Optogenetics

Optogenetics is the integration of optics and genetics to control precisely defined events within specific cells of living tissue even within freely moving animals, with the temporal precision (millisecond-timescale) needed to keep pace with functioning intact biological systems, and with suitably fast and precise methods for reading out the effects of optogenetic control.

In 2012 Gero Miesenböck was awarded the |336 InBev-Baillet Latour International Health Prize for "pioneering optogenetic approaches to manipulate neuronal activity and to control animal behaviour."

In 2010 Karl Deisseroth was awarded the inaugural HFSP Nakasone Award for "for his pioneering work on the development of optogenetic methods for studying the function of neuronal networks underlying behavior."

In 2010, optogenetics was chosen as the Method of the Year across all fields of science and engineering by the interdisciplinary research journal Nature Methods (Primer on Optogenetics, Editorial Commentary ). At the same time, optogenetics was highlighted in the article on “Breakthroughs of the Decade” in the scientific research journal Science Breakthrough of the Decade; these journals also referenced recent public-access general-interest video Method of the year video and textual SciAm summaries of optogenetics.

History
The "far-fetched" possibility of using light for selectively controlling precise neural activity (action potential) patterns within subtypes of cells in the brain was articulated by Francis Crick in his Kuffler Lectures at the University of California in San Diego in 1999. An early use of light to activate neurons was carried out by Richard Fork and later Rafael Yuste, who demonstrated laser activation of neurons within intact tissue, although not in a genetically-targeted manner. The earliest genetically targeted method, which used light to control genetically-sensitised neurons, was reported in January 2002 by Boris Zemelman and Gero Miesenböck, who employed Drosophila rhodopsin photoreceptors for controlling neural activity in cultured mammalian neurons. In April 2005, Susana Lima and Miesenböck reported the first use of genetically-targeted photostimulation to control the behaviour of an animal. They showed that photostimulation of genetically circumscribed groups of neurons, such as those of the dopaminergic system, elicited characteristic behavioural changes in fruit flies. Beginning in 2004, the Kramer and Isacoff groups developed organic photoswitches or "caged" compounds that could interact with genetically-introduced ion channels.

In August 2005, Karl Deisseroth's laboratory in the Bioengineering Department at Stanford including graduate students Ed Boyden and Feng Zhang (both now at MIT) published the first demonstration of a single-component optogenetic system, beginning in cultured mammalian neurons. using channelrhodopsin, a single-component light-activated cation channel from unicellular algae), whose molecular identity had been confimed in November 2003. Channelrhodopsin (ChR)  allowed millisecond-scale temporal control in mammals, required only one gene to be expressed in order to work, and responded to visible-spectrum light with a chromophore retinal that was already present and supplied to the ChR by the vertebrate tissues. The surprising experimental utility of this single-component "microbial opsin" approach was quickly proven with many additional microbial opsin classes and in a variety of animal models ranging from behaving mammals to classical model organisms such as flies, worms, and zebrafish. The "optogenetic" terminology was coined in 2006, and since 2005 hundreds of laboratories around the world have employed microbial opsins to study complex biological systems (references below).

Description
Millisecond-scale temporal precision is central to optogenetics, which allows the experimenter to keep pace with fast biological information processing (for example, in probing the causal role of specific action potential patterns in defined neurons). Indeed, to probe the neural code, optogenetics by definition must operate on the millisecond timescale to allow addition or deletion of precise activity patterns within specific cells in the brains of intact animals, including mammals (see Figure 1). By comparison, the temporal precision of traditional genetic manipulations (employed to probe the causal role of specific genes within cells, via "loss-of-function" or "gain of function" changes in these genes) is rather slow, from hours or days to months. It is important to also have fast readouts in optogenetics that can keep pace with the optical control. This can be done with electrical recordings ("optrodes") or with reporter proteins that are biosensors, where scientists have fused fluorescent proteins to detector proteins. An example of this is voltage-sensitive fluorescent protein (VSFP2). The hallmark of optogenetics therefore is introduction of fast light-activated channels and enzymes that allow temporally precise manipulation of electrical and biochemical events while maintaining cell-type resolution through the use of specific targeting mechanisms. Among the microbial opsins which can be used to investigate the function of neural systems are the channelrhodopsins (ChR2, ChR1, VChR1, and SFOs) to excite neurons. For silencing, halorhodopsin (NpHR), enhanced halorhodopsins (eNpHR2.0 and eNpHR3.0), archaerhodopsin (Arch), Leptosphaeria maculans fungal opsins (Mac), and enhanced bacteriorhodopsin (eBR) have been employed to inhibit neurons (see Figure 2), including in freely-moving mammals.

