September 7, 1999

SCIENTIST AT WORK / Dr. Joe Z. Tsien

Of Smart Mice and an Even Smarter Man

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By NICHOLAS WADE

PRINCETON, N.J. -- A certain amount of disorder has broken out around Dr. Joe Z. Tsien, the biologist who announced last week that he had created a smarter strain of mouse by genetically altering a gene for memory.

	Laura Pedrick for The New York Times
	Dr. Joe Z. Tsien with a supersmart mouse he genetically engineered in a conference room at Princeton University. The research may shed light on human intelligence.

Patients call seeking help. Individuals of enhanced imaginations warn that the mice may escape and take over the planet. Television crews patrol the halls. His voice-mail box has overflowed.

But Dr. Tsien, seemingly the only scientist on the Princeton campus who, on a warm summer day, is wearing a tie, ignores the chaos and a phone that rings every couple of minutes. In soft tones he describes the remarkable journey that has led him from Wuxi, a small town near Shanghai, to the position of having made a significant, maybe decisive, contribution to understanding the nature of memory and intelligence.

Dr. Tsien (pronounced chee-YEN) says he did not begin to consider the wider implications of his work until just before his article was published. He engineered his smarter mice for purely academic reasons, to address and perhaps solve the question of how memories are laid down in the brain.

But the mice turned out to be smarter as well as having better memories, lending an unexpected new dimension to the experiment.

Although many arguments with psychologists doubtless lie ahead, Dr. Tsien believes that learning, memory and intelligence are all intimately related because, as his smarter mice demonstrate, "a common unifying mechanism underlies them all."

And because mice and people use the same basic mechanism of memory, the smarter mice could well shed much light on the nature of human intelligence.

Dr. Tsien's result, as he is the first to note, rests on knowledge and techniques developed by other scientists.

He describes his experiment as "obvious" -- at least in retrospect. His achievement lies in the fact that, in a highly competitive field of biology, he was the first to conceive of the experiment and to see that it could be decisive.

He also carried it out in a particularly convincing way. "Extremely nicely done," was the verdict of Dr. Eric R. Kandel, a leading biologist at Columbia University and the former laboratory chief of Dr. Tsien.

The idea that led to the smarter mice was no lucky break. Rather, it was a feat for which Dr. Tsien had been preparing intensively for many years, including seven years of postdoctoral education.

In Wuxi, where his father was a clerk and his mother an accountant, he was the only person to enter college from his high school, one attached to a fabric plant. But the college was a good one, the East China Normal University in Shanghai, and he decided to do doctoral studies in the United States.

"In 1986, China was still very closed, so we really had no idea about the United States," Dr. Tsien says in describing how he picked a college. He chose the University of Minnesota because it offered to waive the application fee, which he could not afford, and because the Chinese characters for Minnesota translated invitingly to "clean air blue sky."

Having recovered from the surprise of finding the clean-air-blue-sky state so cold, he developed an interest in neurophysiology and the instruments then available for monitoring the electrical signals transmitted by brain cells. "I got fascinated by seeing a nerve cell fire. They are talking -- what does that mean?" he says.

A long apprenticeship was necessary before he could begin to parse that language. He did his Ph.D. thesis with Dr. Lester R. Drewes of the University of Minnesota, helping him conduct studies under a Defense Department grant on how the warfare agent sarin blocks the transmission of nervous signals.

Receiving his Ph.D. in 1990, he was accepted as a postdoctoral student by Dr. Kandel's laboratory.

There he worked on identifying genes that are active in rats' brains during memory formation.

"I got a more systematic education in neuroscience. I got to see how a big lab operates," Dr. Tsien said.

He then moved to another leading neuroscience laboratory, that of Dr. Susumu Tonegawa at the Massachusetts Institute of Technology. Dr. Tonegawa won the Nobel Prize in Physiology or Medicine in 1987 for research on the genetic control of the immune system, and later switched to the study of learning.

In Dr. Tonegawa's lab, Dr. Tsien worked with so-called knock-out mice, animals from which a gene has been deleted.

The idea is to learn what a gene does by excising it and seeing what defects the mouse develops. He became interested in the brain cell component, known as the NMDA receptor, suspected of being central to the memory mechanism. The receptor consists of parts made by several genes, the chief part being specified by a gene called NR1.

	Princeton University
	A mouse whose memory was enhanced through genetic manipulation took a learning test recently in a laboratory at Princeton University.

Using advanced genetic techniques, he decided to create a mouse lacking the NR1 gene in the cells of its forebrain.

Creating the mouse took two and a half years and, for a postdoctoral student, was a substantial risk. If the experiment failed, there would be no result worth publishing.

In the end, he was able to knock out the gene in just the cells of the hippocampus, a brain module dedicated to learning and much studied by neuroscientists. "I think a god looked on me very kindly," he said, referring to the element of luck in creating such a valuable research tool.

