I am currently working on the effects of cholesterol on heart muscle cells (cardiac myocytes). This week I cultured heart cells that I extracted from experimental mouse models (read: I removed the hearts from several mice, broke down the heart into individual cells, and grew them in petri dishes with a nutritious broth). I'd never seen these kinds of cells under the microscope before and boy, did I get a neat surprise!
As you know, the heart pumps blood by contracting muscles in the wall of the heart. The force of the contraction pushes blood out of the heart and into the arteries to be carried off to our extremities. To review a little biology, remember that the structural hierarchy in the body is as follows:
Organ (heart) ==> Tissue (heart muscle) ==> Cell (cardiac myocyte)
Macroscopic ==> Microscopic
Although we see the entire heart contract as one muscle tissue, the contractions are actually taking place on the cellular level. This means that individual cells contract independently. It is the function of the brain and the nervous system to coordinate thousands of cellular contractions into one big muscle contraction of the whole heart.
When you culture cardiac myocytes in a dish, you can observe the individual contractions. Indeed, long after you have extracted the cells from a mouse, the cells are still beating on their own. Incredible! And it makes experimenting a lot easier...
Here's an example of what I can see under the microscope:
Truly amazing!
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nice video. cardiology is one of my fav subjects so far.
ReplyDeleteHI I too work with cardiac myocytes (neonatal rat), can I ask what broth do you use and how do you mince the heart tissues? thanks for the terrific video.
ReplyDeleteJust noticed the comment.
ReplyDeleteI use different media mixtures:
For neonatal mouse myocytes:
1) Harvest in cold Hanks Balanced Salt Solution (HBSS)
2) I only mince the right and left ventricular tissue (all other xs tissue is removed)
3) I mince into small pieces (isolating the ventricles) using scalpel --and I do this directly in the buffer and wash several times with HBSS. Chop/separate as quicly as possible.
4) sequential digests with a Trypsin/HBSS solution at 37 C. *This is what really breaks it into individual cells
5) re-suspend and pre-plate cells for 1-3 hours in DMEM + 10% fetal bovine serum and 5% horse serum to remove nonmyocytes.
6) Count viable myocytes (trypan blue)
8) seed viable cells onto fibronectin-coated plates
9) overnight in DMEM + 10% fetal bovine serum, 5% horse serum and pen/strep.
In my experiments, I use DMEM + 2% horse serum + 2& fetal calf serum + pen/strep
I only use mouse cells, not rat. Not sure if that makes a difference.
I also isoloate adult mouse cardiomyocytes. Different dilutions for the media...
Oh, and thanks Vidya for coming to the site! Let me know if you have any other questions!
ReplyDeleteYou too Cristin, thanks for the comments! I appreciate your visits!