optogenetics

Scared by the light

neurophilosophy — Fri, 02 Jul 2010 14:25:47 +0000

Scared by the light

WHO could have guessed that a protein isolated from pond scum would transform the way researchers investigate the brain? The protein, called channelrhodopsin (ChR), is found in algae and other microbes, and is related to the molecule in human photoreceptors that captures light particles. Both versions control the electrical currents that constantly flow in and out of cells; one regulates the algae's movements in response to light, the other generates the nervous impulses sent along the optic nerve to the brain. Unlike its human equivalent, the algal ChR controls the currents directly because it forms a pore that spans the cell membrane. When expressed in neurons, it renders the cells sensitive to light, and they can be switched on or off very precisely using lasers.

This discovery led to the emergence of a new field called optogenetics. Early studies showed that the technique can be used to control the behaviour of small organisms such as nematode worms and fruit flies. Last year, Karl Deisseroth's group at Stanford University demonstrated, for the first time, that it can also be used to control reward and motivation behaviours in mice. Josh Johansen of the Center for Neural Science at New York University and his colleagues have now taken this one step further. Working in collaboration with Deisseroth, they show that optogenetics can also be used to induce a simple form of associative learning called fear conditioning.

Fear conditioning typically involves associating an innocuous stimulus such as an audible tone with an aversive stimulus such as electric shock. With repeated pairings, the animal quickly learns an association between the two, and subsequently expects to receive a shock whenever it hears the tone. As a result, it will exhibit a fear response (freezing) when the innocuous stimulus is presented alone. The initial association between the two stimuli is thought to be due to synaptic plasticity in the amygdala - the inputs related to each stimulus converge on the same group cells, causing the connections between them to be strenthened. As a result, a memory associating the innocuous stimulus with the aversive one is encoded, but this quickly fades, or becomes extinct, if the innocuous stimulus is then repeatedly presented alone.

Johansen and his colleagues substituted the electric shock with pulses of laser light, delivered to the lateral nucleus of the amygdala through a fibre optic cable. This was done in three groups of rats - in one, the light pulses were delivered at the same time as a tone; in another, just before; and in the third, just after. 24 hours later, they played the tone back, and tested the animals' responses to it. The first group of rats froze when they heard it, biut the other two did not, showing that the light pulses were sufficient to induce fear conditioning, but only when delivered at the same time as the tone. This supports the view that associative learning occurs when inputs representing the two stimuli coincide on the same neurons.

The observed fear responses were, however, significantly smaller than those seen when electric shocks are used, or when the central nucleus of the amygdala is electrically stimulated. This might be because the optical stimulation activated cells in the lateral nucleus indiscriminately, interfering with proper encoding of the memory. A more likely explanation is that the lateral amygdala alone is insufficient to generate a full fear response, and that other components of the brain's fear circuitry are also required. One of these other components is the central nucleus of the amygdala, whose role in fear conditioning overlaps with, but is distinct from, that of the lateral mucleus.

"The central nucleus is especially involved in controlling responses," says senior author Joseph LeDoux, "while the lateral nucleus is especially involved in sensory processing and association [between the two stimuli], and is the initial site of plasticity. The central nucleus is also a site of plasticity, but seems to be downstream, which is why we focused on the lateral nucleus." LeDoux's group is now introducing ChRs into different cell types in the amygdala, in order to examine the contribution of each to fear conditioning. They are also investigating the effects of various neuromodulators on synaptic plasticity in the amygdala.

Optogenetics is still in its infancy, and will undoubtedly become more advanced with time, enabling researchers to probe the cellular mechanisms of fear conditioning in greater detail. Indeed, a group from the Howard Hughes Medical Institute has just reported using the technique in combination with two-photon microscopy, not only to target single cells in the hippocampus, but also to selectively activate distinct subcellular compartments such as the axon and dendrites. Such advances will eventually provide a more detailed understanding of how the amygdaloid nuclei are connected to one another and to other parts of the fear circuit, as well as how information related to fear conditioning is processed by individual cells and neural networks.

Related:

Johansen, J., et al. (2010). Optical activation of lateral amygdala pyramidal cells instructs associative fear learning. Proc. Nat. Acad. Sci. DOI: 10.1073/pnas.1002418107.

neurophilosophy Fri, 07/02/2010 - 10:25

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neuroscience

optogenetics

amygdala

brain

classical conditioning

fear

neuroscience

optogenetics

Thanks for the post. Excellent job of describing the study. We were really pleased with the way this project turned out. There have been a number of interesting optogenetic studies of plasticity, but relatively few that have related such plasticity to actual behavioral learning. Fear conditioning is a good model for doing this kind of work because of the excellent control the experimenter has over the relevant stimuli. And because we've learned so much about the role of the lateral amygdala in fear conditioning, this was an ideal brain area to test usefulness of this approach. Now we can get on with the really interesting questions using this approach.

a free access description of a LeDoux experiment and the first comment is by the actual Joseph LeDoux. hmmm....I think the internet has become unbeatable.

I'm a lay person "fan" of neuroscience through experience of brain tumor, under-utilized high intelligence, etc. etc. I'm the opposite of your mice, and seek the light, even when my reward is pain. Maybe that's why I'm an artist (opera singer).

