The Universe Within (11 page)

The rats clearly had set times of activity and rest and of hunger and satiety. Richter began to fiddle with things a bit. Once he had a reasonable idea of their activities, he started to leave the lights on in the lab overnight. Then he would turn the lights off for days on end. The activity patterns of Richter’s rats stayed on track. The rats responded in the way the caveman
Siffre did years later.

With that simple observation, Richter saw in these rats
the question that was to become his lifelong obsession: What was the basis for these innate daily rhythms? What controlled them?

Richter turned to studying blind rats. With no obvious cues to perceive light and dark, these animals retained set daily patterns of sleep and feeding. He postulated that there must be something inside, some clock that keeps things ticking along. If so, where in the body did it reside?

The first candidates were the glands that secrete hormones controlling heart rate, breathing, and other bodily functions. Richter gave the rats chemicals to disrupt their hormone balance, then altered their diets, and later he even took out the glands to see what happened. He performed more than two hundred of these kinds of experiments over the years. And in every case, he got the same result. No matter which part he removed, or which hormone was blocked, daily patterns of activity just ticked along normally.

Richter turned his attention to rats’ brains and nerves. He systematically lopped off pieces of the rats’ brains to see the effects. In 1967—forty-eight years after his adviser’s gift of the cage—Richter removed one little patch of tissue at the base of the brain. Excision of a fleck of cells not much bigger than a grain of rice obliterated virtually all of the rhythmic behaviors in the rats. The rats’ internal clocks were in a tiny field of cells right behind the eyes, later named
Richter’s patch.

How could a bunch of cells keep time? Answers to that question were to elude Richter and his brute-force approach of removing pieces of the body. The key came from an entirely different approach to science.

Genetic
mutants are the stuff of biology: six-toed cats, two-headed snakes, and conjoined goats are not just curiosities; they
have the power to tell us the fundamentals of how bodies are built and how they work. While
mutants tell us which genes have gone awry, their scientific importance lies in how they reveal what normal genes can do.

Let’s say you find a mutant animal that lacks eyes. Clearly there is some genetic difference between mutant and normal animals that causes eyes to be missing. There is a lot of work to be done to isolate the actual genetic defect. But by doing breeding experiments between the mutants and the normal animals, you can ultimately identify the gene itself and, if you are really persistent, discover the actual stretch of DNA involved. Knowing the DNA is a key to understanding the molecular machinery that controls how eyes are built. The same is true with virtually every gene that acts to build bodies.

Scientists have scoured populations looking for mutants. Careers have been made, and
Nobel Prizes won, by cataloging mutants or by finding just the right one with an extra toe, different jaw, or oddball eyes, limbs, or heart. While the rewards can be high, this approach is often like rolling the dice. Some mutations crop up only once in every 100,000 individuals. Unfortunately, the creatures we are most interested in,
mammals like us, are the hardest to work with, because they take a longer time to develop than other creatures, they take more resources to rear, and they spend most of the critical phases of their development hidden from the outside world inside the female.

Because of these difficulties, for the past hundred years, flies have been the best animals in which to study genes. As opposed to mammals or reptiles, for example, you can get lots
of flies: they reproduce and develop fast. With such a steady supply of embryos and an entire community of scientists doing the research and sharing data on them, an enormous number of different kinds of mutants have not only popped up in labs but been cataloged, described, and stored for others to study.

But why wait for nature to produce mutants when you can make them? The process of mutating DNA is relatively straightforward, on paper at least. First, you take some
flies and treat them with chemicals or radiation that interrupts the copying of their DNA. With altered DNA, a variety of mutants emerge in the next generation. In flies, the wait from one generation to the next is only about a day. Then scan the mutants for whatever trait you are interested in. Some will have bizarre legs, others mutant eyes, and so forth.

By the late 1960s,
Seymour Benzer’s laboratory at Caltech had become a hub for the study of mutant flies of all sorts. A young student,
Ronald Konopka, entered with a plan and the desire to take some risks making mutants. Benzer’s lab was interested in looking at behaviors, and by the time Konopka set to work, Benzer’s group had already accumulated mutants with odd reproductive and courtship dances.

Konopka had a plan to use this same approach to unravel the genetic workings of the clocks in the body. Smart money at the time was against his success. Most scientists thought the body’s clock was just like a stopwatch and way too complex to dissect easily.

Konopka had come to the right place to make inroads into this problem. Seymour Benzer had a personal motive to understand body clocks. He was a notorious night owl, working late hours in the lab, yet his wife was in bed soon after dinner. Perhaps mutant flies could provide help to a marriage where the partners interacted with each other only at dinnertime.

Konopka spent what must have seemed like a near eternity looking at developing flies. Flies hatch from eggs as maggots, feed for a bit, then develop hard cases around their bodies from which they later emerge as metamorphosed flies. Flies emerge from their cases in early morning, at the time of day when it is normally coolest. This behavior is a manifestation of their internal
clocks; flies reared in
artificial light-dark cycles will always emerge at the end of a dark cycle, when their
brain is wired to expect the cool of dawn.

