The AM ester will be cleaved and allow the dye to stay inside the cell even after washing the cells. However, if your cells highly express multi-drug exporters, the dye will be extruded over time and the signal will become weak.

Dear Jordan, thanks for your answer.

Do you know if cell cutures can resist alive after puting the dye? Is it possible to register the same culture over time?

Depends on what you mean by "over time."

You can maintain a calcium indicator in cells for many hours and continuously observe them. However, there are many factors which play into the toxicity of a dye, including the fact that you will need to use blue light to image the dye, which will cause phototoxicity in your cells. So, you need to only collect data over the meaningful period and frequency for your research question.

For example, if you care about changes that take place over minutes or tens of minutes, then don't image every 5 seconds.

I want to record for periods no longer than five minutes, but at 50, 70 and 100 days of culture, because the networks changes its activity during that time. Could be possible to record the same culture?

I think its extremely unlikely to expect stain to stick around for more than a couple of days.