Optogenetics in 1000 words or less

Before the start of our department’s postgraduate research conference, I gave a 20 minute presentation on the methods of systems neuroscience. The idea was to try and provide a more general idea of what was involved in experimental neuroscience than could be squeezed into the beginning of the talk about my research. At the end of the presentation, I was asked which technique was my favourite. I’ve been trained in in vivo electrophysiology so that has become my weapon of choice; however, since first learning about optogenetics, I’ve fallen in love with the most sci-fi of neuroscience methods.

So, having missed the opportunity to wax lyrical about it then and inspired by Neuroskeptic’s fMRI in 1000 words, here is…

Optogenetics in 1000 words or less

In a 1979 issue of Scientific American, Francis Crick (of DNA fame) wrote a wish list of research methods to help understand how the brain processes information. One of those wishes was a method to gain control over some neurons whilst “leaving the others more or less unaltered.”

Traditionally, to investigate the functional connectivity of structures in the brain, a researcher would insert an electrode into an area of interest and pass an electric current through it to activate the neurons in the immediate area, and measure the results in another structure. However, this is not a particularly precise method – not only is the effective extent of the current difficult to predict exactly, but the current could potentially activate any and all neurons that are in the vicinity, as well as the connections of potentially irrelevant neurons that happen to pass close by.

However, perhaps most importantly, structures in the brain usually contain several different types of neuron acting together in a circuit, which may normally function in specific patterns; an increase in the activity of neurons in group A might reduce the activity of neurons in group B, or neurons in group X might only respond to activity in group Y or Z, but not both. In cases such as these, simultaneous activation of some or all neurons in all of the groups is not just unrealistic, but would potentially provide a misleading picture of their functional interaction.

Just under 35 years after Crick’s article, research groups began isolating proteins from light-sensitive algae. In 2005 Karl Diesseroth and colleagues used these developments, and ticked off Crick’s wish by demonstrating optogenetics in mammals. The technique involves inserting genes from light-sensitive organisms into neurons using a virus. The virus can be tailored to infect a particular subpopulation of neurons – for example one releasing a particular neurotransmitter. That’s the -genetics part.

The genes coded for light-sensitive proteins (‘opsins’) that were expressed in the cell membranes of the neurons. Most of the opsins used in optogenetics are ion channels; tiny gates in the membrane of the neuron that control the flow of electrically charged ions into and out of the neuron. The light gated ion channels are activated directly by a light flash from an implanted optic fibre. The light causes a conformational change in a light sensitive molecule, which in turn causes a conformational change in the tranmembrane proteins. This change opens the channel and allows the movement of ions. The flow of ions changes the electrical potential across the cell membrane which might, if sufficiently large, cause the neuron to fire. That’s the opto- part, and possibly the coolest thing to come out of neuroscience ever.

Diagram of the proteins of channelrhodopsin-2

Since Diesseroth’s experiment, a toolkit of techniques has been developed to deliver and turn on genes for whole a range of opsins of microbial, algal and other origins. The different opsins are sensitive to different wavelengths of light and provide different functions. For example Channelrhodopsin-2 (ChR2), sensitive to blue light (~480 nm), is a cation channel which depolarises the neuron. Halorhodopsin (NaHR), sensitive to yellow light, is a chloride specific ion pump which hyperpolarises it. Together with other excitatory (Volvox Channelrhodopsin – VChR2) and inhibitory (Archaerhodopsin – (Arch), and fungal opsins such as Leptosphaeria maculans opsin – (Mac)) opsins, subpopulations of neurons can be turned on and off with millisecond accuracy, enabling the frequency of their activity to be precisely controlled.

The effects of activating channelrhodopsin-2 and halorhodopsin (from h+ magazine)

The technique has been further developed by customising existing opsins to serve other functions, for example fusing opsins with G-protein coupled receptors to manipulate the concentration of intracellular signalling molecules such as cGMP and cAMP (Kim, 2005, Airan, 2009), or replacing one of the proteins in a channel with a fluorescent marker that can be used to locate the modified neurons. All of these techniques allow neural circuits to be precisely probed at the high speeds needed to understand neuronal communication, as well as investigating the fundamental assumptions behind proxy measures of neural activity such as fMRI.

As well as fundamental research, optogenetics also has potential for clinical treatments such as Parkinson’s disease. In one of the treatments for Parkinson’s disease, deep brain stimulation, electrical pacemakers are used to provide signals to structures which have lost their input due to the death of dopamine releasing neurons. The greater specificity of optogenetics has provided a greater understanding of why DBS works, and what can be done to improve its effectiveness. The technique is also providing insights into the neural mechanisms involved in autism, schizophrenia, drug abuse and depression.

Using optogenetics to manipulate reward

The seemingly endless advances in research technology amaze me. The thought that the electrochemical activity of cells in the body underlies an infinitely complex range of behaviour fascinates me. Optogenetics combines the two in the most impressive way. Since the development of the technique, it has been taken up in hundreds of laboratories, and entered the mainstream so far so fast that in 2006 Jay Leno made a joke about using optogenetics to control a fly to pester George W Bush.

Although the technique is still in its early stages (and there are suggestions that it might not be as perfectly selective as first assumed), it is still a huge leap forward for neuroscience research, and it promises to open up a whole new world of information about neural function.

Crick FH (1979) Thinking about the brain. Sci Am 241: 219–232

Early research

Zemelman BV, Lee GA, Ng M, Miesenböck G (January 2002). “Selective photostimulation of genetically chARGed neurons”. Neuron 33 (1): 15–22. PMID 11779476.

Banghart M, Borges K, Isacoff E, Trauner D, Kramer RH (December 2004). “Light-activated ion channels for remote control of neuronal firing”. Nat. Neurosci. 7 (12): 1381–6.doi:10.1038/nn135610.1038/nn1356. PMID 15558062

2005 optogenetics paper

Boyden ES, Zhang F, Bamberg E, Nagel G, Deisseroth K (September 2005). “Millisecond-timescale, genetically targeted optical control of neural activity”. Nat. Neurosci. 8 (9): 1263–8.doi:10.1038/nn152510.1038/nn1525. PMID 16116447

Optogenetics with customised opsins

Kim JM, Hwa J, Garriga P, Reeves PJ, RajBhandary UL, Khorana HG (February 2005). “Light-driven activation of beta 2-adrenergic receptor signaling by a chimeric rhodopsin containing the beta 2-adrenergic receptor cytoplasmic loops.”. Biochemistry 44 (7): 2284–92. doi:10.1021/bi048328i. PMID 15709741.

Airan RD, Thompson KR, Fenno LE, Bernstein H, Deisseroth K (April 2009). “Temporally precise in vivo control of intracellular signalling”. Nature 458 (7241): 1025–9.doi:10.1038/nature0792610.1038/nature07926. PMID 19295515

Cross-posted on Science Brainwaves Brain & Behaviour blog

About these ads