Think about traffic flow in a city – there are stop signs, one-way streets, and traffic lights to organize movement across a widespread network.
Now, imagine what would happen if you removed some of the traffic signals.
Among your brain’s 86 billion neurons are the brain’s own version of stop signals: inhibitory neurons that emit chemicals to help regulate the flow of ions traveling down one cell’s axon to the next neuron.
Just as a city without traffic signals would experience a spike in vehicle accidents, when the brain’s inhibitory signals are weakened, activity can become unchecked, leading to a variety of disorders.
In a new study published in GLIA on March 11, Virginia Tech neuroscientists at the Fralin Biomedical Research Institute at VTC describe how the common Toxoplasma gondii parasite prompts the loss of inhibitory signaling in the brain by altering the behavior of nearby cells called microglia.
The Centers for Disease Control and Prevention estimates that 40 million Americans have varying levels of Toxoplasma infection, although most cases are asymptomatic.
Commonly passed to humans via exposure to farm animals, infected cat litter, or undercooked meat, the parasitic infection causes unnoticeable or mild, to flu-like symptoms in most healthy people.
But for a small number of patients, these microscopic parasites hunker down inside of neurons, causing signaling errors that can result in seizures, personality and mood disorders, vision changes, and even schizophrenia.
“After the initial infection, humans will enter a phase of chronic infection. We wanted to examine how the brain circuitry changes in these later stages of parasitic cyst infection,” said Michael Fox, a professor at the Fralin Biomedical Research Institute and the study’s lead author.
The parasite forms microscopic cysts tucked inside of individual neurons.
“The theory is that neurons are a great place to hide because they fail to produce some molecules that could attract cells of the immune system,” said Fox, who is also director of the research institute’s Center for Neurobiology Research.
Fox and his collaborator, Ira Blader, recently reported that long-term Toxoplasma infections redistribute levels of a key enzyme needed in inhibitory neurons to generate GABA, a neurotransmitter released at the specialized connection between two neurons, called a synapse.
Building on that discovery, the scientists revealed that persistent parasitic infection causes a loss of inhibitory synapses, and they also observed that cell bodies of neurons became ensheathed by other brain cells, microglia.
These microglia appear to prevent inhibitory interneurons from signaling to the ensheathed neurons.
“In neuropsychiatric disorders, similar patterns of inhibitory synapse loss have been reported, therefore these results could explain why some people develop these disorders post-infection,” Fox said.
Fox said the inspiration for this study started years ago when he met Blader, a collaborating author and professor of microbiology and immunology at the University at Buffalo Jacobs School of Medicine and Biomedical Sciences, after he delivered a seminar at Virginia Tech.
Blader studied Toxoplasma gondii and wanted to understand how specific strands of the parasite impacted the retina in mouse models.
Working together, the two labs found that while the retina showed no remarkable changes, inhibitory interneurons in the brain were clearly impacted by the infection. Mice – similar to humans – exhibit unusual behavioral changes after Toxoplasma infection.
One hallmark symptom in infected mice is their tendency to approach known predators, such as cats, displaying a lack of fear, survival instincts, or situational processing.
“Even though a lot of neuroscientists study Toxoplasma infection as a model for immune response in the brain, we want to understand what this parasite does to rewire the brain, leading to these dramatic shifts in behavior,” Fox said.
The parasite forms microscopic cysts tucked inside of individual neurons.
Future studies will focus on further describing how microglia are involved in the brain’s response to the parasite.
Among the research collaborators is Gabriela Carrillo, the study’s first author and a graduate student in the Translational Biology, Medicine, and Health Program. Previously trained as an architect before pursuing a career in science, Carrillo chose this topic for her doctorate dissertation because it involves an interdisciplinary approach.
“By combining multiple tools to study infectious disease and neuroscience, we’re able to approach this complex mechanistic response from multiple perspectives to ask entirely new questions,” Carrillo said.
“This research is fascinating to me because we are exposing activated microglial response and fundamental aspects of brain biology through a microbiological lens.”
The study’s other contributing authors include Valerie Ballard, a Roanoke Valley Governor’s School high school student; Taylor Glausen, a graduate student working in Blader’s laboratory at the University at Buffalo; Zack Boone, a Virginia Tech undergraduate student; Cyrus Hinkson, a fourth-year Virginia Tech Carilion School of Medicine student; and Elizabeth Wohlfert, an assistant professor of microbiology and immunology at the University at Buffalo.
Toxoplasma gondii (T. gondii) is an obligate neurotropic protozoan parasite that forms cysts in some tissues, including the brain of warm-blooded mammals like humans . Cats are the final hosts of this parasite, and human infection occurs often via the ingestion of oocyst in water or tissue cyst in undercooked or raw meat .
Toxoplasma gondii infects about 25–30% of the people in developed and developing countries . Most of T. gondii infections in immunocompetent individuals are asymptomatic.
