So the second thing the researches wanted to test was whether the electrical stimulation was stimulating glial cells to secrete inhibitory chemicals. To do this they introduced channel rhodopsin (this protein activates cells wheras the halorhodopsin inhibits them) using viruses but put it under a glia-specific promoter so instead of excitatory cells expressing the protein it was the surrounding glial cells. Here are the results:
With all studies like this the researchers must include histological verification that the proteins they’re trying to express are, indeed, being expressed, and in the right places. Here we see a stain for GFAP (a glial-specific protein) in green and the channel rhdopsin in red. In the overlay it is clear that there is strong colocalization. In short, the protein is where it should be. It’s very important to check but fairly repetitive and uninteresting so I’ll neglect it in the next few figures.
The proteins are activating proteins so you might expect to see an increase in spiking, but… we don’t. We don’t because the cells creating those large spikes are the neurons, and the neurons were not being activated by the light. Instead the light was activating the glia which secreted chemicals that inhibited the surrounding neurons resulting in an overall decrease in firing.
This treatment had no effect on parkinson’s symptoms.
The researchers found that causing the excitatory cells in the STN to fire at normal rhythms (by supplying light pulses to transfected cells at those normal rhythms) also had no therapeutic effect.
At this point the researchers take a moment to prove that the light they’re supplying is really hitting the places they think it is. They take small samples of brain tissue and simply shine their lasers through it, measuring the intensity at various distances. The light-activated proteins being used require a light intensity of about one mW/ square mm.
The top graph shows that the two colors of light being used each are sufficiently intense after passing through up to 1.5 mm of tissue. Below shows what cells have been activated by the light. Neurons produce the protein c-fos when they are activated which can later be stained for. By measuring how far from the epicenter of the light application c-fos appears the researchers can determine what volume of tissue could have its light-sensitive proteins activated. It turned out that about 1 cubic millimeter of tissue received bright enough light. The viral injections that introduce the DNA for these proteins affect about one cubic millimeter, so any infected cell can be successfully activated by this light application technique.
Up next: an optogenetic interference that had therapeutic effect. Stay tuned!
This past winter semester I took a class about recent studies into complex behavior. The majority of the papers we read were about fruit flies’ mating habits and other model organisms, investigating the very basics of how flies decide who to mate with, for example. The last portion of the class we individually prepared presentations about papers often having to to with mammals and less fundamental research. I thought I’d share my presentation on a paper ( Gradinaru et al. 2009) whose goal was using optogenetics to determine why deep brain stimulation (DBS) actually works. So DBS stimulation, in essence, involves sticking an electrode down into the brain to the subthalamic nucleus (STN), a part of the brain involved in motor control. The electrode injects current at a high frequency and this often eliminates Parkinson’s symptoms, but nobody knows why.
Electrical stimulation affects all cells nearby, be they neuron bodies or glial cells or even axons passing near the electrode, so even though it is known where DBS occurs it’s harder to say which cells being affected by DBS are having a therapeutic effect. The leading hypotheses regarding the are that excitatory cells in the STN are inhibited by the electric stimulation, that astroglia (nearby support cells) are activated and secrete a chemical which inhibits excitatory cells in the STN, and that the cells of the STN are firing in the wrong pattern. To attempt to tease these different possibilities apart from one another the investigators used optogenetics in hemiparkinsonian rats. So firstly, hemiparkinsonian rats are rats that have half (hemi) Parkinson’s, which is achieved by applying a chemical to one side of their brains’ motor circuit, causing a lesion which leads to Parkinson’s symptoms only in one direction. One result of this is that the rats “rotate” or walk in circles sometimes. DBS can be given to these rats which eliminates their rotations and other Parkinson’s symptoms. Optogenetics is the introduction of light sensitive ion channels/pumps into cells using (in this case) viruses. The damaging parts of the viral DNA are removed and replaced with genes coding for these light-sensitive proteins and these genes are under the control of DNA regulatory elements called promoters which are only activated in certain cell types. The result is that you can inject a small amount of these viruses into an area that you’d like to stimulate, the viruses insert the DNA into all the cell bodies around that area (the viruses don’t tend to inject into axons passing through), but only cells capable of driving the promoter (cells that produce the necessary transcription factors) that has been injected will actually make the protein. The protein produced will find its way to the cell membrane where it is ready to activate or deactivate that cell upon application of light of the appropriate wavelength (color). I’ll go over one figure now and leave the rest for later. Here are the results from Figure 1:
CaMKIIα::eNpHR refers to the viral DNA setup used. It was Halorhodopsin (eNpHR) under the calmodulin-dependent kinase type II promoter (found specifically in excitatory cells (those that use glutamate as their neurotransmitter)). The black band across the middle is an electrical recording of the area being stimulated with light. Voltage is on the Y axis, time on the X. The yellow bar shows when yellow light is applied (halorhodopsin is most sensitive to yellow light). The graphs at the bottom show the same thing.
