Sunday, November 18, 2007

[LJ Repost] My experience with lab mice

Recently, I visited my friend Rachael. We were talking about our respective summer activities, and since I was working in a neurobiology lab at the time, the topic of animal research came up. Rachael is many things, including (1) highly empathetic; (2) hyperenthusiastic about animals and the environment; (3) very protective of, and caring for, the same. The Rachael I knew back in elementary school probably would have had a fit at the thought that her friend could be involved in something as barbaric as animal research. But we're not eight years old anymore, and instead of going on a long animal-rights diatribe, she just asked me how I dealt with it. I couldn't really give a good answer in realtime, without a chance to think through it. I said something along the lines of "I just push it aside. Yes, it's disturbing, but we try to be as humane as possible, and I push it aside."

It is difficult to deal with the suffering and death of research animals.

I've spent two summers under grad students in a certain neurobiology lab at Stanford. Luckily, the only organisms we worked with were mice and bacteria. (And do I ever feel sorry for the people who work with monkeys.) This post will be about mice. Bacteria are not worth discussing.

Actually, the first time I worked with mice was the summer/fall of 2005, when I interned in a different neurobiology lab, under a scientist named Helen. I only went in once a week, and I hadn't taken biology yet so I hardly knew anything about what we were studying, couldn't really make a substantive contribution...blah blah blah. I'd had pet hamsters before, but that was the first time I had to deal with mice in a research setting.

I remember how freaked out I was the first time I watched Helen handling the mice. To pick them up, you pull them up by the tails and then scruff them. To obtain a very small tissue sample for genotyping, you cut off a bit less than a millimeter of their tail, with a razor blade. To give them each a unique identifying number, you use a predefined code that involves cutting off toes (and sometimes punching holes in ears, though we didn't use that). The mice would get frightened, squeak, writhe, urinate, try to escape, try to bite, and eventually, bleed. Each mouse didn't bleed much, but I was quite surprised at the amount left on the workspace when we finished numbering and tailcutting a couple cages' worth of mice.

Thankfully, that was as much as I had to do while working with Helen. I did do some sectioning and staining of brains, but they had already been dissected out, and I was able to think of them as just tissue. Meat. I suppose I knew, academically, that someone had had to kill mice and dissect them to obtain these brains, but I was one step removed from that and it didn't really hit me. They were just excised brains, just grey lumps in a dish; they didn't look at me with cute little button eyes and squeal, demanding to be put back in their cage with all their paws intact.

At one point, Helen and I walked past a guy who was doing a more elaborate procedure. The mouse was on its back on a styrofoam block, pinned by the paws -- essentially crucified flat. Its chest was cut wide open, and I could see its heart beating. There was blood everywhere, and there were lots of tubes going back and forth from the mouse to this little whirring apparatus in a corner. It would have thoroughly blown my mind, but I didn't catch more than a brief glimpse. Anyway, except for that, my mouse involvement was relatively pain-free in that lab.

I got a bit of a shock when I started in the second lab, in summer 2006 (working with a grad student named John). We were doing a project that involved acutely isolating cortical astrocytes. Translation: killing young mice, dissecting out their cerebral cortices, and processing these in various complicated ways to finally end up with an isolate of single cells of a certain type. That procedure (`prep' for short) takes pretty much all day.

Hah, I'd thought tailcutting and toecutting were bad? We were working with very young mice, recently weaned or not yet weaned. I think the age range was roughly from 1 to 10 days. They were still pink and didn't even have their eyes open yet. With adult mice, you gas them first, but to kill a young mouse, you behead it with scissors. (I think the protocols are similar for rats, except sometimes you use this guillotine-looking thing. I never used that, or saw it used.)

You behead the mouse with scissors, with about inch-long blades. The body twitches, and the head falls onto the table, and it twitches too. Blood wells and drips out of the body, reddening about a square inch of the absorbent pad you do dissections on. (You get quite a bit more blood from an adult mouse.) You discard the body in a biohazard bag. Meanwhile, the head is sitting on the table, looking for all the world like a live mouse, except there's empty space behind its neck. The jaw opens and shuts, which makes the head rock back and forth. It takes about ten seconds to go quiescent, and then you unroof the skull and dissect out the cortex.

Sometimes, when you put the scissors up to their neck, they squeal and put their little hands up, and grasp the blade, and you can't behead them without cutting off their fingers.

The first time I watched John do it, and the first time I did it myself.....I'd like to say I felt faint, or nearly threw up, or something overt like that. But I just felt a deep sort of quiet horror that didn't lend itself to being expressed that way. I didn't feel a physiological effect, like faintness or nausea; just quiet horror and mental revulsion. But I wondered, what was wrong with me? Why wasn't I more upset? As unpleasant as the experience was, I wanted it to feel more unpleasant. I didn't want to feel the beginnings of numbness and desensitization.

I did an awful lot of preps that summer, about two per week on the average. And don't get me wrong, I enjoyed most of the tissue culture work, the part where you're working with bits of tissue or cells in a tube, instead of with (mostly-)entire animals. The summer's work as a whole was fun and challenging and rewarding and all sorts of other awesome adjectives. But I was doing two preps a week, often under tight time constraints, so I didn't spend much time hyperventilating over dead mouse pups. I didn't actually become numb, but I grew much better able to put the horror aside quickly and move on to the next thing that needed to be done. And then after five minutes, the disturbing part was over, and there were just tissue bits in a dish. Meat.

