Guest post: Neuroscience through Optogenetics

A guest post from Hugh Pastoll, my good friend and long-time intellectual sparring partner. Hugh's introduction follows, and then his article. 

Michael and I met while studying PPE at the University of Cape Town. Like him, I’ve completely changed direction since then and am now doing a PhD in Computational Neuroscience at the University of Edinburgh.

As part of my postgraduate studies I’ve been fortunate enough to use an exciting new and truly revolutionary technology known as optogenetics. Optogenetics permits fine-grained control of brain activity with light, dramatically increasing the range of interesting experiments we can do. Since it is likely that it will soon become the technology of choice for investigating brain function, Michael has invited me to give a short primer on optogenetics in general and channelrhodopsins in particular.

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As a computational neuroscientist I am ultimately motivated by understanding how neural activity determines behavior. Frustratingly, for a long time even attempting to answer this sort of question has been pretty much impossible. This has been a major barrier to understanding how brains work… until recently.

To see why we have been stuck, imagine that I want to test the hypothesis that some pattern of neural activity causes a particular behavior. In order to test this hypothesis I’d need to conduct an experiment where I manipulated the animal’s neural activity and observed its behavior (simply noticing that the pattern and behavior both occur when I give the animal a stimulus only establishes correlation, not causality). Now, we’ve been able to control neural activity for quite a while - the sticking point was that we weren’t able to do it with the millisecond fidelity, neuron type specificity and sub-cubic-millimeter spatial precision we need to test most of our important hypotheses.

To illustrate, say I hypothesize that synchronized firing of excitatory neurons in the subthalamic nucleus at 20 Hz is responsible for akinesia (deficit in movement initiation). Testing this typical hypothesis would require me to synchronize only sub-thalamic excitatory neurons without changing their overall firing rate or affecting activity in nearby brain areas while the animal is behaving. I can’t think of any way we would have to able to accomplish this with drugs, electrical stimulation or any other standard technique for controlling neural activity.

Thanks to the recent development of optogenetics, though, such control is not only possible, but relatively easy. I can’t really exaggerate how completely cool this is - it is going to allow the field of computational neuroscience to hit its stride and start delivering the kinds of insights we need to understand what’s really going on in the brain.

So how does optogenetics work? To understand this, you need to know how ion channels control action potentials in neurons. Very briefly, ion channels are specialized protein channels that, when open, conduct ions (charged molecules) across cell membranes. The brief rise in membrane potential during an action potential is due to positive ions rapidly moving from the outside to the inside of a cell. Channelrhodopsins are ion channels that open when you shine blue light on them! This means we can force the membrane potential of a neuron to become more positive and generate an action potential. This is the ‘opto’ part of optogenetics.

Channelrhodopsin-2 (ChR2 - the most useful original kind) was first described in a species of green algae called Chlamydomonas reinhardtii in 2002 and found to work in mammalian neurons. Since then, genetic engineers have found that strategically mutating different amino acids changes the kinetics of the channel (how quickly it opens and closes). So, now there are different versions that allow different types of control. The fastest type (named ChETA ) opens in about 2 milliseconds and closes after about 5ms; fast enough to pulse blue light at 200 Hz and have the neuron fire at virtually every pulse. Another type (ChR2-C128S) usually takes minutes to close but shuts off very quickly if you shine green light on it. This means it can act as a kind of bi-stable on-off neuron switch. With such fine-grained control we can manipulate neuron spiking in pretty much any way we like.

Now for the ‘genetic’ part of optogenetics: Different kinds of neurons make different kinds of proteins. Since channelrhodopsin is a protein, we can use the cellular machinery that determines whether a protein is expressed in a neuron to restrict channelrhodopsin expression to a specific type of neuron.
This allows us to make one type of neuron in an area fire, without directly disrupting the normal activity of other types in the same area, giving us the neuron sub-type specificity we need for our experiments.

Furthermore, we can restrict channelrhodopsin expression to a very small area of the brain. Since we know that genes code for proteins, if only cells in one area have the channelrhodopsin gene only those cells in that area will respond to light. We can accomplish this by infecting a group of neurons with a non-replicating retrovirus that carries the channelrhodopsin gene. This gene will then be integrated into the genome of the infected neurons and expressed, introducing channelrhodopsins with spatial specificity.

However, although this combination of temporal, neuron sub-type and spatial specificity will enable a wide range of experiments, even more is possible. Another class of membrane proteins, known as halorhodopsins, have the opposite effect to channelrhodopsins. Halorhodopsins are not passive channels - they actively pump negative ions into a cell when illuminated with yellow light, making it more negative and stopping it from firing. Additionally, proteins that pump positive hydrogen ions out of cells to make their interior more negative have been described recently. These proteins are more effective than some types of halorhodopsins at preventing neurons from firing and different types respond to a different light colors – allowing researchers to pick colors that don't interfere with other rhodopsins the animal may also be expressing.