After about two hundred tries at making mutants, Konopka found one batch of flies that were completely messed up: some emerged too early from their cases, others too late, and still others at random times during the day. The differences in time emergence almost certainly reflected some sort of genetic defect. Here was a defect that reflected a possible clock mutation.

The lab bred each type of fly selectively with another of the same type, making whole family lines of the different mutant flies. With these lines, Konopka and Benzer set the stage for understanding the molecular machinery that keeps time.

Like any good gene, Konopka’s makes a
protein that does the real work in the body. Knowing the gene means that you can ask the important question: Where and when is the protein active?

In normal flies the amount of protein peaks in the early night, then drops to almost nothing during the day. Knowing this normal
rhythm allowed Konopka and Benzer to make sense of the mutants. In the early hatchers, the protein levels peaked early. In the late hatchers, they peaked late. And what about the proteins in the flies with no rhythm? No working protein. The activity of the protein matched exactly what you would expect from one that was behind the daily rhythm.

By now a number of other labs were digging into the problem. Using powerful technologies of fly genetics, they isolated the DNA and found a number of other genes that work in this system. With each new gene, a finer understanding of the fly’s biological clock emerged.

A clock, even the biological one that Konopka and Benzer found, works something like a pendulum that swings back and forth at a set pace. The regular movement provides a basis to measure units of time. Looking at the clocks inside bodies means seeing
molecular equivalents of pendulums inside cells. Here, the major timekeepers come about from chemical chain reactions in which each step occurs at a rate set by the laws of physics and chemistry. When DNA is activated, it makes interacting proteins that are transported through the cell and perform a number of tasks, one of which is to activate portions of DNA again, letting the cycle begin anew. This pendulum-like swing of chemical changes is defined by how fast proteins are made, combine, travel through the cell, and ultimately interact with DNA.

Mutant flies opened up not only a way to see genes that control biological clocks but also a new way to think about people.

About ten years after Konopka and Benzer’s discovery, a graduate student,
Martin Ralph, was setting off to see if there were genes in
mammals that controlled body clocks. He and his adviser in Oregon were working with Syrian
hamsters, running
them on treadmills to monitor their activity. Normal hamsters had rest and activity times, measured by how long they ran on a treadmill, that totaled to about twenty-four hours.

When each new shipment of hamsters came to the lab, Ralph would run them on his treadmills, hoping for some oddball pattern of rest and activity to emerge. He was making a huge bet that a
mutant would randomly show up in one of his weekly shipments.

One day Ralph took the newly received batch of hamsters, put them in the cages, and let them run. To his joy—and relief, no doubt—one individual had a total daily rest and activity cycle that was dramatically different from twenty-four hours. This one hamster functioned on a twenty-two-hour cycle. When he bred the hamster with others, he found the offspring also had the shorter daily rhythm of rest and activity. The aberrant
clock of this hamster was caused by a mutant gene.

When Ralph and others looked in more detail at the activity of the gene, they saw that the major effect of the mutant was a little patch of cells inside the brain. The mutation affected cells inside
Richter’s patch. Ralph’s bet had paid off.

The plot thickened in the early 1990s when a patient entered a
sleep clinic in Salt Lake City complaining of an odd problem. She got a regular eight hours of sleep each night. But no matter how hard she tried to stay awake, she always fell asleep at 7:30 p.m. With a normal eight hours of sleep, that meant she was waking at 3:30 a.m. Her clock ticked at a normal pace, only everything was shifted. Sound familiar?

Entering a sleep clinic means being probed, outfitted with patches, and connected to machines as you try to slumber, all in an effort to put some numbers on bodily functions—breathing, heart rate, body temperature, and so on—during the night. Every test showed that there was nothing strange about this woman’s health or behavior.

Shortly after the testing began, the patient revealed the critical
fact that other
family members shared the same sleeping timetable. There were several puzzles to the family’s sleeping habits. Other members of her family rose very early, but they lived in different places. One fact after another led the researchers to suspect that the cause didn’t lie in their homes, diets, or local environments.

The researchers assembled a full pedigree, mapping the family tree against which relatives rose early and which did not. Then the pattern emerged: here was a classic
genetic trait, just like the fly
mutants.

Looking at a family tree doesn’t reveal the actual change to DNA itself. That level of understanding requires taking cells from the body, by swabbing the cheek of each family member, and comparing the DNA inside the cells between those who rise early and those who wake at more normal hours. If there is a genetic difference, it should be seen in some part of the DNA that consistently varies between the risers.

Isolating the DNA of the early risers and comparing it with that of the normal sleepers showed that the altered sleeping pattern was caused by a very precise change to one gene. Genes and proteins leave their fingerprints everywhere. Once you know the sequence and structure of a gene or protein, it is a relatively simple matter to search for them in other cells. This trail led from the DNA of mutant human early risers, to that of
Martin Ralph’s early running Syrian
hamsters, ultimately to the gene that caused
Konopka and
Benzer’s
flies to hatch early. Each species has its own specialized protein interactions inside the clock, but the basic genes and principles that drive the molecular pendulum are all there. Flies, hamsters, and people share parts of their clocks, their genes, and their histories.