Wonderful article. I've been reading your work in the last weeks and I really appreciate your effort. LeDoux writing here is just a plus! Keep on the good stuff. ;)

Three notable scientists (Peter Hegemann, Georg Nagel, and Ernst Bamberg) actually won The Wiley Prize in Biomedical Sciences for their work on channelrhodopsins.

I am a long time member of the "Neuroscience" fan club. When I see a simple way to use light as amygdala food I do it. I have set up a reading station by installing a 120 watt grow lamp above and behind each shoulder. By treating myself as a plant I stay warm and avoid S.A.D. It also keeps my eyes happy.

Lights, genes, action

ddobbs — Wed, 26 May 2010 11:27:37 +0000

Lights, genes, action

Two or three years ago, Emory neurologistÂ Helen Mayberg, whose experiments using deep-brain stimulation for depression I check in on now and then,Â told me that Karl Deisseroth's work using light to fiddle with brain circuits had huge potential both as a replacement for DBS and for much else. As Lizzie Buchen ably reports in Nature, that potential is now being realized.

This is a very slick tool that seems almost too far out to actually work. It lets you use light to turn brain circuits on and off at will, and with great precision. It's not simple to construct. But once constructed, it works simply. And because it can isolate tightly defined circuits and turn them on and off, it has huge potential both as a research tool (Hm; what doesÂ this circuit do?) and as what amounts to a reversible ablation tool. DBS is reversible â a huge asset if things go awry, and a great way to test for placebo effect â but you can't target it as precisely as you can optogenetics and it inflicts a greater intrusion into skull and brain.ï»¿

Using a hybrid of genetics, virology and optics, the techniques involved enable researchers to instantaneously activate or silence specific groups of neurons within circuits with a precision that electrophysiology and other standard methods do not allow. Systems neuroscientists have longed for such an advance, which allows them their first real opportunity to pick apart the labyrinthine jumble of cell types in a circuit and test what each one does. "It has revolutionized my approach to science," says Antonello Bonci, a neurophysiologist at the UCSF Ernest Gallo Clinic and Research Center in Emeryville who began using the technique in 2007. "It can clarify unequivocally the role of specific classes of cells, and solve controversies that have been going on for many, many years." Among the clarifications sought is the precise function of 'place' cells, hippocampal neurons that fire only when an animal finds itself in a specific location; another is the function of complex activity patterns observed when an animal is paying attention or executing a movement.ï»¿

"God's gift to neurophysiologists," MIT's Robert Desimone calls it. You can get the rest at Nature, where they've recently made all news items, and news features like this one, open access.

ddobbs Wed, 05/26/2010 - 07:27

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Brains and minds

Genetics & genomics (incl behav genetics)

medicine

deep brain stimulation

Optogenetic fMRI

neurophilosophy — Fri, 21 May 2010 01:50:54 +0000

Optogenetic fMRI

OF all the techniques used by neuroscientists, none has captured the imagination of the general public more than functional magnetic resonance imaging (fMRI). The technique, which is also referred to as functional neuroimaging and, more commonly, "brain scanning", enables us to peer into the human brain non-invasively, to observe its workings and correlate specific thought processes or stimuli to activity in particular regions. fMRI data affect the way in which people perceive scientific results: colourful images of the brain have persuasive power, making the accompanying data seem more credible.

Functional neuroimaging is used widely by researchers, too, with tens of thousands of research papers describing fMRI studies being published in the past decade. Yet, a big question mark has been hanging over the validity of the technique for over a year and, furthermore, the way in which fMRI data are interpreted has also been called into question. Using a novel combination of fMRI and a recently developed state-of-the-art technique called optogenetics, researchers now provide the first direct evidence that the fMRI signal is a valid measure of brain activity.

fMRI measures brain activity indirectly, using signals generated by the flow of blood around the brain. It is based on the assumption that increased blood flow to a particular region of the brain is related to activity in that region, because the cells within it require a supply of oxygen to generate nervous impulses. But several recent studies challenged this assumption. The most important of these, by Yevgeniy Sirotin and Aniruddha Das of Columbia University, showed that the brain pre-empts itself by increasing the flow of blood to regions that might become active in the near future, but that this anticipatory blood flow is not always needed. This prompted the question of whether or not fMRI actually measures what researchers have always claimed it measures.

The new study, published online in the journal Nature, finally answers this question, by confirming that the fMRI signal is indeed closely correlated with increased brain activity. Senior author Karl Deisseroth published one of the first papers describing the optogenetics technique, back in 2005. That paper showed that light-sensitive proteins called channelrhodopsins, which had been isolated from algae several years earlier, can make the neurons sensitive to light when shuttled into them. As a result, light pulses of the appropriate can be used to control the activity of individual, specified cells, on a millisecond timescale. The technique was quickly applied to controlling simple behaviours in small organisms such as nematode worms and fruit flies; more recently, it has been used to successfully control complex functions, such as reward behaviors and memory, in mammals; it has even been used to restore vision in blind mice.

