For the first time, researchers have developed a way to view stem cells in the brains of living animals, including humans—

The Birth of a Brain Cell: Scientists Witness Neurogenesis
For the first time, researchers have developed a way to view stem cells in the brains of living animals, including humans—a finding that allows scientists to follow the process neurogenesis (the birth of neurons). The discovery comes just months after scientists confirmed that such cells are generated in adult as well as developing brains.

"I was looking for a method that would enable us to study these cells through[out a] life span," says Mirjana Maletic-Savatic, an assistant professor of neurology at Stony Brook University in New York State, who specializes in neurological disorders such as cerebral palsy that premature and low-weight babies are at greater risk of developing. She says the new technique will enable her to track children at risk by monitoring the quantity and behavior of these so-called progenitor cells in their brains.

The key ingredient in this process is a substance unique to immature cells that is neither found in mature neurons nor in glia, the brain's nonneuronal support cells. Maletic-Savatic and her colleagues collected samples of each of the three cell types from rat brains (stem cells from embryonic animals, the others from adults) and cultured the varieties separately in the lab. They were able to determine the chemical makeup of each variety—and isolate the compound unique to stem cells—with nuclear magnetic resonance (NMR) spectroscopy. (NMR helps to determine a molecule's structure by measuring the magnetic properties of its subatomic particles.) Although the NMR could identify the biomarker, but not its makeup, Maletic-Savatic speculates it is a blend of fatty acids in a lipid (fat) or lipid protein.

After pinpointing their marker, the team ran two tests to determine the method's sensitivity and accuracy: First, they injected a bevy of stem cells into a rat's cerebral cortex, an outer brain layer where neurogenesis does not normally occur. They then passed an electric current through the animals' brains; electric currents induce neurogenesis in the hippocampus, a forebrain structure that is one of two sites (the other being the subventricular zone) where new neurons are believed to arise.

After performing each procedure, the team used NMR spectroscopy to capture images of the living rats' brains. There was, however, too much visual interference on the scans to find their biomarker. The researchers called upon Stony Brook electrical engineering professor Petar Djurić to help them come up with an algorithm to cut through the clutter and glean a clear picture of their target compound.

With the analytical method helping to decode their scans, they could clearly see increased biomarker levels in the cortex after a neural stem cell injection. Similarly, after the animals were given electric shocks, levels of the compound clearly went up in the hippocampus.

The team next turned its attention to humans, enlisting 11 healthy volunteers, ranging in age from eight to 35, who each spent 45 minutes in an NMR scanner. Hippocampal scans turned up more of the marker than the cortical images. In addition, the older subjects showed lower levels of the biomarker than younger ones (a finding consistent with earlier studies). "This is the first technique that allows detection of these cells in the living human brain," says Maletic-Savatic.

Fred Gage, a genetics professor at the Salk Institute for Biological Studies in La Jolla, Calif., and co-author a 1998 report in Nature Medicine that announced the discovery of neurogenesis in the adult human brain, praises the new approach. "It seems that they are measuring proliferation rather then maturation based on their results," he says. "It will be important for them to knock down neurogenesis in a mouse and show that [this] signal disappears to confirm the causal link with neurogenesis.

If the new work is replicated and confirmed, it may allow for faster diagnosis and tracking of myriad psychiatric and neurological conditions. Among them: chronic depression. Study co-author Grigori Enikolopov, an associate professor of molecular biology at Cold Spring Harbor Laboratory in Long Island, N.Y., showed last year that antidepressants lead to new nervous system cells, raising questions about the role these cells play in the causation of the ailment.

"Although we are only just beginning to test applications, it is clear that this biomarker may have promise in identifying cell proliferation in the brain, which can be a sign of cancer," Enikolopov says. "In other patients, it could show us how neurogenesis is related to the course of diseases such as depression, bipolar disorder, Alzheimer's, Parkinson's, MS, and post-traumatic stress disorder."


Neurogenesis

Young Cells in Old Brains
The paradigm-shifting conclusion that adult brains can grow new neurons owes a lot to Elizabeth Gould's rats and monkeys.
Reunion weekend at Princeton University, and the shady Gothic campus has been inundated by spring showers and men in boaters and natty orange seersucker jackets. Tents and small groups of murmuring alumni dot the courtyards. Everything proper, seemingly in its place. In Green Hall, however, the same order does not prevail. Elizabeth Gould's laboratory is undergoing construction, and the neuroscientist herself would not be mistaken for an alum: her plaid blue workman's shirt hangs loosely and unbuttoned over a T-shirt and jeans, and she confesses she often feels out of place on the conservative campus.

Against a backdrop of tidy ideas about the brain, Gould and her colleagues have been messing things up and, in the process, contributing to some of the most exciting findings of the past decade. Her work--and that of several other neuroscientists--has made clear that new neurons are produced in certain areas of the adult brains of mammals, including primates. Moreover, these cells can be killed off by stress and unchallenging environments but thrive in enriched settings where animals are learning, and they may play a role in memory.

