VOLUME 12, NUMBER 19 • APRIL 6, 2001

Researchers Light the Path of the Brain’s Feeding Circuit in Mice

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A novel technique that uses a virus tagged with a green-glowing jellyfish protein has enabled scientists to visualize the feeding circuit in mice. The method may be useful in studies of other complex circuits in the brain. The findings are reported in the March 30 issue of Science by a team of researchers from The Rockefeller University, the Howard Hughes Medical Institute, Princeton University and the University of California at San Diego.

The scientists have shown that key neurons that play a role in regulating food intake and that respond to the hormone leptin also receive inputs from neurons in a number of other brain regions.

"Gross connections between neurons in the hypothalamus have been known for decades," says lead author Jeff DeFalco, a postdoctoral researcher in the Laboratory of Molecular Genetics at Rockefeller. "This new technique is exciting because, for the first time, we can identify circuits involving specific classes of neurons."

Leptin, a hormone that plays an important role in regulating food intake and body weight, is produced mainly by fat cells and signals nutritional information to the brain. In general, an increased amount of fat leads to the production of more leptin and a decreased amount of fat leads to a decreased amount of leptin. An increase or decrease in the level of leptin elicits a set of responses that act to return weight to the starting point. Studies in animals have shown that increased leptin reduces food intake and decreased leptin increases food intake. Leptin exerts these effects by changing the activity of a neural circuit in the brain.

To trace the brain’s feeding circuit, the scientists inserted a green fluorescent protein (GFP) marker, which normally glows green, into a Pseudorabies virus. Pseudorabies virus is an animal virus that will spread from one nerve cell to the next only if the cells are in synaptic contact with one another. In the past, this virus has been used to trace neural circuits. However, the virus normally infects cells indiscriminately and has thus been of limited value for tracing the neural connections of specific nerve cells. In this paper, the authors created a viral strain that would be activated in specific cell types but remain inactive in others. This new strain is activated when it infects a cell that expresses another gene known as the Cre recombinase.

The researchers generated two strains of mice in which the Cre recombinase enzyme was co-expressed in nerve cells that express either NPY or the leptin receptor. These two cell types are known to play an important role in regulating feeding behavior. The leptin receptor is the molecule that receives leptin’s signal, while injections of NPY increase food intake in mice.

Jeffrey Friedman is the Marilyn M. Simpson Professor and director of the Starr Center for Human Genetics at The Rockefeller University.

The researchers next injected the Cre-dependent, GFP-tagged virus directly into a region of the hypothalamus called the arcuate nucleus. This is a brain region where NPY and the leptin receptor are expressed. After injection, the virus first spread throughout the cells of the arcuate nucleus itself, causing the neurons containing the Cre recombinase gene to glow green (from the GFP). The virus then spread from these cells to other cells that were in contact with it, and so on. As the virus infects each neuron, it amplifies itself, allowing the researchers to trace the pathway of neurons leaving the hypothalamus.

By looking at sections of the mouse brain taken at various times after infection, DeFalco and his colleagues were able to establish a hierarchy of neuronal signaling. They found that, in addition to sensing leptin levels, the nerve cells in the hypothalamus receive inputs from other brain regions, including sites that play a role in modulating emotion, olfaction and higher brain functions. Further studies of the molecular mechanisms by which these neurons alter the activity of the cells that express NPY and the leptin receptor could lead to the identification of new molecules that regulate feeding.

"The precise delineation of the architecture of the neural system that controls feeding behavior is necessary if we are to understand the molecular mechanisms that control weight," says co-author Jeffrey M. Friedman, Marilyn M. Simpson Professor and head of the Laboratory of Molecular Genetics at Rockefeller and an investigator at the Howard Hughes Medical Institute. "This new method allows us for the first time to directly visualize some of the components of this neural system."

DeFalco’s and Friedman’s co-authors are Hongyan Liu and Xiaoli Cai at Rockefeller; Mark Tomishima and Lynn Enquist at Princeton; and Jamey D. Marth at the University of California at San Diego.

This research was supported by the National Institute of Diabetes and Digestive and Kidney Diseases, part of the federal government’s National Institutes of Health. Friedman is the Marilyn M. Simpson Professor and director of the Starr Center for Human Genetics at Rockefeller.