NIMH · Light Switches Brain Pathway On-and-Off to Dissect How Anxiety Works

Science Update • April 18, 2011

Light Switches Brain Pathway On-and-Off to Dissect How Anxiety Works
Turns Cowering Mice into Instant Adventurers

Scientists, for the first time, have switched anxiety on-and-off in active animals by shining light at a brain pathway. Instinctively reclusive mice suddenly began exploring normally forbidding open spaces when a blue laser activated the pathway – and retreated into a protected area when it dimmed. By contrast, anxiety-like behaviors increased when an amber laser inhibited the same pathway.

Researchers, supported in part by NIMH, used a virus, genetic engineering and fiber-optics to control the pathway in the brain's fear center with millisecond precision.

"Our findings reveal how balanced antagonistic brain pathways are continuously regulating anxiety," explained Karl Deisseroth, M.D., Ph.D., of Stanford University, a practicing psychiatrist as well as a neuroscientist. "We have pinpointed an anxiety-quelling pathway and demonstrated a way to control it that may hold promise for new types of anti-anxiety treatments."

NIMH grantees Deisseroth, Kay M. Tye, Ph.D., and colleagues, report on their findings March 17, 2011 in the journal Nature.

Optogenetic alchemy
Anxiety disorders are the most common type of psychiatric illness, affecting more than 1 in 4 people at some time during their lives. To understand the neural basis of these disorders, researchers are studying the workings of circuitry in the fear center, called the amygdala, in rodents.

Deisseroth's team has pioneered a method, called optogenetics, of experimentally activating brain activity with light. They incorporate a protein borrowed from light-reactive organisms to make brain tissue similarly light-responsive. Previously, they used this tool to activate particular types of neurons. The new study is the first to use it to reversibly manipulate a specific projection of a neuron (see picture below). It's also the first time the technique has been used to study anxiety as opposed to fear – a generalized state versus a transient reaction to an immediate threat.

The researchers borrowed a gene that codes for a light-sensitive protein from algae and delivered it to the amygdala pathway via a virus. In the algae, the protein's function is to activate a pathway that causes the organism to swim toward blue spectrum light. Hence a blue light now activated the amygdala pathway. When they wanted to inhibit the pathway in response to light, they similarly borrowed a gene from a light-responsive bacterium that codes for a protein that inhibits a pathway in response to a particular spectrum of light — in this case amber — and infected the amygdala pathway with that gene.

When the researchers optogenetically activated whole neuronal cell bodies in the amygdala, it increased anxiety-like behavior: mice hunkered down in a protected corner of a maze and wouldn't venture into more exposed areas. These and related findings led the researchers to hypothesize that they would get the same effect if they narrowed the focus of the activation to just a specific neuronal projection (see picture below).

A post-doc's eureka! moment
But it turned out that the opposite was true.

When they activated the projection with the blue laser, the engineered mice suddenly seemed to summon the courage to explore the more exposed parts of the maze that they would normally avoid (see video below).

"I was quite surprised. We did not see aversion. We did not see fear. We did not see any of these things I expected to see," said Tye, whose post-doctoral study is supported by a NIMH-funded training grant. "I suddenly got this huge, dramatic effect of reduction in anxiety-related behaviors and I had to follow it up. So I pretty much dropped my original ideas of what I was going to study during my fellowship and started pursuing this."

When the researchers blocked activity in the projection with the amber laser, the animals showed even more anxiety-like behavior than they usually do. The experiments hint at how the brain is able to regulate anxiety levels — on a millisecond timescale — by dialing activity up and down in such antagonistic amygdala pathways.

Futuristic anxiety treatment?

Tye said she and Deisseroth plan to follow up with further dissection of anxiety pathways. She also hopes to examine whether such optogenetic manipulations, sustained over hours or days, might induce long-lasting adaptations — perhaps for weeks –– in the set-points of anxiety pathways.

A future anxiety disorder treatment that might similarly target such specific pathways could, theoretically, quell anxiety instantly without producing unwanted side effects, such as drowsiness, often experienced with current anti-anxiety medications. For patients with severely debilitating anxiety, a treatment something like deep brain stimulation for depression, but more precisely targeted at a specific pathway, might someday be feasible, she suggested.

"Everything else in your brain should be unperturbed, because the manipulation would be so specific," explained Tye.

Video (see the original text please) shows a mouse under "optogenetic" control while in an anxiety-producing situation. Being in elevated, open spaces makes mice anxious. So, in this "elevated-plus maze," the mouse normally stays in the arms with high walls; it normally won't venture into arms with low walls. However, this mouse has been genetically engineered to have an anxiety-quelling pathway in its fear hub activate when a blue laser shines on it via the fiber-optic cable. At those times (when the blue text appears), the animal gains courage and ventures into the normally scary places. Video speeds up a 15 minute session 10-fold.

Source, Kay M. Tye, Ph.D., Stanford University

Researchers were surprised to discover that activating the whole cell body of an amygdala neuron increased anxiety in mice, while activating just one of its projections had the opposite effect. So unraveling the secrets of how anxiety works might require dissecting the action of each such pathway individually, say the researchers.

Reference
Amygdala circuitry mediating reversible and bidirectional control of anxiety. Tye KM, Prakash R, Kim SY, Fenno LE, Grosenick L, Zarabi H, Thompson KR, Gradinaru V, Ramakrishnan C, Deisseroth K. Nature. 2011 Mar 17;471(7338):358-62. Epub 2011 Mar 9. PMID: 21389985



full-text:
NIMH · Light Switches Brain Pathway On-and-Off to Dissect How Anxiety Works


Nature. 2011 Mar 17;471(7338):358-62. Epub 2011 Mar 9.
Amygdala circuitry mediating reversible and bidirectional control of anxiety.
Tye KM, Prakash R, Kim SY, Fenno LE, Grosenick L, Zarabi H, Thompson KR, Gradinaru V, Ramakrishnan C, Deisseroth K.
SourceDepartment of Bioengineering, Stanford University, Stanford, California 94305, USA.

Abstract
Anxiety--a sustained state of heightened apprehension in the absence of immediate threat--becomes severely debilitating in disease states. Anxiety disorders represent the most common of psychiatric diseases (28% lifetime prevalence) and contribute to the aetiology of major depression and substance abuse. Although it has been proposed that the amygdala, a brain region important for emotional processing, has a role in anxiety, the neural mechanisms that control anxiety remain unclear. Here we explore the neural circuits underlying anxiety-related behaviours by using optogenetics with two-photon microscopy, anxiety assays in freely moving mice, and electrophysiology. With the capability of optogenetics to control not only cell types but also specific connections between cells, we observed that temporally precise optogenetic stimulation of basolateral amygdala (BLA) terminals in the central nucleus of the amygdala (CeA)--achieved by viral transduction of the BLA with a codon-optimized channelrhodopsin followed by restricted illumination in the downstream CeA--exerted an acute, reversible anxiolytic effect. Conversely, selective optogenetic inhibition of the same projection with a third-generation halorhodopsin (eNpHR3.0) increased anxiety-related behaviours. Importantly, these effects were not observed with direct optogenetic control of BLA somata, possibly owing to recruitment of antagonistic downstream structures. Together, these results implicate specific BLA-CeA projections as critical circuit elements for acute anxiety control in the mammalian brain, and demonstrate the importance of optogenetically targeting defined projections, beyond simply targeting cell types, in the study of circuit function relevant to neuropsychiatric disease.

PMID:21389985[PubMed - indexed for MEDLINE]
Amygdala circuitry mediating reversible and bidire... [Nature. 2011] - PubMed result