Blood Brain Barrier Crossing by a Parasite

Visualizing a Parasite Crossing the Blood Brain Barrier

An estimated 30 percent of the world's population is chronically infected with the parasite Toxoplasma gondii. It's known that "Toxo" can affect the brain, even influencing the behavior of its hosts. But scientists have debated exactly how the parasite crosses the blood-brain barrier, a physical obstacle intended to keep pathogens out of the brain.

Now, researchers have identified how the parasite makes its way in. Using a powerful imaging technique that allowed the scientists to track the presence and movement of parasites in living tissues, the researchers found that Toxoplasma infects the brain's endothelial cells, which line blood vessels, reproduces inside of them, and then moves on to invade the central nervous system.

A few different theories have been considered to explain how Toxoplasma could enter the brain. Some believe the parasite squeezes between the barrier cells, while others think the parasite goes directly through a cell. Another idea is the Trojan horse hypothesis, in which the parasite hitches a ride across the barrier while hidden inside an infected host cell.

Researchers used mice that had been specially bred to express a fluorescent green protein in their endothelial cells. They then infected the mice with modified Toxoplasma that expressed a red fluorescent protein.

After a week, they saw endothelial cells in the brain that were infected, as well as evidence that the parasite was reproducing inside those cells. Two weeks post-infection, they saw that parasites appeared in the brain tissue adjacent to the endothelial cells.

In additional experiments, they were able to visualize parasites bursting out of infected endothelial cells, thereby introducing the parasite into the brain.

The researchers also wanted to revisit the Trojan horse hypothesis, to see if, as had been proposed, infected monocytes, a type of immune cell, might be responsible for carrying the parasite into the brain. To test this, the team infected monocytes with a form of Toxo, labeled red, that can't reproduce, then introduced those cells into mice. If the monocytes were indeed acting as a Trojan horse, the scientists would expect to see the parasite breach the blood-brain barrier. But they only saw infected cells within blood vessels, and these cells were not able to cross the blood brain barrier.

To further illuminate the mechanism by which Toxo infects and disseminates through the body, the researchers looked specifically at levels of free parasites, that is, parasites that had not already infected or become engulfed by a host cell.

They were surprised to see that a significant portion, around a third of the mouse's total parasite load, existed as free parasites in the blood. This presence of free parasites was, however, transient. By 10 days after infection, most mice had no free parasites in their blood.

"From a treatment perspective," author said, "that means if a pregnant woman gets infected for the first time, there is a fairly short period of time when the parasite can cross the placenta and affect the fetus. That tells us that targeting these stages in the blood during this narrow window could be effective at preventing congenital transmission."

As a final test to see whether parasites could directly access the brain from the blood, the researchers infected mice with a mixture of normal parasites and mutants that was unable to reproduce, each labeled in different colors. They then showed that only the normal, reproducing parasite made its way into the functional brain tissue.

"This shows that the parasite has to replicate in order to spread from the blood into other tissues," author said. "That could mean a drug that blocks replication could be effective at preventing dissemination."