This week I’ve
realised that, despite it being my birthday tomorrow, I’m going to have to
stifle the usual excitement I feel on such a day, especially my 21st,
behave like a proper adult and focus on the job at hand. Sorry to bore you all
with science again, but this is kind of a big deal I have on my hands. I’ve
already explained that the main focus of our lab is to explore ways to “heal”
damaged nerve cells, and this solo project will focus on the use of some
samples that a collaborator in France has sent to Lindy. Just playing with
hugely expensive compounds sent halfway across the world, no biggie…
In short, I’m testing the potential antioxidant effects
of calixarenes on stressed PC12 cells. That probably doesn’t make very much
sense to anyone (including myself the first time I heard it), so I’ll do my
best at explaining. PC12 is a cell line derived from the rat adrenal medulla
(center of the adrenal gland in the kidney). The purpose of cell lines in
general is to study how body cells would react with the convenience of being
able to culture them in a dish, allowing a much quicker rate of study than if
you were to raise them in animals individually. The special thing about PC12s
is that, as well as being robust and able to take a lot of bashing around/changing
conditioins etc, they allow us to stress them and monitor their recovery with
treatment in the confidence that cells of the nervous system would show the
same improvements in a live animal (in
vivo). Clever, eh?
As for the calixarenes
coming from France, they’re the most interesting part because so little work
has been done on them in this field to date. I say little, I mean maybe 3 or 4
papers worldwide. They’re organic molecules (made primarily from carbon,
hydrogen, nitrogen and oxygen) and, most importantly, their name is derived
from the word “chalice” because they’re shaped like little cups. This is
biologically significant because they can hold small molecules, including drugs
and other therapeutic agents, and deliver them to our traumatised nerve cells.
This could have huge implications in the medical world; we want to start with
the first step, however, and see what happens when they get to cells without
anything inside them and whether this alone has any positive effects.
So, on with the big
plan. This morning, I took my PC12s out of incubation, calculated the volume
required to give 10,000 cells per well and transferred this miniscule amount to
100 wells along with media. Hello armache. Overnight, they’re going to “get
happy and comfortable in their new home” (everyone here talks about cells like
they’re children) and should be ready for the second part of the experiment.
Tomorrow, I’ll begin by treating them with a very high dose of glutamate.
Glutamate is a neurotransmitter, a group of compounds that neurons use to
communicate with each other by releasing them from their terminals to be
received by their neighbours. Although the release of neurotransmitters is
essential in coordinating behaviour, exposure to too much is highly toxic to
cells. Some of my stressed cells will then receive calixarene at various
concentrations for thirty minutes and the rest won’t. Lastly, I’ll stain all
the cells with a dye that fluoresces in the presence of any oxidative stress
caused by the glutamate and a plate reader will translate this into numerical
values. Then comes the statistical analysis to see whether adding the
calixarenes actually helps the cells in any way, but I suspect that will be a
very hungover, post-birthday-celebrations Thursday job. It’s all go in
Experimental and Regenerative Neurosciences!
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