Until recently, dogma held that mature brains were static: no cells were born, except in the olfactory bulb. One of the cornerstones of this understanding came from studies by Pasko Rakic of Yale University, who examined macaque monkeys and found no evidence of the creation of nerve cells, a process called neurogenesis. The prevailing view has since held that primates--and, indeed, mammals in general--are born with all the neurons they are going to have. Such neural stability was considered necessary for long-term memory. So in the late 1980s when Gould, who was then researching the effect of hormones on the brain as a postdoctoral fellow in the laboratory of Bruce S. McEwen at the Rockefeller University, saw evidence of new neurons in the rat hippocampus, she was perplexed. Gould knew from the pioneering work of Fernando Nottebohm, also at Rockefeller, that neurogenesis occurred in adult birds--canaries and zebra finches, for instance, grow nerve cells to learn new songs--but she and her lab mates knew of no mammalian parallel. "We were really puzzled," she recalls. "It wasn't until we delved far enough back into the literature that we found evidence that new neurons are produced in the hippocampus."

Those earlier studies had never been widely noticed. Beginning in the 1960s Joseph Altman, now professor emeritus at Purdue University, and neurologist Michael S. Kaplan independently recorded neurogenesis in rats and other mammals. They saw growth in the olfactory bulb, in the hippocampus--a region important to memory--and, most strikingly, in the neocortex, which is the part of the brain involved in higher thinking. "But nobody picked up on the results," Gould says. "It is a classic example of something appearing before its time."

In her work with rats, Gould verified that when she altered the normal hormonal bath the hippocampus received, cells died and, apparently to compensate, more cells were born. "That was really the beginning of my interest in neurogenesis and my realization that it happened," she says. "But at that time, to be perfectly honest, I was more interested in solving the puzzle of my own data and not so much into saying, 'Hey, this is a really cool phenomenon that has been overlooked and that has a lot of meaning.'" Her first papers on the phenomenon, published in 1992 and 1993, did not attract much attention.

Gould went on to do experiments clarifying aspects of neurogenesis. She found that stress suppressed the creation of neurons and that lesions in the hippocampus triggered the development of new cells--something she considers significant because it implies that the brain can heal, or be induced to heal, after injury.

In 1997 Gould, who grew up in Huntington, N.Y., moved to Princeton as an assistant professor. Over the next few years she and her co-workers reported that new neurons survived if animals lived in complex environments and learned tasks, findings also documented in mice by Fred H. Gage of the Salk Institute for Biological Studies in La Jolla, Calif. Gould then observed that new neurons are found not just in the rat hippocampus but also in those of marmoset monkeys and macaques. News of neurogenesis in primates, including confirmatory work by Rakic in macaques and by Gage in the human hippocampus, catalyzed widespread interest because it introduced the possibility of repairing the brain and elucidating memory formation.

For Gould, the sudden splash of attention has been disorienting--and she does not relish it, particularly when it takes her away from her experiments. She says she is happiest in the lab, working under the microscope with brain slices, which she finds beautiful and which recall a childhood interest in being an artist. And she has liked being in a quiet field of research, one she chose when studying psychology at the University of California at Los Angeles. "I have no interest in doing experiments that someone else is going to do a month later if I don't get around to it," she says. "You have to pick things to do that are really intriguing to you, things that you are really curious about--not just because you want to publish on them before anyone else does."

Her curiosity is taking her in several directions these days. An outstanding question centers on what role new hippocampal neurons play. Do they establish new circuits or memories? Or do they replace old neurons in established circuits? This year Gould and her colleagues reported that the neurons are involved in the creation of trace memories--memories important to temporal information. "We had evidence that the new cells were affected by learning, and this is evidence that the new cells are necessary for learning," Gould explains. She now intends to do similar studies in marmosets, to see whether her discoveries about rats will prove true for primates.

Gould is also repeating and extending work of a few years ago in which she found neurogenesis in the neocortex of macaques, a finding that remains controversial and that would be highly significant because of the importance of the cortex. Although no one has published a replication so far, William T. Greenough of the University of Illinois says Gould's findings "do not surprise me. We have unpublished data in rats that support the same thing."

In addition, Gould has begun investigating the role of sleep deprivation in neurogenesis, an interest triggered by the birth of her third child last year. "I never really thought about the sleep aspect until I wasn't getting any," she says, laughing. And she is intrigued by the possibility that much of what we have come to understand from laboratory settings may be skewed.

"Our laboratory animals are very abnormal," Gould notes. "They have unlimited access to food and water, and they have no interesting cognitive experiences at all. We know that if you house an animal in that setting, most of its new neurons will die within a few weeks after they are produced." Gould is designing environments that are closer to the ones rats and marmosets experience in the wild, hoping to get closer to the truth about the brain. "It really raises the issue of whether a lot of the things we are looking at are really deprivation effects."

Potentially shifting another paradigm doesn't faze Gould. "There has to be some fresh perspective, something new that you can bring to the work that other people wouldn't see," she says. "Otherwise you are not making a real contribution, and you might as well just step aside and find something else to do."