Scientist sheds light on workings of internal clock

By Chris Emery, Baltimore Sun, August 21, 2006

Andrea Meredith's mice have a terrible sense of timing. "It's as if they can't tell the difference between day and night," said Meredith, a neuroscientist at the University of Maryland School of Medicine.

Usually, rodents roam at night and sleep all day, even if kept in total darkness. But keep Meredith's mice in the dark, and they will hop onto their exercise wheels regardless of the hour.

The odd behavior is the result of a change that Meredith engineered in a microscopic structure of their brains. The neuroscientist has been exploring possible links between the structure of cells known as "clock neurons" and specific behavioral patterns in the mice, experts say.

What she has found sheds new light on the part of the brain that acts as an internal clock, generating circadian rhythms in animals and humans. It may be a key step in understanding and treating sleep disorders, jet lag, depression and other human health problems linked to errant daily rhythms, experts say.

"People want to know, how does that internal clock tell the rest of the organism how to operate?" said Jennifer Loros, a professor at Dartmouth Medical School, who studies circadian rhythms in fungus. "This begins to connect the link between rhythmic [nerve activity] and rhythmic behavioral output."

The first documented evidence of circadian rhythms dates to 1729, when French scientist Jean-Jacques d'Ortous de Mairan found that mimosa plants kept in a dark closet still opened their leaves during the day and closed them at night.

Later, similar self-generated rhythms were found in humans and most animals. Blood pressure, body temperature, digestion, libido and other basic functions in humans and mice vary according to a near 24-hour daily cycle. (Such patterns have not been found, however, in cave fish and other creatures that never see the light of day.)

There are practical reasons to have what amounts to a self-winding internal clock.

"If you have a little field mouse living in its burrow, it doesn't have any light, but it still needs to know when to go up and seek food," said Meredith. "If it goes up at the wrong time, there are predators there and it gets eaten."

In recent decades, researchers found that circadian rhythms in mammals are generated by the suprachiasmatic nucleus (SCN), a small region of the brain composed of clock neurons.

One key study by University of Oregon scientists involved golden hamsters, whose internal clocks usually follow a strict 24.1-hour cycle. When they replaced the SCN of a normal golden hamster with one from a mutant hamster that followed a shorter 22-hour schedule, the previously normal rodent adopted the mutant schedule.

Sunlight usually helps a creature's biological clock keep time. Light-sensitive nerves in the eyes connect to the clock neurons and match internal time to the outside light cycle. This tie to the environment becomes evident when a person switches time zones quickly and jet lag sets in.

"The light-dark cycle shifts, but the clock hasn't shifted yet," Meredith said.

Experts believe the brain's ability to generate its own rhythm, even in the absence of light, stems from a "core clock," a cycle in the clock neurons' genetic machinery that repeats about every 24 hours.

While scientists know where in the brain the clock is located and think they understand the genetic mechanism of the core clock, little is known of how the clock controls behavior, experts said.

Meredith decided to study this problem after moving to Stanford University in 2001 to take a position as a postdoctoral researcher. Meredith, now 32, received a bachelor's degree in biology at the University of Maryland, Baltimore County, at age 20, and a doctorate at the University of Texas Southwestern Medical School, in Dallas.

At Stanford, Meredith studied tiny gates, known as ion channels, embedded in the outside wall of the clock neurons. Ion channels allow electrically charged ions such as sodium and potassium to flow in and out of the neurons. This flow forms the basis of brain function by allowing electrical signals to travel quickly from one cell to another.

Meredith specifically studied gates known as BK channels that control large flows of potassium out of the cell. Prior research suggested that BK channels were controlled by the core clock, so it made sense to Meredith that the channels might play some role in the generation of circadian behaviors.

To make the connection, Meredith, a self-described scientific jack-of-all-trades, brought a mixed bag of new scientific tools and techniques to an old problem.

The first step required her to combine genetic engineering with behavioral science. She engineered mice that had no functioning BK channels in their clock neurons, then watched how they acted. When exposed to light, the engineered mice behaved just like those with BK channels: They ran on their wheels at night and slept during the day. But when the engineered mice were kept in the dark they went haywire.

"We saw a very dramatic difference," Meredith said.

Their strict schedule loosened and they roamed their cages and ran on their wheels at erratic times. Their daily amount of activity stayed about the same, but it was spread out more evenly over the 24-hour period.

Meredith had, for the first time, established a link between specific circadian behaviors in the mice and an ion channel on clock neurons.

But the jump from cellular structure to behaviors was a large one. She still needed to find out how her engineering had affected the intermediate step in the process: the electrical signaling in the brain. This phase of the study required her to switch gears and study the electrical properties of nerve cells.

She found that the clock neurons in her engineered mice generated signals different from those in normal mice. Moreover, the odd patterns of signaling corresponded to the odd patterns of behavior in the mice.

Meredith and her colleagues at Stanford had connected the dots between the BK channel and mice behavior patterns. In the engineered mice, the core genetic clocks seemed to be working fine, but the clock appeared to be unable to communicate well with the parts of the brains where actions such as wheel running were generated.

"The signal for time is no longer being transmitted to the legs," Meredith said. "It's like you have an actual clock and you put a piece of tape over it so you can't see the dial anymore."

The results of her study were published in June in the journal Nature Neuroscience. Her work appears to be the first to link a specific cellular structure in the clock neurons to specific circadian behaviors patterns they generate, said Roberto Refinetti, a psychologist at the University of South Carolina and editor of the Journal of Circadian Rhythms.

"The clock itself has been much-studied," Refinetti said. "The novelty of this is being on the output side."

Understanding the cellular mechanisms involved in the control of circadian rhythms could open the door to fixing problems linked to the daily cycles, Refinetti and other experts say.

People's core genetic clocks may be idiosyncratic, research suggests, giving credence to the idea of "night owls" and "morning people." In fact, most people, if isolated from sunlight and other external timing cues, will soon switch to a near 25-hour cycle. Most people thus find it easier to adjust when traveling west than east - it's easier to cope with a longer first day of travel than a shorter one.

Many suffer from sleep disorders because their internal clock is out of sync with the daily schedule imposed by society, work and family.

"With many sleep disorders, if you could choose your own work time you wouldn't have the problem," said Refinetti.

Therapies that target the clock neurons, he said, could help night owls adjust to waking at the crack of dawn or help morning people work the graveyard shift.

Meredith, who accepted a position as an assistant professor at the University of Maryland this summer and is continuing her research there, hopes her work will open the door to therapies for other problems related to circadian rhythms, such as obesity and heart attacks, which often occur in the morning and correspond to hormone-induced increases in blood pressure.

She gave jet lag as another example. Research suggests that people who fly across time zones adjust to their new environment at an average rate of about one day per hour of time change.

"Maybe you could take a drug and cause yourself to shift through jet lag much faster," she said.

Mental disorders and cancer have also been linked to dysfunctional biological clocks, said Dartmouth's Loros.

"There are so many things in the human body affected by circadian rhythms," she said. 'This is just the tip of the iceberg."

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Hypnogogia

"O Sleep, rest of all things, mildest of the gods, balm of the soul..."

(Iris to Hypnos. Ovid, Metamorphoses)