I am trying to do some functional assays on primary smooth muscle cells from young mice. When I say young I mean newborn esophagus or 3 week old aortic smooth muscle. There are not many cells for functional assays so I take them into cell culture for a while until they grow confluent, and then I try to stimulate them with an agonist (carbachol) and visualize contraction under the microscope. The problem appears to be that these cells lose their contractility really fast and I can't see any response to agonist, although smooth muscle actin expression is still quite high (so they are still 'smooth muscle cells'). I culture them in DMEM + 10% FBS on plastic. I read that primary SMCs will rexpress contractility makers if starved for 3 days, but this hasn't helped my situation--maybe this trick is human-specific?

Does anyone have any tips on how I can keep these cells contraction-competent in culture?