involved in energy generation and protein synthesis, such as aspartate, pyruvate and alpha-ketoglutarate. This result suggests that the cell is very metabolically activated. We believe this is because the viruses require more energy to replicate,” explains Peron.

In another experiment, astrocyte cultures were placed in a device capable of measuring glucose and oxygen consumption – a technique known as respirometry. The analysis confirmed the more accelerated metabolism of the infected cells.

“Since it is a central nervous system, it called our attention to the fact that glutamine is lower, as it is the raw material for the synthesis of glutamate and about 90% of the synapses are mediated by this neurotransmitter. Apparently, therefore, the infection causes an imbalance of energy which, in turn, leads to an imbalance in the levels of glutamate. It is possible that this alters the functioning of neurons, but it is something that still needs to be tested”, says the professor at ICB-USP.

When infected astrocytes were treated with a drug capable of blocking glutaminolysis, viral replication was reduced – there was a decrease in both the concentration of viral RNA and the amount of Sars-CoV-2 particles present in the culture medium.

In in vivo tests, hamsters were infected intranasally and the presence of the virus in the central nervous system was monitored up to 14 days later. It was possible to observe that, as occurred in vitro, the infection induced the production of inflammatory cytokines and also caused alterations in the profile of brain proteins.

“We observed the presence of viral particles in the hippocampus and cortex – two regions rich in glutamate. We also saw changes in several proteins related to carbon and glutamine metabolism. This makes us think that something similar is happening in humans and maybe this is the origin of symptoms such as memory loss, cognitive impairment, difficulty concentrating and mental confusion”, says Peron.

Martins-de-Souza comments that, in tests with human astrocytes, a reduction in glutamine had already been observed. “These new findings confirm that glutaminolysis is an important process for viral replication. We are, therefore, talking about a target in the brain that can be explored in the search for therapies”, he says.

For Peron, something more likely to be tested in the short term is the treatment of neurological sequelae of Covid-19 with drugs capable of modulating glutamate-mediated synapses. This type of medication is already used in patients with Alzheimer’s.