The chart on the left and the black bar in the right shows that the rotations were unaffected by the light. The red bar shows that electrical stimulation (DBS) was indeed effective (as was known and expected).
The conclusion from this is that deep brain stimulation is not effective (at least solely) because it inhibits the activity of excitatory neurons in the STN.
The more astute among you may have noticed that I haven’t posted anything here for a while. I aim to change that. I aim to change it because I think being able to write things that people want to read is a valuable skill.
A massive portion of my time is spent learning things. I learn lots for my classes, but in my leisure time I read science and technology blogs and non-fiction books, and one of my favorite things is talking to people about all these interesting things I find out about. Hopefully this blog can be yet another venue for talking about interesting things with interested people!
Now a little catch-up.
- I’m back in Michigan.
- I no longer start to say “xie-xie” instead of “Thank you” or “ni-hao” instead of “hello.”
- I am still very good at using chopsticks
- My major is now Molecular/cellular/developmental biology with some biomedical engineering classes thrown in (on course for a BME masters)
Last semester I took a programming class, multivariable calculus, and thermodynamics. I’m currently in E+M physics, moral philosophy of contemporary issues, bioreactions, and genetics and molecular biology of advanced behavior (a literature-based biology class).
My plan is to explain some of the papers we’ve been learning about in my biology class which have been focused on a variety of complex behaviors like courtship, motor control, circadian rhythms in the mouse and fruit fly but seem to have at least equal weight on the techniques used to carry out the research which are really incredible. I’ll do my best to explain them as they come up.
This past week I attended the 2nd Beijing International Symposium on Computational Neuroscience at Tsinghua University. It was a one day conference all about computational neuroscience and it was a really great day.
You’re probably wondering what computational neuroscience is. I wasn’t exactly sure myself, but after a day’s immersion in it I feel able to describe it. Computational neuroscience is looking at things that brains do and trying to figure out a way that they might be doing them using computational models. For example, the last speaker of the day was examining how people learn to do a series of steps that results in varying rewards depending on the steps chosen. His models said that people, and animals, partially use a full and detailed prediction of each choice, the choices that the first choice makes available, and so on and logically say “well if I want such and such a result I can just follow this series of steps.” He asserts that with more experience animals begin to abandon the perfectly logical, but cognitively taxing, exact method and substitute a method of choosing wherein each possible choice at a given stage just has a sort of feeling attached to it that says “this choice seems to work out pretty well” and for each subsequent step the animal just picks each option that seems like it will work well, without explicitly planning. So he has this model and can simulate how animals would behave if they were following one or the other of these two methods or a hybrid of them and he then puts animals (people) through a test and sees whether he was able to accurately predict their choices using his computer model. And voila, he has done computational neuroscience. He now has a model that, while it is not really how the brain is operating, does a good job of approximately achieving the same result as a real brain in this particular circumstance.