One thing that helped was perverse, macabre humor. It made me realize anew what awful things we had to do, while helping me cope and smile. I guess everybody got into the black humor to some extent -- I sure wasn't the one who put up the "Dr. Kevorkian wants YOU to keep the euthanasia area clean" poster. A lot of people were in the habit of referring to older pups as `pupcorn', because they would jump around enthusiastically when you opened the cage, and they could easily jump high enough to escape if you weren't careful. But my favorite was a random thing that happened to me. Immediately after beheading the pup, you cut away the skin on top of its head using small scissors. John's small scissors squeaked. The first time I used them, I jumped about a mile all "OMG OMG OMG it's still alive it squeaked WTF OMG AAAAAAAACCCCKK", until I realized that the mouse head had no vocal tract, so it couldn't possibly be squeaking. After that, I could always laugh at myself during a dissection.

This summer (2007), I was back in the same lab, working with a different grad student (Lynette). I didn't have much trouble re-acclimating to the lab, the preps, and the mice dissection. The deep breathing, the numbing, and the humor -- all that kept working, thankfully.

I watched another student do a perfusion, which involves pumping out all a mouse's blood and replacing it with saline solution, then pumping that out and replacing it with preservative. This turned out to be the same procedure that I saw back when I was with Helen, with the mouse crucified, heart beating, all that. I watched a good part of it. Got a little nauseous, but not really that much. I knew the mouse was unconscious and completely unaware of what was happening to it; everyone takes great care to make sure that the animals are well and completely anaesthetized. Sure, it bothered me, but not overly much; and pretty soon the spinal cord was being extracted, and it was back to the "it's just tissue" stage.

I watched Lynette euthanize four cages of mice. (I participated obliquely, by carrying cages and suchlike; I passed up the offer to participate directly.) We put a cage into a gas chamber and turned on the CO2. The mice gradually slowed down and got quiescent, and eventually they went from asleep to dead. You could tell they were dead when their eyes turned green. After gassing the mice, you had to verify that they were dead. This meant holding the tail with one hand, and pinching the neck with the other, and breaking their necks. I passed up on the offer to try my hand at this, though I had the feeling I probably should have gone for it.

While I watched the euthanasia, I tried not to block it out or push it aside. I wanted to feel horror and revulsion. I wanted to be upset. I made a point of looking at their faces when Lynette said she always tried to avoid it. Things like that. I wanted to make sure I hadn't gone numb.

The next time we had to euthanize mice, we might have had as many as 50, all piled in a cage. Normally, when you're CO2ing them, they just sort of get sleepy, move slower, and then go unconscious. This time, about 3/4 of the way there, a lot of them suddenly moved/twitched at the same time. I don't know if it was coincidental, but it was scary. I participated concretely this time, by breaking the necks of the last six or so. They're so small and soft and fragile, and their necks break so easily. It's easier than breaking a toothpick.

It's not arbitrary. These mice are killed because they are not useful. They don't carry enough copies of the mutation we need. Some of the mothers get in the habit of having litters and then eating them. (By the way, that's a natural mechanism: when the environment is lousy for raising pups, the mother will eat them to conserve protein, protein being not exactly abundant in a mouse's natural diet.) Litters are the wrong age at inconvenient times. One of the females was pregnant, but the pups would be too old by the time anyone could do anything useful with them, just because of everyone's schedules. No one keeps extra mice around for their own sake. It's a hassle to take care of them, and it wastes funding better spent on expensive experimental apparatus and reagents. I accept all of that.

(I would have offered to adopt them, but I'd asked about that at Helen's lab. For one thing, mice with poorly-understood mutations make dangerous pets. For another, it's a bad idea for mouse researchers to keep pet mice of any kind, in case the pet mice get a disease and the researcher carries it into the mouse colonies at the lab. Apart from all of that, the sheer volume of mice involved would be prohibitive.)

Relatively speaking, I felt OK killing mice for a prep, because they were dying for a reason, and their cells were being put to good use. Even part of them lived on; though we killed the mice, we spent every effort coaxing their glial cells to grow and thrive. They were contributing to science, and might one day contribute to human medicine. But the euthanasia was useless and pointless, and that was what got me. These mice were not being used for anything. They were just extra, and there was no room for keeping extra mice.

Fortunately, there isn't any kind of machoness culture around this in the lab. You don't get ridiculed for being upset about the animals suffering and dying. At most, you get a few odd looks, and a suddenly solicitous mentor. We spent half an hour discussing how badly we felt for the people in another lab who had to kill a monkey. They had known this monkey, had worked with it for months, had trained it to do things, had even named it. And now they had to kill it and extract its brain. We were very glad we were not them.

This post is a combination of two LJ posts. Here are the comments to the first and second original posts.


  1. CO2? Why don't they flood the box with Nitrogen instead? Anoxia through O2 starvation is demonstrably painless. Many research pilots go through it, to the point of unconsciousness, and the general comment on recovery is "Did something happen?"
    CO2 overdose triggers the breath reflex, which O2 starvation does not, at least in humans.

  2. I was planning to actually do research into this, and I did do quite a bit of Googling and skimming. But until I have more time (read: January), I'm going to have to punt on this very worthwhile question.
    Honestly, I have no idea why they don't do nitrogen. Pretty much all what I found was protocols: they tell you how to do things, but not why things are done that way. And I don't quite know who to ask about where animal procedures originate.

  3. I hate killing mice for research too. It really eats at me. Fundamentally, life is life regardless of whether it is human or mouse. Then again, the research is designed to ask questions that could lead to significant advancements in our understanding. It's an ethical dilemma for sure... very tough.

    I think CO2 is used more than N2 because CO2 is more dense than air and so it's easier to fill a box with a high concentration.

  4. someday the mice will use us for their experiment..because we have the same antibody etc..


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  6. Did you ever do any research on O2 starvation? Seems like a much more humane way to put animals to sleep without suffering.

  7. Did you ever do any research on O2 starvation? Seems like a much more humane way to put animals to sleep without suffering.