This time, Deisseroth and his colleagues injected a virus carrying the channelrhodopsin gene into the primary motor cortex of mice. The gene was engineered so that it would be expressed specifically in the large pyramidal cells that send axons down into the spinal cord and control movement. The mice were then placed into a custom-made MRI-compatible cradle fitted with a stereotaxic frame to keep their heads still. With the animals inside the scanner, the researchers delivered light pulses to the virus injection site, using an optical fibre inserted through a hole in the skull. In this way, they could activate specified subsets of cells in the motor cortex and simultaneously monitor the fMRI signals generated by them.

The researchers observed fMRI signals (above right) in the motor cortex 3-6 seconds after delivering pulses of light to the motor cortex. The signals originated in the area into which the light pulses were delivered (indicated above by the asterisk) and spread away from it, lasting for about 20 seconds before returning to a baseline level. By contrast, no fMRI response was seen in control mice injected with a salt water solution instead of the ChR2 gene (left). Activity in a part of the brain called the thalamus, which receives connections from the motor cortex, was also observed. This downstream activity was initially weaker than that seen in the cortex, but after a 5-second delay showed a very similar pattern, and reliably followed the cortical activity by several hundredths of a second.

The activity pattern observed in the thalamus again demonstrates that fMRI can accurately measure the activity of groups of neurons. It also shows that the combination of optogenetics and fMRI can be used to investigate how activity in one region of the brain alters activity in distant regions via long-range connections - the 5-second delay is consistent with network activity that modulates the output of the cortex and activity in the thalamus. By introducing ChR2 into the thalamus and performing simultaneous optical stimulation and neuroimaging, the researchers also revealed hitherto unknown details about the pathways connecting the thalamus and motor cortex.

The thalamus is thought of as a 'relay station', which receives sensory information en route to the cortex, and provides the cortex with feedback. The feedback pathways are mostly ipsilateral, that is, they project to the sensory cortical areas on the same side of the brain. The thalamus also sends and receives information to and from the motor cortex, but the former pathways are thought to project to motor areas on both sides of the brain, because control movement involves co-ordination between the two sides of the body. The experiments confirmed this, by showing that optical stimulation of one side of the thalamus evoked activity in the motor cortical areas on both sides.

Previous attempts to investigate the relationship between blood flow and neuronal activity have involved using microelectrodes to stimulate small groups of cells whilst simultaneously performing functional neuroimaging. This method has limitations, because electrical stimulation activates not only those cells targeted by the electrodes, but also distant cells whose axons traverse the stimulated brain region. In these neurons, the electrodes elicit nervous impulses that travel backwards along the axon, causing the cell body to generate further impulses. The resulting activity can therefore potentially confound the fMRI signal, because it is unrelated to that of the targeted cells.

Optogenetic fMRI largely overcomes this problem. Optical stimulation will activate ChR2-expressing axons in the targeted area, and the resultant nervous impulses will be back-propagated. So although some unwanted activity might be observed, it will be related to the cells of interest, because only those cells will be sensitive to light. As well as confirming the validity of fMRI data, this initial description of optogenetic fMRI shows that it is a very powerful technique for investigating neuronal activity at both the local and the global level. The new study does not, however, rule out the possibility that fMRI also detects other unrelated signals. Many questions still remain about exactly how the fMRI signal is generated but, using optogenetics, researchers may soon begin to answer them.

Related:

Lee, J., et al (2010). Global and local fMRI signals driven by neurons defined optogenetically by type and wiring. Nature DOI: 10.1038/nature09108. [PDF]

Boyden, E., et al. (2005). Millisecond-timescale, genetically targeted optical control of neuronal activity. Nat. Neurosci. 8: 1263-1268. [PDF]

McCabe, D. P. & Castel, A. D. (2008). Seeing is believing: The effect of brain images on judgments of scientific reasoning. Cognition: 107: 343-52. [PDF]

neurophilosophy Thu, 05/20/2010 - 21:50

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Gleanings: Illusions, genomics, Freud, lighting the brain, and eating alone

ddobbs — Wed, 12 May 2010 06:55:42 +0000

Gleanings: Illusions, genomics, Freud, lighting the brain, and eating alone

Traveling. But here's what I'm reading during train, plane, and bus rides -- and over meals:

Gravity-defying ramps take illusion prize.Â This contest always produces fascinating stuff. This time, the ball rolls up. Video here.

ï»¿

Vaughan Bell ponders cortisol, dopamine, neuroplasticity, and other things that set off his bullshit detector. Riff launched from a post from Neuroskeptic on cortisol and childcare scare stories, equally read-worthy.

Dan Vorhaus does a wonderful round-up of reactions and implications stemming from the news that genetic testing is coming to Walgreens. Best blog-post title cited there: "Chapter 38 of the Sky is Falling," a fine post by genomeboy Misha Angrist.

From Mark Bittman's blog, Suzanne LenzerÂ On Eating Alone. Particularly appropriate while traveling. And while I wish I were sharing these meals with my better half, I'm with Lenzer and Stephen B. Johnson in taking a real pleasure in eating good food while reading a good book:

The truth is eating alone is a treat. Now, when I'm in my chair or on my stool, menu in hand, I get to think about what I'm going to enjoy eating and drinking all by myself, ponder what I'm going to think about or read that I haven't had time forï»¿.

Last night it was gnochhi and The Selfish Gene. (On which more another time.)

