I decided to research toxoplasmosis, because I was curious about its effects on behavior, especially since it is a very common disease (approximately one-third of humans worldwide are infected) and its preferred host is the housecat. I found a research study (David et al, 2016) investigating the effects of Toxoplasma gondii infection on the brain. They did several different experiments using mice during the course of the study. They looked at the morphological and molecular effects of infection by examining slices of the brain at different stages of infection, tested for levels of glutamate and amino acids in the brain, observed the effects of the drug ceftriaxone, and did a behavioral test.

First, they wanted to see if the morphological effects of infection remain once the disease enters its chronic phase. Toxoplasmosis is generally regarded as benign once the initial infection has passed, although it remains latent for the lifetime of the host. The research team began by examining scanning serial electron microscopy images of astrocytes (a type of brain cell that swells as part of the immune response) at three, six, and twelve weeks. The disease is considered chronic after the third week. The images shown in the paper depict a slight swelling after three weeks and significant swelling at six weeks (with the cell endfeet almost six times the width of those of the original cell). At twelve weeks, the swelling is somewhat reduced, although still much greater than in an uninfected brain. This shows that the immune response remains long after the initial infection.

Next the research team wanted to see if the change in astrocyte morphology also led to a change in function. One of the normal functions of astrocytes is to remove extracellular glutamate using two transporters, GLT-1 and GLAST. Then enzyme glutamine synthetase (GS) converts the glutamate to glutamine inside the cell. Therefore, to see if astrocyte function was impacted by T. gondii infection, researchers compared the levels of GLT-1, GLAST, and GS in infected and uninfected brains. The figures presented in the paper show that GLT-1 levels dropped after one week and continued to decrease throughout the course of infection. GS levels initially showed no change, but then dropped to about half normal levels after two weeks and stayed at about that level. GLAST levels showed no change to a slight increase over normal levels. This shows that the astrocytes were unable to perform their normal task of regulating extracellular glutamate due to infection.

Next, the researchers wanted to investigate the sources of the behavioral changes seen in other studies of toxoplasma infection. They decided to examine the prefrontal cortex, an area of the brain related, to fear, anxiety, and decision-making. First they stained infected brain samples for B-III-tubulin, a component of neuron cytoskeletons. Images in the paper clearly show reduced B-III-tubulin density in the infected brain. Next, they stained for neurons themselves, to see if the reduction in B-III-tubulin was a result of neuron death or reduced function. The next images show no change between infected and uninfected brains, indicating no loss of neurons. Next they looked for dendritic spines, loss of which occurs in disorders such as autism and Fragile X syndrome. The images show fewer spines after six weeks than in the uninfected brain. Loss of dendritic spines is associated with loss of synapses, so they research team next stained for a presynaptic marker called VGlut-1. Images of infected brains show less VGlut-1 activity than uninfected brains.

The researchers hypothesized that the changes observed in the prefrontal cortex could be a result of glutamate excitotoxicity, meaning that glutamate levels had reached a pathological level, something that occurs in neurodegenerative diseases such as multiple sclerosis. To measure glutamate levels over the course of infection, researchers inserted probes into the frontal cortex of infected and uninfected mice and made measurements of glutamate and twenty amino acids. The figures show low glutamate levels for the first three weeks of infection, and then a rapid, continuous increase thereafter.

Because their experiments showed that glutamate levels increased in the brain at the same time that GLT-1 (responsible for transporting glutamate into the cell) levels dropped, researchers wanted to see if a drug designed to increase GLT-1, ceftriaxone, would have an effect. First they wanted to ensure that the drug did not interfere with the immune response, and in the figures shown, the brains of those infected look similar whether or not they received treatment. Next, they compared GLT-1 and glutamate levels of treated and untreated individuals using the same tests done previously. The results for treated individuals appear close to uninfected than infected, untreated individuals, showing that ceftriaxone is effective at restoring GLT-1 funtion. Those treated with ceftriaxone also showed restoration of neuronal function in the prefrontal cortex.

The last test done was a behavioral test, using uninfected, untreated infected, and infected treated with ceftriaxone. The images shown show that infected mice, regardless of treatment, spent more time in the open areas of a maze than uninfected mice, indicating decreased anxiety. Figures show no change in mouse velocity or distance traveled regardless of infection/treatment status. EEG readings show results for the treated mice that fall between the uninfected and untreated infected groups. These results indicate that more research needs to be done into the behavioral effects of toxoplasmosis, as restoring glutamate levels does not fully restore neuronal activity.

This research is related to cells because it looks at infection effects on two types of cells, astrocytes and neurons. It also examined enzyme functionality, cell communication, transporters, and structural components such as the cytoskeleton. The images in the paper were of cells or cell components. Ultimately, any disease must have an effect at the cellular level and toxoplasmosis is no exception.

 

David, C. N., Frias, E. S., Szu, J. I., Vieira, P. A., Hubbard, J. A., Lovelace, J., . . . Wilson, E. H. (2016). GLT-1-dependent disruption of CNS glutamate homeostasis and neuronal function by the protozoan parasite toxoplasma gondii. PLoS Pathogens,12(6) doi:http://dx.doi.org.proxy.lib.odu.edu/10.1371/journal.ppat.1005643