At age 1, Lillian Courtney could walk by holding onto
furniture, say 20 to 30 words and even string words
together. But her parents began to notice that she'd
become quiet and subdued. Then, her limbs seemed wobbly. In a matter of days, Lily lost almost all motor control and nearly all her language.
Over the next seven months, Lily tried to fight back. "She was screaming, manic, head-banging and hitting, sleeping no more than two hours a day," says her mother, Leslie. "Obviously, it would be scary to lose all your skills."
Lily was eventually diagnosed with Rett syndrome, a neurologic disorder affecting girls almost exclusively, that is marked by a rapid regression in language and motor skills, cognitive impairment and sometimes autism-like behaviors. It typically begins to manifest between 6 and 18 months of age—a time when cognitive and motor development normally surge.
Rett used to be seen as a degenerative disease, with a gradual, irreversible loss of brain cells. But recent research indicates that cells aren't lost, and that the brain is structurally normal. It's the synapses—the points of communication between brain cells—that are unhealthy. At the age when Rett typically strikes, synapses are developing rapidly: New ones are formed, existing ones are strengthened and irrelevant ones are abandoned, based on input the brain gets from the environment. But in Rett syndrome, this process seems to go awry.
"In simplistic terms, the brain works by learning to make synapses," says Omar Khwaja, MD, PhD, Lillian's neurologist at Children's Hospital Boston, who launched the clinical Rett Syndrome Program last year. "When you make a synapse, you have to stimulate it. But if the synapse is intrinsically slow, doesn't react well to its normal stimuli, or is very fragile, or if conduction across the synapse is impaired, it'll disappear."
Nature deliberately sets aside times in development when the brain is "plastic" and able to update its synapses readily, providing the right balance for learning, socialization and motor function. Increasing evidence suggests that this rewiring process is disrupted not only in Rett syndrome, but in a variety of other disorders that develop after birth, such as autism and schizophrenia.
In several laboratories at Children's, researchers are probing the complex pathways that orchestrate synapse development and brain rewiring, seeking ways to make it work again. The most recent work offers hope, hinting that some broken links could be repaired.
A surprising reversal
Synapses can become compromised in many ways. An incoming signal from a neighboring nerve cell may fail to propagate an electrical impulse in the receiving cell. Or this impulse may fail to spark the usual chain of signals that travel to the cell nucleus, turning on genes. Or something may go wrong in the signal chain itself, so that genes that need to be turned on stay silent. As a result, communication back to the cell surface—telling the cell to strengthen a synapse, make a new one, eliminate a synapse or make a different kind of synapse—is lost. (See figure below.)
In Rett syndrome, the component that's broken is a master switch called MeCP2; it controls a group of genes that initiate synaptic changes. Normally, when relevant input comes into the cell, MeCP2 is chemically altered, causing it to turn the genes on. This stimulation is missing in Rett syndrome.
A recent study from Scotland, however, provided tantalizing evidence that Rett symptoms could be reversed—arguing strongly against it being a progressive, untreatable disease. The researchers worked with mice that had the MeCP2 gene disabled, and had developed advanced Rett-like disease. Using genetic tricks, they turned the MeCP2 gene back on in the brain—and saw a striking recovery in the mice.
Could MeCP2 be switched back on in girls with Rett syndrome? Not yet, but Khwaja hopes to launch a pilot trial of a hormone that MeCP2 indirectly regulates, called IGF1, that may be under-produced in the brains of people with Rett syndrome. In the laboratory, IGF1 has been shown to enhance synapse maturation, and in mice missing the MeCP2 gene, treatment with IGF1 ameliorated their Rett-like disease.
Leslie hopes Lillian, now 5½, can take part in the trial, which hasn't yet been funded. In the meantime, Lily is receiving physical therapy and a form of intensive instruction called Applied Behavioral Analysis, involving seemingly endless repetition—a strategy that aims to build up synapses through constant behavioral reinforcement. "The only way she retains any skill, such as turning the pages of a book," Leslie says, "is to do it over and over and over."
MeCP2 and IGF-1 are just two of many factors in the brain that maintain synapses. The Children's laboratory of Michael Greenberg, PhD, who recently became chair of Neurobiology at Harvard Medical School, studies molecules in the cell that collect information at the synapse, process it, and rally more than 300 genes that set synaptic changes in motion. His lab is systematically seeking genes that MeCP2 acts on, which could provide additional targets for treatment, and has also discovered other master switches. Other Neurobiology labs at Children's, such as those of Takao Hensch, PhD, Michela Fagiolini, PhD, and Chinfei Chen, MD, PhD, are studying these molecules to see what happens in cells and in live animals when they're disrupted.
Another master switch, called MEF2, controls about 150 other genes, and appears to be involved in restricting the number of synapses, a process as important as building synapses. "Too many synapses could result in too much competing information—no one strong signal is getting through," says Chen, whose lab is studying synaptic connections in the brain's thalamus by measuring the electrical currents they generate. (The thalamus is a central relay station that transmits most sensory signals to higher brain areas, so it is important to get these connections right.) The wiring may be disrupted in some cognitive disorders, and Chen's early work suggests that thalamic connections are abnormal in mice with Rett syndrome—too abundant and too weak.