InÂ Illuminating the brain, Lizzie Buchen reports on the growing use of light to turn different brain areas on and off. This carries significant potential as a cleaner, safer, easier, even more flexible replacement for the sort of deep brain stimulationÂ (DBS) modulators used to treat Parkinson's and in Mayberg et alia's experimental depression treatment discussed in yesterday's post.

Paul Broks on 150 years of Sigmund Freud Some useful reminders.

And via Vaughan Bell's most recent weekly round-up,

ï»¿AlthoughÂ autism is usually thought of as a disability, aÂ New Scientist article discusses the fact that the condition can be associated with various cognitive advantages.

ddobbs Wed, 05/12/2010 - 02:55

Tags

Brains and minds

culture of science

Genetics & genomics (incl behav genetics)

I love eating alone, too. MFK Fisher, the 1940s food writer, has some wonderful passages about eating alone with pride and pleasure in her memoir, 'The Gastronomical Me'...most notably, aboard a ship making a transatlantic crossing. I think about her every time I go to a cafe with a book or magazine.

Optogenetics controls brain signalling and sheds light on Parkinson's therapy

neurophilosophy — Tue, 24 Mar 2009 05:50:04 +0000

Optogenetics controls brain signalling and sheds light on Parkinson's therapy

Optogenetics is a newly developed technique based on a group of light-sensitive proteins called channelrhodopsins, which were isolated recently from various species of micro-organism. Although relatively new, this technique has already proven to be extremely powerful, because channelrhodopsins can be targeted to specific cells, so that their activity can be controlled by light, on a millisecond-by-millisecond timescale.

A group of researchers from Stanford University now report a new addition to the optogenetic toolkit, and demonstrate that it can be used to precisely control biochemical signalling pathways in the mouse brain and to manipulate complex reward-related behaviour. They have also used the existing channelrhodopsins to probe the neural circuitry implicated in Parkinson's Disease and thus gain a better understanding of why deep brain stimulation is effective in treating the disease.

The new work comes from Karl Deisseroth's lab, which has been instrumental in developing channelrhodopsins as tools for studying the nervous system. In the first of two papers, which appeared online last week in the journal Nature, Deisseroth and his colleagues describe how they manipulated the learning behaviour of mice using chimeric α₁- and β₂-adrenoceptors. These receptors are coupled, on the inner surface of the cell membrane, to small enzyme molecules called G-proteins; when a catecholamine neurotransmitter (adrenaline, noradrenaline or dopamine) binds to the receptor, the G-protein is released, and sets off to initiate multiple complex biochemical signalling pathways inside the cell.

The researchers replaced the neurotransmitter binding site with the light-sensitive part of rhodopsin, the vertebrate homologue of the microbial proteins, then injected the chimeric receptors directly into the nucleus accumbens of mice, which lies underneath the cerebral cortex and is commonly referred to as the "reward centre" or "pleasure centre". The animals were then made to perform a conditioned place preference test. This normally involves placing the animals into a chamber, and giving them a reward in a specific location. After repeated pairings of the location and the reward, the animals learn an association between the two, and subsequently spend more time in that location than in others.

In this case though, the researchers did not reward the mice with food pellets or a sugar solution. Instead, they used pulses of light, delivered to the nucleus accumbens via an optical fibre (below). Thus, whenever the mice moved into the designated location, the light activated the chimeric adrenergic receptors, and switched on the biochemical signalling pathways which would normally be initiated when the animals are given a real reward. Remarkably, the optical stimulation was sufficient to replace a real reward, such as pellets of food or a sugar solution, and as a result, the mice spent more time the following day in the location at which the stimulation had been delivered than in other parts of the chamber.

The second paper describes a set of experiments designed specifically to address the question of how deep brain stimulation (DBS) exerts its therapeutic effect on patients with Parkinson's Disease, the movement disorder associated with degeneration of dopamine-producing neurons in the midbrain. DBS is an experimental technique involving the implantation of stimulating electrodes into the brain. In Parkinsonian patients, the subthalamic nucleus is targeted. However, the circuitry in this region of the brain is complex, and neurons within it can respond to stimulation by either increasing their activity, decreasing it, or, over prolonged periods of time, both. Exactly why DBS is so effective in alleviating the symptoms of some Parkinsons' patients therefore remains unclear.

This set of experiments, described in the current issue of Science, was carried out in mice lacking dopaminergic neurons in the right half of the midbrain. The cells were destroyed with 6-hydroxydopamine, a neurotoxin which enters neurons via the dopamine and noradrenaline reuptake transporters, and therefore selectively destroys cells which synthesize these neurotransmitters. The mice treated in this way exhibited deficits in the limbs on the left side of their body, which are controlled by the right side of the brain, and as a result display rightward rotations when they move.

Deisseroth's group expressed channelrhodopsins in various cell types in and around the subthalamic nucleus, to investigate the effect of stimulating each of them in turn. First, one type of channelrhodopsin, which inhibits nerve cell activity when activated, was targeted to excitatory neurons in the subthalamic nucleus. These cells are thought to become overactive in Parkinson's, and it is widely believed that DBS may effectively ameliorate the symptoms of Parkinson's by acting on them. However, supressing their activity with light of the appropriate wavelength had no effect on the behaviour of the animals.

