2016
DOI: 10.1152/ajpheart.00175.2016
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R-CEPIA1er as a new tool to directly measure sarcoplasmic reticulum [Ca] in ventricular myocytes

Abstract: In cardiomyocytes, [Ca] within the sarcoplasmic reticulum (SR; [Ca]SR) partially determines the amplitude of cytosolic Ca transient that, in turn, governs myocardial contraction. Therefore, it is critical to understand the molecular mechanisms that regulate [Ca]SR handling. Until recently, the best approach available to directly measure [Ca]SR was to use low-affinity Ca indicators (e.g., Fluo-5N). However, this approach presents several limitations, including nonspecific cellular localization, dye extrusion, a… Show more

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Cited by 23 publications
(23 citation statements)
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“…In this study, only indirect and global ROS measurements were used although development of local ROS measurements could provide further insights. For local [Ca 2+ ], targeted probes are available but cell culture 60–62 complicates the application as culture affects TT ultrastructure.…”
Section: Discussionmentioning
confidence: 99%
“…In this study, only indirect and global ROS measurements were used although development of local ROS measurements could provide further insights. For local [Ca 2+ ], targeted probes are available but cell culture 60–62 complicates the application as culture affects TT ultrastructure.…”
Section: Discussionmentioning
confidence: 99%
“…Once caffeine was washed out, the RyR inhibitor ruthenium red (10 M) was applied to stop Ca 2ϩ release and allow measurement of the rate of luminal [Ca 2ϩ ] ER recovery. At the end of each experiment, the R-CEPIA1er signal was calibrated by addition of the Ca 2ϩ ionophore ionomycin (Iono) (21). ER Ca 2ϩ recovery, monitored by [Ca 2ϩ ] ER accumulation, was analyzed to determine the maximum ER Ca 2ϩ uptake rate (i.e.…”
Section: Structural and Cellular Determinants Of Serca Conformationmentioning
confidence: 99%
“…where F is the R-CEPIA1er fluorescence; F max and F min are the fluorescence level at 10 mM Ca 2ϩ /Iono before and after depletion of ER Ca 2ϩ with caffeine (10 mM), respectively. The R-CEPIA1er Ca 2ϩ dissociation constant (K d ) is 390 M based on in situ calibrations (21). At the end of each experiment, the R-CEPIA1er signal (F max ) was calibrated with addition of Iono (100 M).…”
Section: Measuring Er Ca 2؉ Uptake In Permeabilized Hek-293 Cellsmentioning
confidence: 99%
“…[Ca 2+ ]ER measurements. [Ca 2+ ]ER was recorded as changes in fluorescence intensity of the genetically encoded ER-targeted Ca 2+ sensor R-CEPIA1er (29). R-CEPIA1er was excited with a 514 nm line of the argon laser and signal was collected at >560 nm.…”
Section: In-cell Calcium Uptake and Spontaneous Calcium Release Methodsmentioning
confidence: 99%