Yet another master switch called Npas4, reported in September in the journal Nature, regulates more than 200 genes that tell the brain to start laying down inhibitory synapses, which are essential for proper brain function. At birth, the rapidly developing brain teems with excitatory synapses, which tend to make nerve cells "fire" and stimulate their neighbors. But if the excitation isn't eventually balanced, it can lead to epilepsy; even diseases like autism and schizophrenia are now thought to involve an excitatory/inhibitory imbalance.
"Early excitatory input is important to make first contacts between neurons," says Hensch, who studies how molecules such as those identified by Greenberg's lab affect larger-scale brain circuits. "But then, at the next stage, you need inhibition."
Timing is everything
Four decades ago, a series of historic experiments at Harvard showed that if a kitten had one eye patched at birth, then later had the patch removed, the animal would be blind in that eye. The eye itself was normal, but the brain couldn't see—it had missed the opportunity to be stimulated with input from that eye. But if the eye was patched for the same length of time later in life, there was no vision loss.
These findings led to the principle that brain circuits are shaped by experience—input from the environment—during well-defined times in development known as critical periods, after which the brain is less adaptable. In humans, critical periods occur in infancy and early childhood, and again in adolescence.
Hensch has adapted the classic experiment to demonstrate the importance of inhibitory synapses in launching critical periods. Studying mice reared in the dark, he and his colleagues recently showed that a factor from the eye, known as Otx2, launches critical periods in the visual system by triggering maturation of certain brain cells that form inhibitory circuits (see figure at right). More recently, focusing on factors that help establish the excitatory/inhibitory balance, Hensch and Fagiolini successfully reactivated critical periods in adult mice.
These studies involved the visual cortex, but Hensch believes that controlling the timing of critical periods elsewhere in the brain could ameliorate disorders such as autism, schizophrenia and attention-deficit hyperactivity disorder, in which critical periods for cognitive, behavioral and language development may be inappropriately accelerated, delayed or prolonged. The brain thus may be rewiring itself at the wrong time, or missing opportunities to rewire when it's getting optimal stimulation.
"If a critical period starts too early, brain function could be frozen in an immature state," says Hensch, who last fall won the highly competitive National Institutes of Health Director's Pioneer Award in 2007. "In children with autism, for example, there's a shunning of sensory input and social interaction just at the time when the brain is looking for this information to set up the circuitry. People with autism may also exhibit savant-like skills—and this may reflect a hyperdevelopment of some parts of brain, while other parts of the brain are slow to get started."
Insights from the Middle East
A recent gene-mapping study from Children's, coupled with findings from Mike Greenberg's lab, provides some of the strongest evidence yet that autism stems from disruptions in the brain's ability to process experience—and that some of these disruptions might be reversible.
A team led by Christopher Walsh, MD, PhD, Children's chief of Genetics and a Howard Hughes Medical Institute investigator, studied 88 large Middle Eastern families with an inherited form of autism, likely owing to marriages between cousins. (Such marriages increase the likelihood of a child inheriting a rare recessive disorder.) To confirm diagnoses made by local clinicians, Walsh's team flew to sites in Turkey, Dubai, Kuwait and Saudi Arabia. They then compared the DNA of family members with and without autism, using a technique called homozygosity mapping to find recessive mutations.
The head-to-head analysis pinpointed five rare, inherited chromosome deletions in the families with autism, which varied from family to family and affected at least a half-dozen identifiable genes, none of them previously known to be involved in autism. Walsh compared notes with Greenberg, and found that several of the genes were on his colleague's list of genes affected by experience. The genes' functions vary, but all seem to be involved in different aspects of synapse formation or refinement.
"These findings suggest the fundamental piece of autism may be this experience-dependent learning," says Walsh. "You can get to the same place—autism—by disrupting maybe 100 different genes that disrupt the same mechanism, but in different ways."
Most intriguingly, only one chromosome deletion Walsh identified actually removed a gene. In most cases, what was lost was a region adjacent to the gene that contains its "on/off" switches—raising hope that therapies could be developed to reactivate the gene.
"We know that autism mutations can disrupt proteins in the synapse, or disrupt the gene that's supposed to be turned on by activity," says Walsh. "But another way you can get autism is by disrupting the DNA that controls the gene. The gene itself is fine; if we could just figure out a way to turn it back on, that would be a tremendous treatment."
Back to school
These recent neuro-genetic discoveries suggest that Rett syndrome, autism and other "disorders of the synapse" are dynamic and amenable to change. Such advances bolster the case for early, intensive intervention in autism, and have fueled Khwaja's efforts to convince school administrators to place girls with Rett syndrome in true educational settings, rather than "holding pens" with limited stimulation.
This fall, Lily started regular kindergarten, accompanied by an aide who helps her communicate (she speaks just a few words, and can sign about a half-dozen more). Through Children's Center for Communication Enhancement in Waltham, she's also trying a computer system that allows her to communicate with her eyes.
"We know the possibility of reversal is there," says Leslie, "so we can't back off on education. We have to prepare her for a day when there may be a breakthrough."