A second possibility is that DBS works by stimulating bundles of nerve fibres which enter the ubthalamic nucleus. To test this hypothesis, the researchers introduced another channelrhodopsin, which increases neuronal activity when activated by a different wavelength of light, into neurons in layer 5 of the primary motor cortex. These cells project down into the spinal cord, but their axons also send branches into the subthalamic nucleus. High frequency pulses of light delivered to these cells rescued the movement deficits of the mice, so that their behaviour was indistinguishable from that of normal mice. By contrast, low frequency stimulation of the same cells was found to worsen the animals' symptoms.

These studies contribute significantly to optogenetic technology and to our understanding of why DBS is an effective treatment for Parkinson's. The first paper demonstrates that optogenetics can be used to activate specific signalling cascades in neurons, and therefore provides a powerful new method for further dissection of the numerous biochemical pathways which modulate neuronal activity. The findings reported in the second paper raise questions about the hypothesis that DBS is effective in alleviating Parkinsonian symptoms because it exerts an effect on excitatory neurons in the subthalamic nucleus, and instead implicate the primary motor cortex as a primary target. They also suggest that DBS is at least partly efficacious because it induces oscillatory activity in the subthalamic nucleus, and that targeting white matter tracts (the bundles of nerve fibres which connect distant regions of the brain) would be effective in producing widespread therapeutic effects.

As for the future of optogenetics technology, the next logical step would be to develop wireless implants. This is exactly what researchers at Brown University's Nanophotonics and Neuroengineering Laboratory are aiming to do. They have already developed a dual-use "optrode" consisting of an array of 100 electrodes which can deliver light to specified neurons and simultaneously record the electrical activity of the cells, and a wireless brain implant which can record neuronal activity and convert the signals into a digital stream of light pulses. Combining the properties of these devices could therefore lead to a wireless optogenetic implant, which, if composed of light-emitting carbon nanotubes, might eventually result in smaller neural prostheses with increased biocompability and hence a longer lifespan than existing devices.

Related:

Airan, R. et al (2009). Temporally precise in vivo control of intracellular signalling. Nature DOI: 10.1038/nature07926.

Gradinaru, V. et al (2009). Optical Deconstruction of Parkinsonian Neural Circuitry. Science DOI: 10.1126/science.1167093.

neurophilosophy Tue, 03/24/2009 - 01:50

Tags

Neurodegeneration

neuroscience

optogenetics

brain optogenetics neurotechnology Parkinson's

neuroscience

optogenetics

Optogenetic therapy for spinal cord injury

neurophilosophy — Tue, 18 Nov 2008 11:18:08 +0000

Optogenetic therapy for spinal cord injury

Optogenetics is a recently developed technique based on microbial proteins called channelrhodopsins (ChRs), which render neurons sensitive to light when inserted into them, thus enabling researchers to manipulate the activity of the cells using laser pulses.

Although still very new - the first ChR protein was isolated from a species of green algae in 2002 - optogenetics has already proven to be extremely powerful - it can be used to switch neurons on or off in an extremely precise manner and so to control simple behaviours in small organisms such as the nematode worm.

Earlier this year, ChR was used to restore vision to blind mice lacking the light-sensitive photoreceptor cells in the retina. And now researchers from Case Western Reserve University in Cleveland, Ohio have used the technique to restore motor function in rats paralysed by spinal cord injuries.

Among the most common types of spinal cord injuries in humans are those involving damage to the cervical area of the spine, which begins at the base of the skull and extends down through the neck. The cervical spinal cord gives rise to the nerves which control the head, neck, arms, upper body and diaphragm, so serious damage to this area leads to complete paralysis, and is often fatal, due to paralysis of the muscles required for breathing.

Jerry Silver and his colleagues used an animal model of cervical spinal cord injury in their study. They sectioned one half of the cord at the upper cervical level, leaving their rats paralysed down on side of the body, and with breathing difficulties because of the half-paralysed diaphragm. During the procedure, they also injected a modified Sindbis virus, containing the genes encoding channelrhodopsin and green fluorescent protein, directly into the ventral grey matter of the spinal cord, where the cell bodies of motor neurons are located. Some of these motor neurons form the phrenic nerve, which innervates the diaphragm and controls breathing

Four days later, the animals' spinal cords were exposed again. The motor neurons were stimulated with blue light from a fibre optic cable, and the electrical activity of the diaphragm recorded. Brief episodic bursts of light were found to induce first erratic and then rhythmic activity, which became synchronised with the respiratory activity recorded from the other side of the diaphragm, aso that the rats could breathe normally. Remarkably, a longer repeating patteren of light was found to induce recovery of breathing which persisted for 24 hours.

Somehow, the spinal cord circuitry had adapted so that it could continue to generate the rhythmic patterns of activity required for respiratory movements long after the light stimulation had ceased. Analysis of the GFP expression pattern in the spinal cord showed that an average of 650 neurons in each experimental animal had been successfully infected with the virus carrying the channelrhodopsin gene. It also revealed that both motor neurons and interneurons had taken up the virus, and that some of the infected cells had processes which extended across to the opposite side of the spinal cord.