So the conference was a whole-day affair. I got there in the morning for coffee and snacks and soon the festivities began. A professor from Tsinghua gave an opening remark that, summarized, went a little like this: “Welcome, we’re glad to have you all. BTW, Tsinghua is developing a neuroscience program and if any of you want to come here will pay you well and if you’re a new professor we’ll hook you up with some sweet stuff. Seriously, we’re really awesome.” It was almost obnoxiously smug sounding, but I would be smug too if I could actually offer that. The day consisted of 4 talks by notable people in the field and 3 periods of time to go look at posters. It was followed by an open question session and a buffet dinner.
I met a nice German masters student who I talked to a fair amount throughout the day as well as a number of Chinese students.
I found the whole affair very interesting as I didn’t know much about computational neuroscience to start with and had never been to a conference like that. It was nice to see people talking about receptive fields and other neuroscience things and then having a whole room of people seem to understand what was being said! I also found it humbling how often people just had to say “I don’t know” to curious questions from the audience. Also, during the discussion section at the end questions were raised like “if we can accurately model all the activity of the brain, does that mean we really understand the brain? Does it matter?” and “Is computational neuroscience going to be helpful in the near future?” and I was struck by how… simple the discussion was. These professional neuroscientists could offer no more credible or more solid arguments than I could and the whole discussion seemed to be pretty much about philosophies. Whether or not it is enough to simply be pragmatic and satisfied with a model that works and how hopeful you were willing to be about the applications of cognitive models to computer programming and robotics.
I really enjoyed the conference. It was great to spend a day hearing about things I hadn’t known about before and it was definitely a break in the routine. I look forward to more such events in the future!
I have much to tell you, dear readers, but I’ll break it up into multiple posts for coherence.
This past weekend me and 13 of my companions set off on a trip to Huhhot for Saturday night through Tuesday morning. Huhhot is the capital of Inner Mongolia, which is not a part of Mongolia but rather a province of China that borders Mongolia. As we left Saturday evening we saw Anya, who was just returning from the hospital for the day. She had been there getting some IVs and otherwise being looked after. Due to her illness she was no longer going to go on the trip she had planned nearly single-handedly and she had told us before that while it was not finalized she was probably going home either Monday or Tuesday morning, which meant that we would not see her again as we arrived back in Beijing Tuesday morning. We said our goodbyes and took some pictures and gave some hugs but had to head out to catch our train.
The train station was unreal. I have never, ever, been in a place so crowded as that. The best way to describe the people moving into the train station was “a crush.” Seriously. I took pictures and videos to prove it.
We had soft sleeper tickets for our train voyage, which made the trip very pleasant. (Dear future China Traveller: train tickets go on sale 10 days prior to the train’s departure, roughly. Get there when they go on sale to be sure you get what you want. On the way to Huhhot we were at the ticket counter (you can buy tickets on PKU’s campus) when the tickets went on sale and got 14 consecutive soft sleeper tickets. Since we were coming back 2 days after we left we had to go a second time to buy the return tickets. We got there about 30 minutes after these tickets went on sale and as a result got a slightly disjointed group of tickets: 2 full rooms of four, 1 room of 3, and I guess another room of 3…) Train tickets come in a variety of types. Depending on the train type there are Hard seats, soft seats, hard sleepers, soft sleepers, and deluxe soft sleepers. Hard seats are like coach airplane seats, only the catch is that there aren’t as many seats as there are tickets, so it’s possible that you’ll be stuck standing for your train ride (as some of Andrew’s group was when they went to Mount Tai). I’m told that this is a horrific and traumatizing experience, and I believe it. Soft seats are fairly nice seats I believe. I don’t really know that much about them. Hard sleepers are rooms with 6 cots in each, they are cushioned with a bamboo mat and there is no locking door on the room in which you’re sleeping. Soft sleepers have 4 beds which are cushioned with a fair twin mattress, they are moderately air conditioned and have a locking door. Deluxe soft sleepers are like a hotel room on a train I think.
The important thing is that we had soft sleepers, and it was a good time hanging out, talking, and playing games until we decided to get to sleep. Most people slept fairly well on the train but we suspect it really wasn’t the most restful of sleeps possibly due to the jostling of the train.