With such powerful optogenetic tools at our disposal we can imagine performing complex experiments, orchestrating neural activity with an array of different color intra-cranial LEDs. Although such experiments will be technically challenging, at the moment it feels like we are only limited by our imagination.

Selected references:
Nagel, G. et al. (2002) "Channelrhodopsin-2, a directly light-gated cation-selective membrane channel," PNAS, doi:10.1073/pnas.193619210.
Boyden, E. et al. (2005) "Millisecond-timescale genetically targeted optical control of neural activity," Nature Neuroscience, doi:10.1038/nn1525
Berndt, A. et al. (2008) "Bi-stable neural state switches," Nature Neuroscience, doi:10.1038/nn.2247
Gradinaru, V. et al. (2009) "Optical deconstruction of Parkinsonian neural circuitry," Science, doi:10.1126/science.1167093
Chow, B. et al. (2010) "High-performance genetically targetable optical neural silencing by light-driven proton pumps," Nature, doi:10.1038/nature08652
Gunaydin, L. et al. (2010) "Ultrafast optogenetic control," Nature Neuroscience, doi:10.1038/nn.2495

Video: Instantiated Turing machine

The Turing machine, first described in Alan Turing's classic paper "On Computable Numbers", is a seminal thought experiment that led directly to the machine you're currently using to read this. The Turing machine was never really meant to be built, but now some guy (not an academic, from what I can tell) has gone and built one, and it's capable of performing actual computations. Awesome.



Note: I've discussed Turing (once) before, and noted he had some daft ideas...

Interruption...

So I'm getting married at the end of the month. Not surprisingly, my wedding entails lots of travelling, organizing, stressing, honeymooning, etc. As a result, blogging until about April 5th will be sparse. Please don't break teh internets while I'm away...

Carnival of the Africans #14

Simon of Amanuensis has put up the 14th edition of the Carnival of the Africans, our humble effort to promote scientific and skeptical blogs from Africa. There are a bunch of very interesting posts to feast on, so do have a look! A selection: Mark from the Grumpy Old Man on the Lancet's retraction of the Wakefield paper and on a quack witchdoctor; Angela from The Skeptic Detective on the South African Society for Paranormal Research; and Simon himself on fMRI evidence for inequality aversion. My contributions to this edition were on deference and the importance of blinding...

We do need hosts for future editions of the carnival, so if you'd like to take on the responsibility (it's fun...), have a look at the schedule, and drop me an email!

African science/skepticism blogrolling for February

The updated African science and skepticism blog roll for February... If you know of blogs not listed here, please let me know. Also: add it to your blog! Do a post like this one! (Email me, and I'll send you the HTML).

• 01 and the universe
• Acinonyx Scepticus
• Amanuensis
  
• ASSAf Blog
• Botswana Skeptic 
• Bomoko and other nonsense words
  
• Communicating Science, the African Way
  
• Defollyant's AntiBlog
  
• Effortless Incitement
• Ewan’s Corner
• Expensive Beliefs **new**
• Geekery
• Grumpy Old Man
• Health Frog **new** 
• Hello Universe, This is Nessie
  
• Ionian Enchantment
• Limbic Nutrition
• Lenny Says 
• McBrolloks
• Meh Blog **new** 
  
• Nathan Bond's TART Remarks
  
• Orion Spur
  
• Other Things Amanzi
• Pickled Bushman
• Psychohistorian
  
• Reason Check
  
• Retroid Raving
• Roy Jobson **new**
• Scorched
• Shadows Hide
• Skeptic South Africa **new**
• Stop Danie Krügel
• Sumbandlila Mission Blog **new**
  
• Synapses
• Tauriq Moosa
• The Joys of Atheism **new**
  
• The Science Of Sport
• The Skeptic Black Sheep
• The Skeptic Detective
• Word of the Blog
  

Picture: Why global warming can't be a conspiracy...

A nice graphic, from Going on a Bear Hunt.

(via PZ Myers).

Encephalon #80: The Twitter Edition

Welcome to the 80th edition of Encephalon (@encephalon_), the world's best mindy/brainy/behaviory blag carnival! Since I've finally joined the whole Twitter party properly (@michaelmeadon), I figured making this the Twitter Edition would be fun. It also features an entirely gratuitous picture of a hot bird (haha), right. So, here are some 'Tweets', all < 140 characters... (The @xxx's refer to the relevant person's Twitter account, if there is one, and the link at the end of each 'Tweet' goes to the blog entry).