The researchers suggest that swiching on the infected motor neurons with the pulses of light had initiated a novel form of synaptic plasticity in the "crossed phrenic pathway". This pathway contains the projections of motor neurons which cross the midline and form connections with their mirror image counterparts on the other side of the spinal cord. Normally, these synapses are weak, so motor neuron activity on one side does not elicit activity on the other., but the strengthening of these crossed pathways seems to have been sufficient to generate rhythmic respiratory activity.

This and other research could one day lead to the development of optical neuroprosthetic devices consisting of remote-controlled light sources (perhaps light-emitting diodes) implanted into the body. Eventually though, optogenetics-based treatments for various neurological diseases seems plausible. The ability to use ChR proteins to both excite and inhibit neurons would have advantages - activating spinal motor neurons or neurons in the brain would be benficial in conditions such as amyotrophic lateral sclerosis or stroke and Parkinson's Disease, respectively, while inhibiting the activity of spinal neurons could prove to be an effective treatment for chronic pain.

Such treatments would of course involve first infecting patients with a genetically engineered virus, which poses major problems. In the meantime, Silver and his colleagues are using the same method to try to restore bladder function to the paralysed rats. They are also looking for ways to prolong ChR expression in the spinal motor neurons, and planning similar experiments in monkeys.

Related:

Controlling animal behaviour with an optical on/off switch for neurons

Brain-muscle interface helps paralysed monkeys move

Alilain, W. J. et al (2008). Light-Induced Rescue of Breathing after Spinal Cord Injury. J. Neurosci. 28: 11862-11870. DOI: 0.1523/JNEUROSCI.3378-08.2008.

neurophilosophy Tue, 11/18/2008 - 06:18

Tags

neuroscience

optogenetics

I just saw a poster on this at SfN! Nice timing.

This has been covered in a recent New Scientist article too: How Broken Nerves Can Be Fixed In a Flash.

Interesting. GFP and ChR will definitely help researchers in managing patients with many types of neurological disorders apart from spinal injury.

Hi I was wondering if you have any idea if optogenetics has benefits for Alzheimer's disease?
Could it possibily stimulate neurones using acetylcholine? hence relieving symptoms of alzheimers such as memory loss

@sarah: Optogenetics is an experimental technique that is being used in animals. Theoretically, it could be used in the development of treatments for various conditions, but this is still some way off in the future.

I'm not sure what you mean by "stimulate neurones using acetylcholine". Channelrhodopsins are light-sensitive proteins which do not have an Ach binding site.

Neuronal light switches

neurophilosophy — Wed, 24 Sep 2008 09:05:50 +0000

Neuronal light switches

The September issue of Scientific American contains an excellent and lengthy article about a state-of-the-art technique called optogenetics, by molecular physiologist Gero MiesenbÃ¶ck, who has been instrumental in its development.

As its name suggests, optogenetics is a combination of optics and genetic engineering. It is a powerful new method for investigating the function of neuronal circuits, based a number of light-sensitive proteins which have recently been isolated from various micro-organisms.

By fusing their genes to promoters which control where they will be activated, researchers can target the proteins to specified cells and thus render those cells sensitive to light. Importantly, each protein is sensitive to a specific wavelength of light and responds to it by changing its three-dimensional structure so that ions can flow into or out of the cell. This conductance can either increase or decrease the electrical activity of the cell, depending on the species of ion and the direction of the flux.

The activity of cells expressing these proteins can therefore be controlled by laser pulses. Individual cells can be made to express combinations of proteins, each sensitive to a different wavelength of light, so that, for example, blue light activates and red light inhibits neuronal activity. This control is remarkably precise - it can be achieved on a millisecond-by-millisecond timescale, such that a single action potential from a train of impulses can be eliminated.

In his article, MiesenbÃ¶ck describes how he and others developed optogenetics from existing techniques which use fluorecsent proteins as cellular dyes. He then discusses the work of a number of researchers who are applying the method to investigate how circuits of neurons generate simple behaviours, and goes on to look at its potential medical applications.

Related:

neurophilosophy Wed, 09/24/2008 - 05:05

Tags

Molecular Biology

neuroscience

optogenetics

Mo has some special skill at finding articles or research that hits targets of being interesting, beautiful and delightful or any combination thereof.

Lead with Sherrington's enchanted loom, by Sherrington's current successor, and the receptor for hard science is lit up and ready to proceed.
My first thought for future potential applicability for optogenetics and medicine was electrical: epilepsy.

My word, there are some clever types out there.

For a minute I thought the world of AI was going to get biological too.

Channelrhodopsin restores vision in blind mice

neurophilosophy — Tue, 27 May 2008 17:45:45 +0000

Channelrhodopsin restores vision in blind mice

New research shows that a protein found in green algae can partially restore visual function when delivered into the retina of blind mice, taking us one step further towards genetic therapy for various conditions in which the degeneration of retinal cells leads to imapired vision or complete blindness.

Normally, light entering the eye falls upon the rods and cones at the back of the retina. These are the photoreceptors: they are packed with a light-sensitive protein called rhodopsin, which initiates an electrical signal when struck by photons (the particles which carry light).

The signals generated by the photoreceptors are transmitted to the bipolar cells, which in turn transmit the signals to ganglion cells, whose axons leave the retina to form the optic nerve. Thus, visual information is carried from the eye to the visual centres of the brain, via the lateral geniculate nucleus, a relay station in the thalamus.

