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Disordering Brain Gives Clues To Brain Disorders: "Knock-Out" Mice Provide Evidence On How Cerebral Cortex Forms

Harvard Medical School

BOSTON--The brain's cerebral cortex has endured decades of poking, slicing, and imaging by scientists--yet has largely remained a black box. Gradually, however, the part of the brain that defines us as human is yielding its secrets to scientists who are trying to pry the box open.

In the January 1997 Neuron, Harvard Medical School researcher Li-Huei Tsai and her colleagues report that mutant mice lacking a certain gene fail to weave the neatly layered pattern that is the trademark of the cerebral cortex. It is the latest in a recent string of findings that put within reach one of the most coveted prizes in neuroscience research: understanding how the cortex unfolds during embryonic development.

The study represents a big step in researchers' efforts to decipher the molecular codes neurons use to communicate with their environment during the formation of the cortex. It may also help reveal the genetic cause of several poorly understood human brain disorders.

The cerebral cortex--the control center for all cognitive, perceptual, and higher motor functions--arises in an embryo according to an intricate, still mysterious plan. In the mouse, where this plan has been scrutinized, the millions of neurons needed to populate the cortex are born over the course of seven days deep inside the brain. From there, successive waves of immature neurons set off toward the brain's surface. Each new group crawls past the older neurons and settles on top of them, just beneath the brain's exterior. Researchers have marveled for years at how the cells gradually build up, from the inside out, a pattern of six orderly layers that mark the mature cortex.

"This process amazes me, but in terms of its molecular control, no one has the faintest idea what is going on," says Tsai, assistant professor of pathology at Harvard.

Like many young researchers who identify the proteins responsible for building biological forms beautifully described by earlier, anatomical analyses, Tsai entered the field via molecular biology. She and others discovered a protein, p35, that occurs only in the nervous system. Better yet, it appears during development and almost vanishes in adulthood; and in cell-culture experiments, it proved necessary for embryonic neurons to migrate.

In the present study, Tsai and her coworkers destroyed the gene encoding p35 in mouse embryos. While appearing normal at first glance, the grown mice behaved slightly abnormally. They were slow, docile, and walked somewhat unsteadily. But most importantly, they suffered fatal seizures.

When the researchers examined the brains of the mice, they were astounded to find nondescript cerebral cortices, devoid of the six distinct layers, says Tsai.

Further analysis showed that the genetic manipulation derailed the newborn neurons' journey to their proper places. Instead of traveling past areas of older neurons, the youngest neurons tended to stop underneath older ones In addition, the wiring of neurons is jumbled in the p35-deficient mice. Their corpus callosum uniting the brain's two hemispheres is barely existent, while other nerve fibers course through brain areas where they do not belong. All this suggests that p35 somehow enables neurons to migrate to the right places and extend axons to make the right connections.

This study may allow previous findings--some going back almost 50 years ago--to fall into place. The completed picture would perhaps show a cascade of events that allows migrating neurons to "read" molecular signposts in their environment and respond to them by turning or stopping. The earlier work includes the description of mutant mice, dubbed reeler, that have similar brain disturbances. Recently, researchers cloned this gene and suggested that its protein might occur near the surface of the embryonic brain, where the youngest neurons stop. Other scientists described a mouse mutation called scrambler, which might play a role in the same signaling cascade. And over the past ten years, scientists have identified additional molecules that guide neuronal migration.

But all of these molecules are outside of cells or on their surfaces, revealing nothing about how neurons process these instructions. Tsai's study is exciting because it provides the first toehold inside the neuron. Moreover, p35 is a protein of known function: It is the activating partner of an enzyme that performs a well-studied biochemical reaction on other cellular proteins. That should make it easier to identify molecules from which p35 receives its cues and others on which it acts, says Tsai.

Interestingly, several human brain disorders have characteristics that lead scientists to suspect that the genes at fault may be involved in the same signaling interactions Tsai studies. One such disorder is lissencephaly. Most children suffering from it are mentally retarded and die of seizures in the first years of life. Two other conditions, called periventricular heterotopia (PH) and double cortex (DC), also appear to stem from defective migration of cortical neurons. The genes for PH and DC are being cloned by Chris Walsh, assistant professor of neurology at Harvard Medical School.

With all the leads provided by mutant mice and human conditions, Tsai hopes that researchers will sort out within the next few years just how neurons navigate the ever changing environment of the developing brain.

This work was funded by the National Institutes of Health.

The authors of the Neuron article, in addition to Tsai, include Teresa Chae and Young Kwon, both of Harvard Medical School, Pieter Dikkes of Children's Hospital, Boston, En Li of Massachusetts General Hospital and Harvard Medical School, and Roderick Bronson of Tufts University, Boston.

Editors, please note: Li-Huei Tsai will be available for interviews at Harvard Medical School.

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