Simultaneous Study And Measurement Of Nuclear Ca～(2+) And Cytosolic Ca～(2+) With New Calcium Fluorescent Ca～(2+) Probe STDBT

Calcium acts as an intracellular second messenger and plays a very important role in biological systems. The concentration change and spatio-temporal distribution of intracellular Ca²⁺ contribute to its messenger function, and the concentration change of intracellular Ca²⁺ can produce and stop calcium signal. Consequently, it is very important to measure subcellular Ca²⁺ concentration and spatio-temporal distribution for understanding calcium signal. At present, most measurements of intracellular Ca²⁺ are accomplished using probes and fluorescent imaging, and probes play a pivotal role. If probes cannot differentiate and load special subcellular region, fluorescent imaging cannot attain satisfied calcium information of the subcellular region. So far there aren't the indicators which are able to target to nucleus and cytosol simultaneously. People used to target [Ca²⁺]_n and [Ca²⁺]_c with recombinant probes by transfecting with complementary DNAs encoding chimaeras bearing nuclear and cytosolic localization signals. But the recombinant probes must be calibrated. However, the cell environments will influence the calibration results. So people always get many controversial results.In this thesis, the investigations include the following several aspects: 1.Confocal microfluorescence and the new fluorescent Ca²⁺ probe STDBT-AM were used to investigate the mechanism of 5-hydroxytryptamino(5-HT) on nuclear calcium and cytosolic calcium dynamics in rat cardiac muscle cells. 2. Study of the regulation of nuclear and cytosolic Ca²⁺ in rat cardiac muscle cells with double targetable probe STDBT-AM. 3. The new fluorescent Ca²⁺ probe STDBT-AM and confocal microscopy were used to discuss Insulin like growth factor-l(IGF-l)-induced cytosolic Ca²⁺ and nuclear Ca²⁺ dynamic mechanisms in cardiac muscle cells.The thesis includes two parts:1. ReviewMethods and investigations for the subcellular (Endoplasmic Reticulum , Mitochondria, Nuclear etc.) measurements were reviewed in detail.2. Simultaneous study of the regulation of nuclear and cytosolic Ca²⁺ with new...