• @edyong209 asks: can a sniff of oxytocin improve the social skills of autistic people? Apparently yes: http://bit.ly/dqVSIV
• @neurocritic on on alien hand syndrome and agency error http://bit.ly/d8D3kY & on neuropsychoanalysis: http://bit.ly/bU3LyD
• @vaughanbell wonders whether you really can be frightened to death: http://bit.ly/d0ehYL
• @podblack on facilitated communication and Belgian Rom Houben. Newsflash: it didn't really work: http://bit.ly/aTHvWy
• @sharpbrains head honcho @AlvaroF interviews Michael Merzenich on brain training, etc. http://bit.ly/5kfG7w
• @sharpbrains says working memory is a better predictor of academic success than IQ: http://bit.ly/5Q3ZJ5
• @somatosphere has a summary of an interdisciplinary UCLA conference on the Cultural and Biological Contexts of Psychiatric Disorder: http://bit.ly/d1J1tT
• @researchdigest says swanky cars enhance men's appeal to women, but not women's  to men: http://bit.ly/dr9krV
• @BrainBlogger on a BMJ article that concluded children with a high IQ have a reduced risk of mortality as adults: http://bit.ly/9T4Hhy
• @BrainBlogger argues the act of speaking in tongues as a verifiable language phenomenon: http://bit.ly/bOlKX2
• @jonahlehrer reveals sportspeople don't maximize gains like game theory predicts: http://bit.ly/bw769F. Von Neumann pissed.

That's it! There doesn't seem to be a host for the next Encephalon yet, so volunteer! 

The Cost of Truth is Eternal Vigilance

A recurring theme on this blog is that it is unwise to rely on 'everyday' or uncritical thinking because our minds are liable to innumerable biases, failures of memory, and so on. An important part of being a good thinker, then, is to submit ideas - and especially our own - to critical scrutiny. I am not, obviously, immune to these biases, in fact, I am as liable to them as anyone else. I do work hard to scrutinize my beliefs carefully, though, and I regularly give up previously held beliefs as a result. To demonstrate not only the dangers of uncritical thinking, but also that I (try to) practice what I preach, here are two recent instances of having to change my mind. Both are pretty unimportant beliefs, but they illustrate the issues nonetheless.

I moved from Johannesburg to Durban in early 2007 and my fiancée did the same in early 2009. Possibly as a result of her comments about how much it has been raining in Durban, I came to believe that 2009 had been an especially wet year: I thought it must be the wettest since I'd moved here. I knew, of course, that the only way to establish this for sure was to look at actual statistics because our memories are flawed and we use the availability heuristic to make inferences about trends. But... I didn't bother to check for a while. When I finally did, it became quite clear that my intuitive sense about Durban's weather was spectacularly wrong. The wonder that is Wolfram Alpha let me create the following two graphs: the first shows the total estimated yearly precipitation (rain, for Durban's purposes) for the last 5 years, and the second shows (I think weekly) rainfall amounts over the same period.

As should be abundantly clear, 2009 is not the wettest year since I moved to Durban, it is in fact the driest. Now, it could be the case that 2009 had less total rainfall, but more rainy days, so I could have been misled for that reason. The second graph, though, is only mildly suggestive on that front and I can find no other data (that's free). So it seems fair to conclude that I was led astray by thinking intuitively when I should have known not to trust my intuitions about trends in complex, variable systems. (For detailed evidence that people are spectacularly bad at thinking statistically, see Kahneman, Slovic & Tversky, 1982).

The second example concerns bias and rather nicely illustrates the importance of blinding. If you had asked me a while ago what the best search engine was, I would have said: "Google - and by a wide margin". Until I found BlindSearch, that is. Branding biases our judgments and Google's brand is so powerful that being objective while knowing which search engine's results you're looking at is extremely difficult. BlindSearch remedies this problem: it lets you search Bing, Yahoo and Google simultaneously, presents the results in three columns, and blinds you to which search engine produced which results. You look through the results, vote for the one you prefer, and then only are the brand names revealed. I've now used BlindSearch dozens of times and a clear pattern has emerged: Google isn't nearly as superior as I once thought it was. While I still tend to prefer Google's results a plurality of the time, Bing and Yahoo do get my vote more often than I would have thought. For the sake of concreteness, here are ten searches I did with my vote listed next to it. I tried to pick topics that were either obscure or controversial to 'test' the search engines, since search terms with obvious results aren't exactly indicative of quality. Also, I verified some of these results by checking whether my vote stayed the same later (it did in all cases).

• Science - Bing
• Islamic terrorism - Difficult call, but Yahoo
• Evolution - Yahoo
• iPad - Google (though it was close)
• Michael Meadon - Google
• Sex - Yahoo (though Bing doesn't work for some reason)
• Homeopathy - Google (by a large margin)
• Jacob Zuma - Bing (close)
• Richard Dawkins - Google (Yahoo sucked)
• Sitcky the stick insect - Google, by far (also wins on Olaf the Hairy)

So that's 2 for Bing, 5 for Google and 3 for Yahoo. Without blinding, my guess would have been that I would have preferred Google 9 times out of 10. Turns out I was wrong. And, contrary to what I'd like to believe, branding works on me too. Bottom line: our biases affect our decisions and our judgments, so when those decisions or judgments are important (which is not the case with search engines), appropriate blinding is vital.

These are just two, small, inconsequential examples, of course. They illustrate an important point though: if you want to be right, you have be be skeptical, self-critical, willing to reconsider and admit error, cautious, and scrupulously careful with facts and arguments. Or, to corrupt a glorious quote misattributed to Thomas Jefferson: the cost of truth is eternal vigilance.