title: Differential mitochondrial calcium responses in different cell types detected with a mitochondrial calcium fluorescent indicator, mito-GCaMP2.
abstract: Mitochondrial calcium plays a crucial role in mitochondrial metabolism, cell calcium handling, and cell death. However, some mechanisms concerning mitochondrial calcium regulation are still unknown, especially how mitochondrial calcium couples with cytosolic calcium. In this work, we constructed a novel mitochondrial calcium fluorescent indicator (mito-GCaMP2) by genetic manipulation. Mito-GCaMP2 was imported into mitochondria with high efficiency and the fluorescent signals co-localized with that of tetramethyl rhodamine methyl ester, a mitochondrial membrane potential indicator. The mitochondrial inhibitors specifically decreased the signals of mito-GCaMP2. The apparent K(d) of mito-GCaMP2 was 195.0 nmol/L at pH 8.0 in adult rat cardiomyocytes. Furthermore, we observed that mito-GCaMP2 preferred the alkaline pH surrounding of mitochondria. In HeLa cells, we found that mitochondrial calcium ([Ca(2+)](mito)) responded to the changes of cytosolic calcium ([Ca(2+)](cyto)) induced by histamine or thapasigargin. Moreover, external Ca(2+) (100 μmol/L) directly induced an increase of [Ca(2+)](mito) in permeabilized HeLa cells. However, in rat cardiomyocytes [Ca(2+)](mito) did not respond to cytosolic calcium transients stimulated by electric pacing or caffeine. In permeabilized cardiomyocytes, 600 nmol/L free Ca(2+) repeatedly increased the fluorescent signals of mito-GCaMP2, which excluded the possibility that mito-GCaMP2 lost its function in cardiomyocytes mitochondria. These results showed that the response of mitochondrial calcium is diverse in different cell lineages and suggested that mitochondria in cardiomyocytes may have a special defense mechanism to control calcium flux.
authors: Yunnan Center for Disease Prevention and Control, Kunming, China. chenminyx@gmail.com
authors: Min
authors: M
authors: Chen
authors: Yanru
authors: Y
authors: Wang
authors: Tingting
authors: T
authors: Hou
authors: Huiliang
authors: H
authors: Zhang
authors: Aijuan
authors: A
authors: Qu
authors: Xianhua
authors: X
authors: Wang
mesh_terms: D000328:Adult; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D006367:HeLa Cells / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D006863:Hydrogen-Ion Concentration; D053078:Membrane Potential, Mitochondrial; D008856:Microscopy, Fluorescence / Q000379:methods*; D008928:Mitochondria / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure*; D051381:Rats
chemical_list: D002118:Calcium