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Fat-free crickets

Today, after I recovered from locking myself out of the house (sooo grateful to the apartment manager for a speedy response), I set about gearing up to extract the lipids out of a test set of radioactive crickets.

The gearing up involved, in part, cleaning out a horrifying fume hood. Let me just say that I eventually sprayed some 409 in there, and that caused some sort of exothermic reaction that I wiped up with paper towels. Scary. There are some incredibly poisonous substances in that particular fume hood. *reflexively changes gloves and washes hands again* Eventually I put down some fresh bench paper and now I have some decent workspace!

The lipid extraction methods here differ from our simplistic methods in Texas. In Texas, we dried the crickets and more-or-less homogenized them with a test tube pestle, then weighed out a portion of the homogenate into some filter paper, scrunched the filter paper closed, and soaked it in 3 changes of chloroform, which is really good at dissolving lipids (good reason to never get it on your skin). Sort of a cricket fat-chloroform tea. Then the remainder was re-weighed, and by the power of subtraction, we'd know how much lipid was in the cricket. Easy peasy.

Now, we're addressing a different, but related set of questions. We want to know, more specifically, how different classes of nutrients are allocated to different metabolic end-products in different nutritional environments (sorry, I know that's a lot of differences, all at once). In particular, we'll be following the fate of glycine. Our major interest is in differences in triglyceride vs. phospholipid synthesis, although that's not the entire story. First I should note that insects don't fuel their high-energy activities in the same way that humans do, with glucose. Instead, insects fuel things like flight with triglycerides. Phospholipid synthesis, in contrast, primarily happens for the sake of producing eggs. In a standardized nutritional environment, the flight-capable long-winged female crickets preferentially divert more nutrients towards triglyceride production, at the expense of phospholipid production. In contrast, the flightless short-winged females have no need for extra triglyceride, so they divert more nutrients towards phospholipid production, and start reproducing sooner during adulthood. This is a now-classic example of a clear and direct life-history trade-off, as a given cricket can only do one thing or the other thing, and cannot maximize both. But now we want to know how this story is modulated by the nutritional environment.

Glycine is an amino acid, so the story is a bit more complicated than phospholipds vs. triglycerides. It can wind up in multiple different places. It can be used as an energy source and respired as carbon dioxide, it can be maintained as protein and diverted to various different places in the body (our interest being ovaries vs. somatic tissues), or its carbon can be converted into either of the two lipid classes described above and stored in either somatic or reproductive tissues. So, to trace the fate of glycine, overall I will need to capture respired carbon dioxide (using filter paper treated with a sodium hydroxide solution), dissect and separate somatic and ovarian tissues, separate out the different lipid classes in somatic and ovarian tissues, and measure the remaining somatic and ovarian protein. On both cricket types, across an array of 13 diets that each contain different total amounts and ratios of protein and carbohydrate.

To put it mildly, I'm going to be busy.

But to begin with, last week I practiced injecting radiolabeled glucose into crickets (labeled with carbon-14), and now I'm practicing lipid extractions on those crickets. Handling chloroform-methanol solutions is somewhat tricky, so I still have a few things to learn, but it's fun to be back in the lab for a bit.


( 3 remarks — Remark )
Mar. 6th, 2015 02:52 am (UTC)
>gearing up to extract the lipids out of a test set of radioactive crickets.

This is simultaneously awesome and a completely horrifying sentence.
Mar. 6th, 2015 12:36 pm (UTC)

I know, right? :-) Maybe I will figure out how to turn this stuff into a superhero movie that I can use to fund my research program.

One thing I'm noticing is that this project is making me science-nerd excited in a way that I haven't been since probably my PhD days. Dunno if it's because biochemistry is awesome or the change of working environment (probably both), but I'm grateful to have the feeling return.

Mar. 7th, 2015 02:57 am (UTC)
I think you also have some spare brain cells left over now that aren't trying to deal every day with the very real and oppressive weight of living in TX.
( 3 remarks — Remark )

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