Nevertheless, in congenital disorders and immunocompromised patients, the infection may lead to the eye, lymph node, and central nervous system diseases [1, 3, 4].
The neurotropic nature and other specifications of T. gondii have made it a potential causative agent for psychiatric and behavioral disorders. T. gondii uses a complicated mechanism to gain access to the brain. When T. gondii reaches the brain, it invades different brain cells, including astrocytes and neurons, where it forms cysts .
According to the evidence, latent toxoplasmosis causes behavioral disorders not only in mice but also in humans [6, 7].
Recently published systematic review articles have proven the relationship between T. gondii infection and some psychiatric disorders such as bipolar disorder [8, 9], schizophrenia [9, 10], and epilepsy .
Among behavioral disorders, depression as the most common mental disorder, is coupled with remarkable morbidity and mortality. According to the Diagnostic and Statistical Manual of Mental Disorder (DSM-V) criteria, major depressive disorder (MDD) is a mental disorder characterized by at least two weeks of feeling low mood and disappointment. It is often accompanied by decreased self-esteem, loss of interest, cognitive performance, delight, dream, appetite, energy level, feelings of worthlessness, changes in weight, and having suicidal ideation and attempt for suicide [12–14].
So far, two systematic reviews have evaluated the relationship between toxoplasmosis and depression. One study has indicated that depression might be associated with microbial infections, such as those caused by T. gondii, human herpesvirus, hepatitis B virus, Chlamydiaceae, and Borna disease virus .
In addition, in a recent systematic review, several psychiatric disorders, namely schizophrenia, bipolar disorder, obsessive-compulsive disorder, and addiction, have been reported to be in association with toxoplasmosis. However, in the mentioned study, no significant relationship was found between depression and toxoplasmosis .
Since the results of the available original articles are contradictory, the aim of this new systematic review was to comprehensively assess the association between MDD and toxoplasmosis adding the latest studies on this matter. This study was also targeted toward the investigation of the susceptibility of toxoplasmosis patients to depression.
This systematic review and meta-analysis aimed to quantify the pooled ORs of anti-T. gondii IgG and IgM antibodies in depressed patients and compare them with those of control groups.
The results obtained from the current study showed that the overall ORs of anti-T. gondii IgG and IgM antibodies in patients with MDD were 1.15 (95% CI: 0.95–1.39) and 1.69 (95% CI: 0.72–3.96), respectively. Based on these results, toxoplasmosis is not considered a risk factor for depressed patients.
There are several reasons accounting for these negative findings:
1) infectious agent does not play an etiologic role in major depression,
2) the infectious agent has attended earlier in individuals’ life but are no longer detectable .
Based on the results of the meta-analysis, the depressed patients had a lower seroprevalence of T. gondii as compared to the controls. Major depression showed no association with anti-Toxoplasma IgG and anti-Toxoplasma IgM.
Our results are inconsistent with those of the systematic reviews evaluating the association between toxoplasmosis and psychiatric disorders, including epilepsy (OR = 2.25) , bipolar disorder (OR = 1.26) , obsessive-compulsive disorder (OR = 1.96) , schizophrenia (OR = 1.81) , and addiction (OR = 1.91) .
On the other hand, our results are in line with those of a couple of previous meta-analyses that showed no association between T. gondii infection and depression [9, 15]. Wang et al. in 2014  evaluated the relationship between different infectious agents and depression. In this meta-analysis, only three studies had addressed the relationship between T. gondii infection and depression. After analyzing various studies, no significant association was found, and the OR was calculated as 1.36. All three studies were included in the current meta-analysis.
In a unique meta-analysis published in 2015 , Sutterland et al. assessed the prevalence of T. gondii infection in several psychiatric disorders using both published and unpublished studies.
They obtained a non-significant OR of 1.2 indicating no general difference between healthy subjects and depressed patients in terms of T. gondii infection prevalence. Since the present study only included the published studies, 7 out of the 9 studies reviewed in the mentioned study were investigated in the current meta-analysis. One of the two non-investigated studies was unpublished, and the other study was a summary of the paper presented at a congress .
In addition, 22 studies were not investigated in the mentioned meta-analysis; however, they were included in the current study due to meeting our inclusion and exclusion criteria.
The advantages of this meta-analysis are as follows:
1) the investigation of a larger number of cases and controls, compared to those of the previous studies that improves the statistical power to assess the association between toxoplasmosis and depression,
2) inclusion of studies from eight other countries in the meta-analysis that intensifies the consistency of the association,
3) analysis of case-control and cross-sectional studies based on OR, and
4) exclusive attention to MDD.
According to Fig 2, the lowest and highest ORs are related to the studies performed by Massa et al.  and Conejero-Goldberg et al. , respectively. This difference could be due to the sole use of postmortem brain samples by Conejero-Goldberg et al. .
In addition, the quality of the mentioned study was low, and the number of the subjects was small. However, Massa et al. used a larger sample size with a different set of psychiatric diagnoses. Moreover, race, educational level, and economic status can result in different ORs .
