A method for imaging gene expression in neurons based on their connections has been developed by researchers at Rockefeller University’s Molecular Genetics lab.
The new technique is called Retro-TRAP. It fuses two approaches to studying the brain. The first is mapping all of its connections and the second is gene expression profiling of neuron populations.
“We hope that Retro-TRAP will be broadly used and provide a more granular understanding of how complex neural circuits function and ultimately lead to better treatments for neurological and neuropsychiatric disorders, " said lead author Jeffrey Friedman.
Understanding Neural Circuits
Neurobiologists frequently discuss their subject using electrical metaphors, but the fact is that the brain is nowhere near as simple as wires and circuits. Neurons can actually behave differently depending on the situation.
“Refinements in neuroscience over time have allowed us to explore how the nervous system works in ever-greater detail, and the approach we have developed continues this trend,” says research associate Mats Ekstrand. “By building on existing techniques, we are now able to take a closer look at the types of cells involved in a particular circuit and what they are doing."
These types of insights could someday help explain why some diseases, for example Parkinson’s Disease, affect certain sets of neurons, or make it possible to exactly target treatments at a dysfunctional neural circuit, instead of saturating the whole brain in a drug.
Translating Ribosome Affinity Purification
The new method is a modified form of a technique known as Translating Ribosome Affinity Purification (hence the acronym TRAP), developed at Rockefeller to pinpoint gene expression using green fluorescent protein to tag protein-assembling machines called ribosomes.
Researchers used Retro-TRAP to introduce green fluorescent protein to the neuron, through a virus that travels backwards from a synapse into the body of a mouse neuron. A small antibody was employed to link the ribosome with the fluorescent protein.
Then, with these fluorescent tags, the researchers pulled out the ribosomes and sequenced the genetic messages passing through them. In this way, they produced a list of active genes.
The team concentrated on the inputs to a well-studied part of the brain, the nucleus accumbens. This brain area integrates information from throughout the brain, including regions involved in executive function, memory, depression, reward-related behavior, feeding and other functions.
“We wanted to target a selected number of inputs into the nucleus accumbens because we figured we might be able to get some molecular clues as to why it is important in regulating so many functions,” said graduate student Alexander Nectow.
Molecular Profiles of Neurons
Retro-TRAP enabled them to create molecular profiles of neurons extending from the hypothalamus and ventral midbrain that project to the nucleus accumbens. The results verified that Retro-TRAP works.
“We hope that Retro-TRAP will be broadly used and provide a more granular understanding of how complex neural circuits function and ultimately lead to better treatments for neurological and neuropsychiatric disorders,” Friedman says.
Additionally, their data contained some new findings. For instance, some neurons in the lateral hypothalamus express the p11 gene involved in depression. After investigation, they found these neurons also tended to express a protein called orexin, a regulator of sleep and feeding. This suggests a molecular association between depression and some of its symptoms.
“Molecular Profiling of Neurons Based on Connectivity.” Mats I. Ekstrand, Alexander R. Nectow, Zachary A. Knight, Kaamashri N. Latcha, Lisa E. Pomeranz and Jeffrey M. Friedman Cell Volume 157, Issue 5, p1230–1242, 22 May 2014