In the retina, visual information passes along two separate and parallel channels, each one most sensitive to different types of light stimuli. Photoreceptors in the ON centre pathway are sensitive to bright stimuli against a dark background, whereas cells in the OFF centre pathway are sensitive to dark stimuli on a light background.

Bipolar cells receive converging inputs from several photoreceptors, and so have similar receptive field properties. ON bipolar cells are most responsive to a light spot on a dark background, and vice versa for OFF bipolar cells. The cells in the retina are therefore well-suited to detecting contrast, which is ideal for recognizing the edges of objects.

Contrast is further increased by a process called lateral inhibition, whereby activity in one pathway reduces activity in the other. In this way, the circuits in the retina carry out simple processing of the visual data entering the eye. They generate a very crude representation of the visual image, which is then sent to the brain for further processing.

The new research, led by Botond Roska of the Friedrich Miescher Institute for Biomedical Research in Basel, Switzerland, and Connie Cepko of Harvard Medical School, is reported online in an advance online publication in Nature Neuroscience. It uses a protein called channelrhodopsin-2 (ChR2), a light-sensitive ion channel that was recently isolated from the green algae Chlamydomonas reinhardtii.

ChR2 belongs to a family of proteins which are also found in bacteria, and are related to the rhodopsin proteins found in the retina of mammals. Unlike their mammalian counterparts, the channelrhodopsins are directly linked to a pore which spans the membrane of the cell, and can open or close in response to light of a specific wavelength and so alter the flow of electrical currents across the membrane. They can therefore confer light sensitivity onto cells that do not normally respond to light.

It is this property of the algal protein that was exploited in the new study. In conditions such as macular degeneration and retinitis pigmentosa, there is a progressive loss of photoreceptors. The researchers therefore reasoned that they could bypass the photoreceptors entirely, and restore visual function by using ChR2 to make the bipolar cells responsive to light.

The study was carried out on a strain of mutant mice which lack a gene that is required for the survival of photoreceptors, and which provide a good model for human retinal degenerative conditions. These animals lose most of their photoreceptors by 4 weeks of age, by which time, light stimulation does not elicit any detectable electrical activity in retinal cells. In other words, by that age, the mice are virtually blind.

The researchers first created a genetic construct consisting of the ChR2 gene fused to the gene encoding a fluorescent protein. This construct was then delivered to the animals' retinas by in vivo electroporation, whereby cells are made transiently permeable to DNA by the application of a small external electric field.

The construct also contained a regulatory DNA sequence called a promoter, which normally drives the synthesis of a glutamate receptor in a specific cell type: the ON centre bipolar cells in the retina. So, when delivered to the retina, the construct may enter various types of cells, but will only be activated in the ON bipolar cells.

In mutant mice lacking photoreceptors, targeted expression of ChannelRhodopsin 2 (ChR2) confers light sensitivity upon bipolar cells. Bipolar cells expressing ChR2 are stained green. Scale bar = 10 micrometers. (From Lagali et al, 2008.)

Once inside, the construct is incorporated into the chromosomes, so that cells containing it synthesize both the fluorescent protein and ChR2. This should make the bipolar cells sensitive to light, and an electrode array which recorded electrical activity in the ganglion cells in response to light confirmed that this was indeed the case.

Thus, in the absence of photoreceptors, the bipolar cells synthesizing Chr2 were sensitive to light, which activated them, so that they generate an electrical signal which is then transmitted to the ganglion cells. The number of signals recorded was found to dependent on the intensity of light, so that the more intense the stimulus used, the more frequent were the bursts of ganglion cell activity.

This was further confirmed anatomically. The parallel ON and OFF bipolar cell streams have slightly different projections, with their processes terminating in two different layers of the retina, and confocal microscopy (above) showed that the cells targeted by the ChR2 construct project to the outer layer only, which is characteristic of ON bipolar cells.

Furthermore, electrophysiological recordings further showed that the responses of the bipolar cells are transmitted to the brain: when the experimental animals were presented with light stimuli, the electrodes detected corresponding activity in cells in the visual cortical areas.

The new ability of the previously-blind mice also affected their behaviour. When placed in a box that is divided into light and dark compartments, the movements of normal mice, but not of blind ones, increase. And when the mutant mice with ChR2-expressing bipolar cells were placed in this apparatus, their movements were comparable to those of normal animals.

Elsewhere, researchers are trying to develop retinal implants for the treatment of eye diseases. A therapy based on ChR2 would have the advantage of not requiring the implantation of foreign metallic objects (electrodes) into the retina. And, because it involves genetic targeting of a specific type of retinal cell, it is an improvement on previous similar work, which has been less selective.

Although the results of the present are very promising, there are several problems. First, the ChR2 protein was introduced into less than 10% of the bipolar cells in the retina, and was not evenly distributed; the bipolar cells were sensitive to a far narrower range of light intensities than are photoreceptors; and they required a far greater intensity to be stimulated into action.

Finally, it should not be assumed that the animal model faithfully reproduces all aspects of eye diseases in humans. It is unclear, for example, how the connections between retinal cells remain intact during the course of progressive degeneration. Undoubtedly, these issues will be addressed in future work.