We arrived in Huhhot and were whisked conveniently to our hostel, the Anda guesthouse, by people from the guesthouse. We settled in briefly and then set off on our “grassland tour.” This began with a harrowing 2 hour drive out to the grasslands. Harrowing because the people of Huhhot laugh in the faces of both death and traffic police and all-together drive like every one of them had a backseat full of dying orphans who they needed desperately to get to a hospital. The main thing that was distressing was passing. My policy about passing other cars is to do so when you’re 100% confident that you will not be blindsided by a car coming around a corner or over a hill or anything like that. Inner Mongolian’s policy about passing is “go for it! We can totally fit three cars abreast on here!” and indeed they can. I even saw four cars abreast on the 2 lane road at times, because why else would there be shoulders than to allow incredibly reckless driving? Our driver and others would consistently pass other vehicles on the two lane road as we went around hills on blind curves. A few times cars would come whipping around head on toward us and only by both drivers cooperative effort, and the car being passed’s complacency, did we not crash. Looking back I’m somewhat disappointed in myself for not firmly requesting the driver to acknowledge his mortality and try to not kill us all, but that wouldn’t have been nearly so interesting…
So after our harrowing journey we arrived at this small outpost in the grasslands where there were a couple of yurts and a stone building that apparently had a kitchen in it. We relaxed in the yurts and had a tasty lunch of lamb (when in a place near Mongolia…) and veggies and other good stuff. After lunch we set off walking on the grasslands for a while. We went over to a nearby herd of cows and a dried up lake bed. It was nice to enjoy the fresh air and clear skies. Following the stroll we went horseback riding which everyone enjoyed quite well, including those who were originally skeptical. The primary complaints were sore butts from some and sore knees from those of us whose stirrups were far too short. The horse were alright, but some didn’t seem to be the healthiest of beasts.
In the evening we had dinner at the hostel and explored the nearby streets. There was a short mongolian singing and dancing put on randomly at the hostel which was sort of cool. We hung around the hostel and talked amongst ourselves as well as with some of the other guests about what to do the next day and all sorts of other things. We met an Irish couple and an English guy who had both been in Huhhot for a few days and had recommendations. It was a fun night.
The following day we hit the city. We visited the 5 pagoda temple, which was a nice little place. We then walked up to the muslim part of the city in search of muslim-type souveniers and stuff but didn’t have great success. After this we attempted to go to the Inner Mongolian museum which we heard rave reviews about and which was recently remodeled, but to our dismay it was closed (it’s closed every Monday, as of the time of this posting). On the plus side it was very nice looking and was probably equally nice inside… We split up after that because some… animosity and confusion was stewing throughout the day amongst our small group. I headed back to the hostel and relaxed and played cards for a while. Eventually, with directions from the hostel staff, Veronica, Tram (Veronica’s labmate who came on the trip with us), Chanan, and I went for a walk. We first visited a nearby park, which was pleasant and park-like. We then went to a market for some shopping, followed by dinner, followed by the train.
On our train ride back our tickets were more scattered and the first thing that happened when we got on was swapping. Two guys traveling together were in different rooms and trying to get in the same room. One’s ticket was in the room with Veronica, Tram, and I, the other was in a room with Chanan, Lester, and a nice lady. After much bustling and some wonderful translating from Ray it was determined that the nice lady would move to the room I was in, and the guy from our room would go to Chanan and Lester’s room. Veronica, Tram, and I had a good time communicating with her, explaining why we were in China, that Veronica’s mom was Chinese but she lived in the US, that we’d never been to China before, and that we were 20/21 years old. After that our vocabulary pretty much ran out and Ray came in and translated for/with us, and after a bit of that he just talked with her in Chinese while Veronica and I amused ourselves with drawing and talking and Tram retreated to her bed to read.
In the morning we learned that as we were arriving in Beijing Anya was boarding her flight to the states. Fortunately we had said our goodbyes before we had left.
Everyone dispersed either to shower at the hotel or straight to their labs and life returned to normal +1 interesting trip in our memories.
