Open AccessImaging Mitochondrial Ca<sup>2+</sup> Uptake in Astrocytes and Neurons using Genetically Encoded Ca<sup>2+</sup> Indicators (GECIs)
Author(s) -
Nannan Zhang,
Zhe Zhang,
Ilker Ozden,
Shinghua Ding
Publication year - 2022
Publication title -
journal of visualized experiments
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.596
H-Index - 91
ISSN - 1940-087X
DOI - 10.3791/62917
Subject(s) - mitochondrion , astrocyte , neurodegeneration , microbiology and biotechnology , biology , cytosol , mitochondrial dna , neuron , neuroglia , neuroscience , biochemistry , central nervous system , pathology , gene , medicine , disease , enzyme
Mitochondrial Ca 2+ plays a critical role in controlling cytosolic Ca 2+ buffering, energy metabolism, and cellular signal transduction. Overloading of mitochondrial Ca 2+ contributes to various pathological conditions, including neurodegeneration and apoptotic cell death in neurological diseases. Here we present a cell-type specific and mitochondria targeting molecular approach for mitochondrial Ca 2+ imaging in astrocytes and neurons in vitro and in vivo. We constructed DNA plasmids encoding mitochondria-targeting genetically encoded Ca 2+ indicators (GECIs) GCaMP5G or GCaMP6s (GCaMP5G/6s) with astrocyte- and neuron-specific promoters gfaABC1D and CaMKII and mitochondria-targeting sequence (mito-). For in vitro mitochondrial Ca 2+ imaging, the plasmids were transfected in cultured astrocytes and neurons to express GCaMP5G/6s. For in vivo mitochondrial Ca 2+ imaging, adeno-associated viral vectors (AAVs) were prepared and injected into the mouse brains to express GCaMP5G/6s in mitochondria in astrocytes and neurons. Our approach provides a useful means to image mitochondrial Ca 2+ dynamics in astrocytes and neurons to study the relationship between cytosolic and mitochondrial Ca 2+ signaling, as well as astrocyte-neuron interactions.