Difference in the sensitivity and specificity of detection methods, different geographic regions, age, gender, and ethnic groups are the main reasons for the difference in the prevalence of T. gondii [6, 9, 53].
Given the incomplete data of the studies assessing the relationship between different variables (e.g., age, gender, ethnic, diet, family history, parasite strain, socioeconomic level) and the prevalence of toxoplasmosis, meta-analysis was not applicable for these variables. Furthermore, due to the lack of the evaluation of these risk factors in some studies, these variables cannot be analyzed; accordingly, this is considered as a basic gap.
For example, the gender agent has an apparent impact on the psychomotor performance of humans . Female sex hormones are known to manipulate dopaminergic actions in some parts of the brain.
The occurrence of degenerative disorders in women, such as Parkinsonism, decreases the protective effect of dopamine . Conversely, higher levels of testosterone in men can cause toxoplasmosis associated with changes in human and animal behaviors .
Therefore, this variable is important in evaluating the relationship between depression and T. gondii and needs to be addressed.
The possible mechanism underlying the behavioral changes correlated with latent toxoplasmosis is the presence and spread of T. gondii cysts in the central nervous system. Berenreiterová et al.  reported that Toxoplasma latent cysts in mice were distributed all over the brain; it would seem possible that parasitic cysts in these areas in humans could alter both frontal and limbic regions which could then result in behavioral and emotional changes.
Tryptophan catabolic shunt and serotonin during reactivation stage of T. gondii infection may contribute to the development of depressive-like behavior . In addition, neuronal function and immune-mediated dopamine and serotonin synthesis may be the mechanism of contribution of T. gondii infection to behavioral disorders .
The production of a large amount of dopamine by parasite increases the destruction of the cyst walls and production of tachyzoites . In a study, the treatment of rats with latent toxoplasmosis by means of a dopamine-2 antagonist resulted in the reduction of their risky behaviors .
T helper 1 (Th1), natural killer cells, and nitric oxide interfere in host response to T. gondii infection. The Th1 cells secrete interferon-γ (IFN-γ) and pro-inflammatory cytokines that are important in impeding tachyzoite replication and preventing the reactivation of the tissue cysts of the central nervous system .
Inflammatory cytokines, such as IFN-c, tumor necrosis factor-a, and interleukin 6, are parts of the inflammatory response to T. gondii, leading to the production of glucocorticoids, which affects neuroplasticity . Reduced serotonin production and tryptophan depletion in the brain may contribute to the incidence of depression caused by such cytokines as IFN-γ that could lead to the activation of indoleamine 2, 3-dioxygenase [64, 65].
Activation of guanosine-triphosphate-cyclohydrolase-1 by IFN-γ and other pro-inflammatory cytokines (e.g., IFN-α and IFN-β) increases the production of neopterin, enhances nitrate concentration through the production of tetrahydrobiopterin, and decreases phenylalanine and tyrosine levels.
Amine acid phenylalanine has a key role in the biosynthesis of norepinephrine and dopamine. Sleep disturbance, fatigue, and disorders of the gastrointestinal and musculoskeletal systems are associated with decreased dopamine synthesis . In a study, it was reported that the signs and symptoms of depression were ameliorated after the treatment of T. gondii infection although patient was unresponsive to the conventional anti-depressants .
There are several restrictions in our research. One of these limitations was the adoption of different sampling techniques, such as Facebook-based snowball method, in various studies. Investigation of different age groups in different studies was another limitation since individuals with a longer duration of Toxoplasma seropositivity could have a higher level of depression.
The type of studies reviewed is also a limitation of this study as the nature of cross-sectional study did not allow us to investigate possible causal relationships between T. gondii and depression.
Also, in most case-control studies, the case and control groups were matched for age and sex, but cross-sectional studies cannot be matched for age and sex and therefore may be one of the reasons for the difference in the significance level of studies.
In some cross sectional studies, despite having a large population-based study sample, the number of participants with depression was still limited, and the lack of significant association of T. gondii with depression despite higher prevalence in seropositivity and higher serointensity may reflect limited statistical power.
Another limitation was that most of the included studies did not have sufficient information on disease status or severity. In addition, the studies had variable quality. Furthermore, a few studies were conducted on limited population, such as pregnant women; therefore, the results cannot be extrapolated to the general population. As the final limitation, the reviewed studies did not evaluate the effect of Toxoplasma strains on depression.
Based on the results of this meta-analysis, no statistically significant association was observed between toxoplasmosis and MDD. However, according to the results of the reviewed studies, especially those of the case-control studies, the potential role of toxoplasmosis in depression cannot be completely ruled out. Therefore, it is necessary to perform further research to clarify the role of T. gondii exposure in the clinical characteristics of MDD and determine the detailed association between T. gondii and dysthymia or mild and moderate depression. In addition, it is recommended to evaluate the effect of the antibody titers on the association between depression and Toxoplasma.