Picture yourself hovering over an alien city with billions of blinking lights of thousands of types, with the task of figuring out which ones are connected, which way the electricity flows and how that translates into nightlife.
Welcome to the deep brain.
Even in an era rapidly becoming known as the heyday of neuroscience, tracing the biochemical signaling among billions of neurons deep in the brain has remained elusive and baffling.
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"It's a new kind of data that no one has been able to get before – a single kind of cell projecting from one deep brain area to another deep brain area during behavior," said Stanford bioengineer and neuroscientist Karl Deisseroth, senior author of the study published online Thursday in the journal Cell.
Deisseroth's team relied on genetics, fiber-optics and a bunch of female mice.
The Deisseroth lab at Stanford had already pioneered the use of optogenetics in neuroscience, a technique that delivers light through a hair-thin probe to stimulate cells that have been modified with a light-sensitive gene. First demonstrated in 2007, optogenetic stimulation not only changed the scale and precision involved in exploring the brain, it allowed researchers to better discern cause and effect, which often were muddled by conventional imaging and detection devices.
Researchers soon began using the technique widely to manipulate the brain cells of laboratory animals. They discovered that stimulating one brain cell had a profound effect on behavior.
Finding out how this works, however, depended on tracing the connections, or "projections," from the stimulated neurons.
The deep brain is a very "noisy" place. The tiny voltage changes that propagate along axons, the slender fibers that extend from the nucleus of a neuron, are difficult to distinguish. Researchers routinely add fluorescent properties to the calcium ions that help drive these voltage change in axons, so they can "see" large-scale evidence of activity. But no one had been able to track that signal in an axon while an animal reacted to the stimulation.
"It's buried in the noise and it's too small to see in a behaving animal," Deisseroth said. "We've never been able to see it. We've never been able to observe how animals normally use projections."
The Stanford team tried a new trick. The researchers delivered the light at a specific frequency by chopping it up with what amounts to a fancy pinwheel. Since the calcium ions fluoresce at the same frequency as the incoming light, the team designed a device to pick up only that signal. That allowed them to follow the signal in real time while they chronicled the animal's behavior. They call the new technique fiber photometry.
The rest was relatively simple rodent play. Lab members placed the probes in the areas they had altered for optogenetic stimulation, set up the detection instruments, then ran trials to test the mouse's reaction to other mice.
They chose two areas involved in reward-related activity -- the ventral tegmental area and nucleus accumbens -- and looked at neurons that rely on the neurotransmitter dopamine to relay signals. Abnormal activity in those areas has been implicated in drug abuse and depression, but those regions are involved in a multitude of complex behaviors. How they might mediate social behavior was unclear.
When the researchers stimulated a type of neuron in the ventral tegmental area, the mouse's penchant for sniffing a stranger increased, while inhibiting the same neurons dampened its curiosity, according to the study. The mice showed no such interest in novel objects, such as a golf ball.
Fiber photometry allowed them to trace the signal to the nucleus accumbens. Next, they directed their optogenetic stimulation to altered dopamine receptors and identified which type was associated with the mouse's changing behaviors. They had captured the firing of neurons deep in the brain, from the source to the destination.
"Nobody had been able to see this traffic along a projection deep in the brain of a mammal," Deisseroth said. "That had not been possible before."
The techniques by themselves are not likely to lead to treatment, Deisseroth said. They will help researchers dig deeper on a finer scale, perhaps to identify genes or receptors that are unique to a projection. Those discoveries would put neuroscience a big step closer to modifying behavior on a tiny scale.