Measuring Intracellular Ca2+ Levels in Plant Cells Using the Fluorescent Probes, Indo-1 and Fura-2

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Abstract
Chlamydomonas in the resting phase of growth has an equal capacity of about 15 micromole O2 uptake per hour per milligram of chlorophyll for both the cytochrome c, CN-sensitive respiration, and for the alternative, salicylhydroxamic acid-sensitive respiration. Alternative respiration capacity was measured as salicylhydroxamic acid inhibited O2 uptake in the presence of CN, and cytochrome c respiration capacity as CN inhibition of O2 uptake in the presence of salicylhydroxamic acid. Measured total respiration was considerably less than the combined capacities for respiration. During the log phase of growth on high (2-5%) CO2, the alternative respiration capacity decreased about 90% but returned as the culture entered the lag phase. When the alternative oxidase capacity was low, addition of salicylic acid or cyanide induced its reappearance. When cells were grown on low (air-level) CO2, which induced a CO2 concentrating mechanism, the alternative oxidase capacity did not decrease during the growth phase. Attempts to measure in vivo distribution of respiration between the two pathways with either CN or salicylhydroxamic acid alone were inconclusive.