How do you measure calcium levels in mitochondria?
By permeabilizing the cells, mitochondrial Ca2+ can be measured using the fluorescent Ca2+ indicator Fluo-4, AM, with measurement of ΔΨm using the fluorescent dye tetramethylrhodamine, methyl ester, perchlorate (TMRM).
What are genetically encoded calcium indicators?
The genetically encoded calcium indicator (GECI), which binds to calcium ions and emits fluorescence visualizing intracellular calcium concentration, enables detection of in vivo neuronal firing activity. Various GECIs have been developed and can be chosen for diverse purposes.
Why is calcium important in mitochondria?
Calcium is thought to play an important role in regulating mitochondrial function. Evidence suggests that an increase in mitochondrial calcium can augment ATP production by altering the activity of calcium-sensitive mitochondrial matrix enzymes.
Where is calcium stored in mitochondria?
A mitochondrion from outside to inside can be divided into four functional areas: the outer mitochondrial membrane (OMM), the mitochondrial intermembrane space, the inner mitochondrial membrane (IMM) and the mitochondrial matrix. Calcium ions mainly distribute in the intermembrane space and the matrix.
Why are calcium ions important for signaling?
Calcium signaling is the use of calcium ions (Ca2+) to communicate and drive intracellular processes often as a step in signal transduction. Ca2+ is important for cellular signalling, for once it enters the cytosol of the cytoplasm it exerts allosteric regulatory effects on many enzymes and proteins.
What is ratiometric imaging?
Ratiometric fluorescence is the method where intensities at two or more wavelengths of an excitation or emission spectrum are measured to detect changes to local environment. Typically, a probe is used that is specifically sensitive to an environmental parameter such as ion concentration, pH, viscosity, or polarity.