Mapping the Brain’s Wiring Diagram

Joint Gladstone-Salk study uses high-resolution technique to decode circuitry that guides brain function
Anatol Kreitzer
Anatol Kreitzer, PhD  [photo: Chris Goodfellow]

June 30, 2013: The father of modern neuroscience, Santiago Ramon y Cajal, is so named because of his pioneering observations of the detailed structure of nerve cells, or neurons. Ramon y Cajal took advantage of a technique called Golgi staining. Golgi staining works by “impregnating” neurons with a black dye that infuses every single process extending from the neuron’s cell body. In this way he could trace, with extraordinary detail, where these processes ended up in the brain. His images of different types of neurons—largely hand-drawn—are over 100 years old, but remain some of the most beautiful ever made. And in addition to being beautiful, they were enormously informative, laying the foundation for our understanding of how brain cells connect to each other.

The power of the brain to control the body and behavior lies in its trillions of intercellular connections called synapses. Synapses occur at the end of long, thin branches, called axons and dendrites, which extend sometimes over great distances to communicate with their target neurons. By bringing together functionally distinct regions of the brain, these connections are essential for coordinating all of the complex tasks our bodies carry out—both those in our consciousness, including vision, hearing and voluntary movement, and those we never even think about, such as metabolic processes, heart rate and blood pressure.

One drawback of the Golgi staining method, revolutionary as it was, is that the dye gets picked up by only a small subset of neurons in a given brain region. Thus, it was impossible to trace and map whole networks of neurons. Since Ramon y Cajal’s time, neuroscientists have been developing better and more sophisticated tracing methods to figure out how our brain is wired together. Recently, a team of scientists at the Gladstone Institutes and the Salk Institute combined an innovative brain-tracing technique with sophisticated genetic tools to ask how one brain region connects to its targets. In so doing, they also reveal clues as to what may happen, neuron by neuron, when these connections are disrupted.

Updating the Methods of Ramon y Cajal
Gladstone investigator Anatol Kreitzer and Salk investigator Edward Callaway have given Ramon y Cajal’s methods a facelift. In the latest issue of the journal Neuron, they combined mouse models with a tracing technique—known as the monosynaptic rabies virus system—to assemble brain-wide maps of cells that connect with the basal ganglia, a region of the brain that is involved in movement and in decision-making. Developing a detailed anatomical understanding of this region is important as it could inform research into disorders that can be traced to basal ganglia dysfunction, such as Parkinson’s disease and Huntington’s disease.

“Taming and harnessing the rabies virus, as pioneered by Dr. Callaway, is ingenious in the exquisite precision that it offers compared with previous methods, which were messier and offered much lower resolution,” explained Dr. Kreitzer. “In this paper, we took the approach one step further by activating the tracer genetically, which ensures that it is only turned on in specific cells in the basal ganglia. This is a huge leap forward technologically, as we can be sure that we’re following only the correct networks that connect to specific kinds of cells in the basal ganglia from other parts of the brain.”

Neuron Figure
Neuronal tracing, then and now. On the left is an image hand-traced by Ramon y Cajal, showing neurons and their branches as revealed by the Golgi staining method. On the right are rabies virus–infected neurons, labeled in red/white, whose labeled branches could be traced to generate brain-wide maps of connectivity.
Degeneration of Neurons is Linked to Movement Disorders
At Gladstone, Dr. Kreitzer focuses his research on the role of the basal ganglia in Parkinson’s and other conditions. Last year, he and his team published research that revealed clues to the relationship between two types of neurons found in the region—and how they guide both movement and decision-making. These two types, called direct-pathway medium spiny neurons (dMSNs) and indirect-pathway medium spiny neurons (iMSNs), act as opposing forces. dMSNs initiate movement, like the gas pedal, and iMSNs inhibit movement, like the brake. The latest research from the Kreitzer lab further found that these two types are also involved in behavior, specifically decision-making, and that dysfunctions of dMSNs or iMSNs is associated with addictive or depressive behaviors, respectively. These findings were important because they provided a link between the physical neuronal degeneration seen in movement disorders, such as Parkinson’s, and some of the disease’s behavioral aspects. But this study still left many questions unanswered.

New System Reveals Details of the Brain
The monosynaptic rabies virus system, developed in 2007, uses a modified version of the rabies virus to “infect” a brain region, which in turn infects the neurons that are connected to it. When the system was applied in genetic mouse models, the team could see specifically how MSNs of the basal ganglia connected motor and reward centers in the cerebral cortex, the outermost layer of the brain. And what they found was surprising.

“We noticed that some regions showed a preference for transmitting to dMSNS versus iMSNs, and vice versa,” said Dr. Kreitzer. “For example, neurons residing in the brain’s motor cortex tended to favor iMSNs, while neurons in the sensory and limbic systems preferred dMSNs. This fine-scale organization, which would have been virtually impossible to observe using traditional techniques, allows us to predict the distinct roles of these two neuronal types.”

Where Do We Go from Here?
We’ve come a long way since the days of Ramon y Cajal. Today, neuroscientists can use light to control the activity of neurons, watch how brain function changes in real time using sophisticated imaging techniques, and—as Drs. Kreitzer and Callaway show—harness the power of both genetics and virally encoded tracers to map neuronal connections. Together, these tools will bring us ever closer to decoding the myriad connections in our brains, and how they operate in health and disease.

“These initial results should be treated as a resource not only for decoding how this network guides the vast array of very distinct brain functions, but also how dysfunctions in different parts of this network can lead to different neurological conditions,” said Dr. Callaway. “If we can use the rabies virus system to pinpoint distinct network disruptions in distinct types of disease, we could significantly improve our understanding of these disease’s underlying molecular mechanisms—and get even closer to developing solutions for them.”