Related:

Lagali, P. S., et al. (2008). Light-activated channels targeted to ON bipolar cells restore visual function in retinal degeneration. Nat. Neurosci. doi: 10.1038/nn.2117. [Abstract]

neurophilosophy Tue, 05/27/2008 - 13:45

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:-), http://www.gravatar.com/smokelesscigarettese smokeless cigarettes for you, rhsd,

Channelrhodopsin_restores_vision.. Ho-o-o-o-t :)

I'm just thinking..WOW! I hope that this pans out to something translatable to humans.

Broken link reported (2 nature neuroscience).

Thanks - it's fixed now.

Thank you for commenting on our work. I only want to point out one error. The electroporated gene most likely does not integrate into the chromosomes. This is important as it might limit the time during which the gene would be expressed. Alternatives, such as viral vectors that do integrate, or at least remain stably associated even if they do not integrate, would be preferable and will be tried.

Control of rodent motor cortex with an optical neural interface

neurophilosophy — Fri, 24 Aug 2007 10:38:33 +0000

Control of rodent motor cortex with an optical neural interface

This year, several research groups have used bacterial proteins called channelrhodopsins to develop a technique with which light can be used to control the activity of nerve cells or the behaviour of small organisms.

For example, Ed Boyden's group at the MIT Media Lab used the method to activate or inhibit neurons on a millisecond-by-millsecond timescale, while Karl Deisseroth and his colleagues at Stanford have created an optical on/off switch that can control the movements of the nematode worm.

Devices employing such technologies could in theory be used in advanced neural prostheses for a variety of conditions, such as epilepsy and depression, and neurodegenerative disorders such as Alzheimer's and Parkinson's. However, there are several major challenge to achieving this.

First, the blue light used for photoactivation of the channelrhodopsin proteins is highly scattered by brain tissue, so, while the technique can be used in nematode worms , using it in larger animals is far more difficult. Secondly, it is not clear excatly how light could be targeted to the specific region of brain in which the channelrhodopsins are being expressed.

Deisseroth's research team has now taken this new technology one step further. In the Journal of Neuroengineering, they describe an optical neural interface which they used to control the whisker deflections in living, intact rats and mice.

Deisserdorf and his colleagues first delivered the channelrhodopsin (ChR2) gene into slices of rodent brain using the previously established method, in order to establish the extent to which light would be transmitted through the tissue.

They cloned the gene and inserted it into a viral vector, together with a genetic element called a CaMKII alpha promotor that drives the protein to be expressed specifically in neurons. Using a fiber optic guide which served as a cannula, the researchers then delivered the gene construct into motor neurons within the brain slices. Antibody staining showed that the ChR2 gene had indeed been delivered to the motor neurons, and electrophysiological recordings confirmed that the cells were activated in response to beams of blue light.

Once they were satisfied that this method could effectively deliver the ChR2 gene to the specified cells and that light could penetrate the brain slices sufficiently, the researchers then set out to develop a method by which the ChR2 gene could be delivered into the brains of living, behaving rodents. They targeted the vibrissal motor system, the part of the rodent brain that controls whisker movements. Thus, the level of optical control could be quantified by the deflection of the deflection.

The neural interface developed consisted of an optical fiber guide, fixed to the skull with a stereotactic rig (see above). The guide was first used as a cannula to deliver the ChR2 construct to into excitatory motor neurons in layer 5 of the vibrissal motor cortex. Then, an optical fiber was inserted into the cannula, so that the light beams were directed at exactly the same cells to which the ChR2 was delivered.

It was found that the interface could effectively control the output of the motor neurons in the vibrissal system. Small magnetic particles were attached to the animals' whiskers, so that their movements could be observed using a magnetic field sensor. Pulses of blue light delivered to the motor neurons via the optical interface caused the whiskers to deflect.

As in the earlier work, the ChR2 construct was delivered to a specific set of neurons. When expressed in the motor neurons, ChR2 made the cells sensitive to blue light, which could then be used to control the activity of the cells on a millisecond-by-millisecond timescale. Deisserdorf's team also used the optical interface to control the whisker movements of mice, whose brains are somewhat smaller than rats.

In the future, such techniques could be used to develop neural prostheses whose optics are either anchored to the inside of the skull or embedded with the brain tissue. Such optical devices would have a number of advantages over those used currently (e.g. the electrode arrays used for deep brain stimulation or the magnetic coils used for transcranial magnetic stimulation). Their size would be comparable to that of the electrode arrays currently being used.

For example, photoactivation can be used to excite specific cells, but electrode arrays cannot. Also, the electrode cables are susceptible to becoming encapsulated by glial cells within months of being implanted; optic fibers would be less so. And, the magnets used for transcranial magnetic stimulation can only deliver magnetic fields superficially; the current work shows that optical interfaces can be used to activate neurons within deep brain structures. Finaly, the long, light and flexible optical fibers would effectively keep heat-generating elements away from target tissues.

Reference:

Aravanis, A. M., et al. (2007). An optical neural interface: in vivo control of rodent motor cortex with integrated fiberoptic and optogenetic technology. J. Neural. Eng. 4: S143- S156. [Full text]

neurophilosophy Fri, 08/24/2007 - 06:38

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