Simultaneous measurement of cytosolic and mitochondrial calcium levels: Observations in TRPC1-silenced hepatocellular carcinoma cells

The measurement of intracellular Ca2+, cytosolic or stored in organelles, i.e., mitochondria, gave valuable data for numerous areas of research. In case of tumor cells, mitochondrial Ca2+ levels play essential roles in apoptosis along with endoplasmic reticulum (ER) Ca2+. In this study, we describe a Ca2+ monitoring system that allows studying both adherent cells and tissues and discuss data obtained from hepatocellular carcinoma cells and rat thoracic aorta by using this system. For this purpose, two apparatus, one for adherent cells and the other for intact rat aorta, were designed and produced. With this system, changes in cytosolic Ca2+ levels following store-operated calcium (SOC) entry induced by sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) blockers were recorded in different hepatocellular carcinoma cells. Furthermore, cytosolic and mitochondrial Ca2+ levels were simultaneously measured in TRPC1-silenced Huh7 hepatocellular carcinoma cells. In addition, the effects of trifluoromethylphenylimidazole (TRIM) on cyclopiazonic acid (CPA)-, serotonin (5-HT)-, and phenylephrine (PE)-induced changes in isometric force and cytosolic Ca2+ levels were determined simultaneously in rat thoracic aorta. The effects of aging on PE-induced responses were also investigated. After SOC entry activation, cytosolic Ca2+ levels were increased, as expected in all hepatocellular carcinoma cells. Mitochondrial Ca2+ levels following CPA-induced ER depletion were significantly (p