Moreover, optogenetic control of well-defined biochemical events within behaving mammals is now also possible. Building on prior work fusing vertebrate opsins to specific G-protein coupled receptors a family of chimeric single-component optogenetic tools was created that allowed researchers to manipulate within behaving mammals the concentration of defined intracellular messengers such as cAMP and IP3 in targeted cells Other biochemical approaches to optogenetics (crucially, with tools that displayed low activity in the dark) followed soon thereafter, when optical control over small GTPases and adenylyl cyclases was achieved in cultured cells using novel strategies from several different laboratories. This emerging repertoire of optogenetic probes now allows cell-type-specific and temporally precise control of multiple axes of cellular function within intact animals.

Optogenetics also necessarily includes 1) the development of genetic targeting strategies such as cell-specific promoters or other customized conditionally-active viruses, to deliver the light-sensitive probes to specific populations of neurons in the brain of living animals (e.g. worms, fruit flies, mice, rats, and monkeys), and 2) hardware (e.g. integrated fiberoptic and solid-state light sources) to allow specific cell types, even deep within the brain, to be controlled in freely behaving animals. Most commonly, the latter is now achieved using the fiberoptic-coupled diode technology introduced in 2007  To stimulate superficial brain areas such as the cerebral cortex, optical fibers or LEDs can be directly mounted to the skull of the animal. More deeply implanted optical fibers have been used to deliver light to deeper brain areas. In invertebrates such as worms and fruit flies some amount of Retinal isomerase all-trans-retinal (ATR) is supplemented with food. A key advantage of microbial opsins as noted above is that they are fully functional without the addition of exogenous co-factors in vertebrates.

The field of optogenetics has furthered the fundamental scientific understanding of how specific cell types contribute to the function of biological tissues such as neural circuits in vivo (see references from the scientific literature below). Moreover, on the clinical side, optogenetics-driven research has led to insights into Parkinson's disease and other neurological and psychiatric disorders. Indeed, optogenetics papers in 2009 have also provided insight into neural codes relevant to autism, Schizophrenia, drug abuse, anxiety, and depression.

It has been pointed out that beyond its scientific impact, optogenetics also represents an important case study in the value of both ecological conservation (as many of the key tools of optogenetics arise from microbial organisms occupying specialized environmental niches), and in the importance of pure basic science (as these opsins were studied over decades for their own sake by biophysicists and microbiologists, without involving consideration of their potential value in delivering insights into neuroscience and neuropsychiatric disease).

Applications
"In vivo" optogenetic activation and/or silencing has been recorded in the following brain regions and cell-types. In vivo and in vitro recordings (by the Cooper laboratory) of individual CAMKII AAV-ChR2 expressing pyramidal neurons within the prefrontal cortex demonstrated high fidelity action potential output with short pulses of blue light at 20 Hz (Figure 1). The same group recorded complete green light-induced silencing of spontaneous activity in the same prefrontal cortical neuronal population expressing an AAV-NpHR vector (Figure 2). The Deisseroth laboratory integrated optogenetics, freely moving mammalian behavior, "in vivo" electrophysiology, and slice physiology to probe the cholinergic interneurons of the nucleus accumbens by direct excitation or inhibition. Despite representing less than 1% of local neurons, these cholinergic cells have dominant control roles, exerting powerful modulation of circuit activity. Furthermore, these neurons are activated by cocaine, and silencing this drug-induced activity during cocaine exposure blocked cocaine conditioning suggesting an important role for these interneurons in drug reward mechanisms.
 * Prefrontal Cortex
 * Nucleus Accumbens
 * Cholinergic Interneurons