We learned last week on Friday that Anya had Epstein-Barr Virus (EBV), the virus responsible for mononucleosis, or “mono.” This means that she has had to go home early. She went home this past Tuesday, the 20th because she was doing better but all involved felt she’d probably be better off being home in case anything further happened.
So Anya has survived, and she is now back in the United States, and we miss her.
It’s now Friday. Anya has been feeling sick since last week some time, which nobody thought to be a big deal because a few people have gotten sick recently and just suffered a couple days of sore throats, stuffy noses or malaise. On Wednesday I ran into Anya leaving the hotel to get some more water and I started chatting with her and immediately noticed that she looked funny. She looked funny because her eyes, and less obviously, her skin, were yellow! Somewhat alarmed I asked her if she realized that she was yellow looking and how long she’d been so and how she felt. She said that she’d noticed but was feeling alright and she’d decided with Han that if she started feeling bad she’d go to the hospital. I told her that the only cause of being yellow, jaundiced, was liver failure, and I urged that she should probably go to the hospital right away since your liver is sort of important. I didn’t hear more about it until later that day, but apparently Han and Frank took her to the hospital that Han does her shadowing at, which is supposed to be the best hospital in China and they looked at her and determined that her bilirubin levels were, indeed, high and therefore something was wrong with her liver. They put her on some drugs (one I.V. every 12 hours (I think) one pill every 8 hours, I’m not sure about what the drugs are, though) to help her liver and started trying to figure out what was causing the whole situation. The best guess was some kind of virus. Her symptoms as far as I know them were being incredibly tired all weekend and at least through Thursday, getting easily exhausted (such as after a 10 minute walk just wanting to lay down and sleep), a slight headache and a tenderness to pressute just below her ribcage. She’s in the international section of the hospital so her doctor speaks passable English, but it’s not 100% fluent. Han is staying with her as much as possible (just going home to sleep) to help translate and deal with crap so Anya can rest as much as possible (Frank may have been there Wed. night as well…). So she spent Wednesday night in the hospital.
Thursday morning they started tests to see if she has a number of likely viruses. One of those tests will be done this afternoon, the last one might take as long as 5 days. Throughout Thursday she continued with the regimen of lots of sleep, lots of water, IV and pill drugs. Han was there with her on Thursday and I found out that we were basically waiting for bacteria to grow again so I went over to visit in the afternoon. After I got off the subway and was walking to the hospital a couple of nice Chinese art students came up to me and started talking and then invited me to their end-of-semester art show. Since I wasn’t really in a rush I went with them and talked with them about how their studies worked and about some of the paintings. They then mentioned that the art was for sale since this was the last day it was on display and the sale of the art helped support the school. I asked how much they were, assuming they would be exorbitantly expensive but was surprised that they were only 10-20 dollars for most of them. I ended up getting a few nice paintings.
So, after the impromptu art fair I got to the hospital, the old side of which has traditional Chinese architecture.
I met up with Han who filled me in on most of the above information while we waited outside Anya’s room since Anya was sleeping. We decided we should make a get-well card (which I just realized I need to take a picture of!) complete with hand-turkeys and virus-punching lymphocytes.
Eventually Anya woke up and we talked with her and asked how she was feeling and all that jazz. She was still tired but starting to get hungry so she went back to sleep while Han and I got some lunch/dinner and brought Anya back some bao zi and porridge. Later on Anya was better-rested so the three of us just talked for a while and Anya had some food. The doctor came in to check on her and it was determined that she’d spend the night in the hospital again since she needed to continue getting IV drugs and the stress of moving back to the hotel and just going to the hospital for IV’s sounded like too much. Han and I left in the evening. Anya spent Thursday night at the hospital, but appeared to be in good spirits.
This morning Han went back to see how she was doing. The doctors were planning to check her bilirubin levels again to see if the drugs appear to be helping, and later in the day the results from one of the virus tests should be in.
That’s all I know at this point, but we’re all hoping she feels better soon.