The mice lacking the NR1 gene in the hippocampus indeed did not remember as well, suggesting the NMDA receptor is important in laying down memories. But the experiment, published in December 1996, was regarded by other experts as less than fully conclusive, because the absence of the NR1 gene could have caused general brain damage not specific to memory.

Dr. Tonegawa became very interested in the mouse, as did Dr. Kandel, because Dr. Tsien had made it with a technique developed in Dr. Kandel's laboratory. The two lab chiefs were also interested in receiving due credit, and discussions ensued between them that were stressful for him, Dr. Tsien recalls.

However, he now had sufficient credentials to set up his own laboratory. "After working with these two powerful people, I wanted to be free," Dr. Tsien says. Two years ago he was appointed an assistant professor at Princeton and was able to set up his own lab. He began to think about how he might try to improve a mouse's memory, rather than sabotage it, because such an experiment would run far less risk of being criticized as nonspecific.

The focus of his thinking was the anatomy of the NMDA receptor. The intricate biological device is shaped like a cylinder or doughnut embedded in the outer wall of certain brain cells.

Usually its central channel is firmly closed. But when the nerve cell receives signals from two other nerve cells at the same time, the NMDA channel springs open, allowing a current to flow into the cell. This current generates a long-lasting change within the cell, making it much more responsive the next time that either of the two other nerve cells is active alone.

This property of the NMDA receptor -- opening when two signals arrive simultaneously -- has long been suspected to be the basic mechanism of memory, because it is a way for the brain to make an association between two events. The exact degree of simultaneity turns out to be very important. In young mice, two signals can arrive as much as one-tenth of a second apart for their coincidence to change the nerve cell. In older animals, the NMDA receptor allows a much narrower window of time for an association to register.

Another known fact about the receptor was that its composition changed with the age of the animal. Its main component is the gene product of NR1, which Dr. Tsien had knocked out in his memory-deficient mice. But the NR1 component works with any of four different partners, which modulate its activity in different ways.

Two of the partners, known as NR2A and NR2B, are particularly important in cells of the forebrain. As animals age, there is a switch from NR2B to NR2A as the preferred partner for NR1.

Abilities in many animals decline after sexual maturity.

Song birds cannot learn new songs. The human mind becomes less flexible at learning new languages. "I am always stuck with my Chinese accent, but if I had come to the United States 12 years earlier I would have learned perfect American," Dr. Tsien says by way of personal example.

As he contemplated these various pieces of information, Dr. Tsien said, it seemed clear that they were related. The natural switch with age of NR1's partner must underlie the increasingly stringent requirement for two signals to arrive simultaneously, and the narrowed window of time must be the reason why older people find it harder to make associations.

But no one had specifically stated the idea in those terms, as far as Dr. Tsien knew. And certainly no one had done the obvious experiment, which was to engineer mice in which the NR2B gene was artificially put into hyperdrive to see if their memories improved.

So Dr. Tsien took a copy of the mouse NR2B gene and linked it to a special piece of DNA, called a promoter, that is active only in cells of the mouse forebrain.

He injected this genetic fragment into fertilized mouse eggs, where it added itself to the mouse's normal complement of genes. Because of the promoter, the NR2B gene was active in cells of the forebrain, adding its product to that produced by the mouse's own NR2B gene.

With all the extra NR2B being produced in the mice's brain cells, the NMDA receptors underwent a subtle but significant change. Instead of staying open for 100 thousandths of a second, as they do in normal mice, the receptor's interval increased to 250 thousandths of a second.

That minute biophysical change, Dr. Tsien says, is what underlies the superior learning skills of the mice. The essence of smartness is an extra 150 thousandths of a second.

Credit for a discovery is often disputed, particularly when the finding is important.

Dabbling with genes, he built a better mouse.

Dr. Tonegawa told reporter for The Star-Ledger of Newark, that Dr. Tsien may have started developing the smarter mice while at the Massachusetts Institute of Technology and accused him of being uncollegial. If the mice were developed in Dr. Tonegawa's lab, M.I.T. would have rights to them and Dr. Tonegawa could exercise a lab chief's claim to a share of the academic credit.

Dr. Tsien said he was "totally surprised" by Dr. Tonegawa's remarks. His smarter mice experiment was conceived and executed entirely at Princeton, he said. Other scientists have patented the NR2B gene but Princeton has filed for a "use patent," the right to use the gene in ways suggested by Dr. Tsien's work.

Dr. Tonegawa, who was traveling in Japan last week, did not respond to a request made through his secretary for an interview.

Dr. J. David Litster, M.I.T.'s vice president for research and the official in charge of disputes over scientific conduct, said that M.I.T. "is not endorsing Tonegawa's claims, certainly not until we know what they are. If it's a dispute between Tonegawa and a former postdoc over credit I don't really think we ought to get involved in that."

Meanwhile Dr. Tsien is waiting to see how the implications of his smarter mice are received by his peers in the neuroscience community and by the public. "To the scientific community this is a small step for a man," he says. "The fundamental question is, 'Is this a giant step for mankind?' "

What does he think? "I don't know," Dr. Tsien replies.