Effects of SO42- Removal on Calcium Dynamics in the Sarcoplasmic Reticulum: Rhod-2 Analysis
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This study investigates the impact of removing SO42- on the calcium concentration within the sarcoplasmic reticulum ([Ca2+]SR) using Rhod-2 fluorescence. Observations indicate minimal cytoplasmic Ca2+ release during peaks, suggesting limited mitochondrial Ca2+ loading. High [Ca2+]SR areas correspond to transverse terminal cisternae, while longitudinal areas exhibit low calcium concentrations. Substitution of SO42- with glutamate and EGTA reveals shifts in calcium uptake dynamics. The findings highlight the intricate balance of calcium signaling in muscle cells and the effects of ionic substitutions.
Effects of SO42- Removal on Calcium Dynamics in the Sarcoplasmic Reticulum: Rhod-2 Analysis
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[Ca2+]SR Rhod-2 Rhod-2 [Ca2+]SR Solution change Note: no great cytoplasmic release at peak of or rise in [Ca2+]SR. Therefore mitochondria cannot be loading cytoplasmic Ca2+. Also, it is obvious that high [Ca2+]SR areas are clearly transversal (SR terminal cisternae) and longitudinal areas (mitochodria) are low [Ca2+]. 081004a Effect of removing SO42- on SR [Ca2+] (SO42- is substituted for glutamate at the second data point in this figure). 081004a
081004a_s132_4_7_rto.tif 081004a_s133_0_11_rto.tif Avg ratio series 133 #0 to 11 Avg ratio series132 #4 to 7 tv,255<rotate(100.*(rto0-0.4),flagrot)>0 B A Difference B-A, > 0 Difference, A-B, > 0
[Ca2+]SR Rhod-2 [Ca2+]SR Rhod-2 [Ca2+]SR Rhod-2 081004b Effect of removing SO42- on SR [Ca2+] (SO42- is substituted for glutamate at the second data point in this figure). [Ca2+]SR Rhod-2
Lots of mitochondria in mag-indo image—could be decreasing estimate of [Ca2+]. The SR emptied over the course of the next few minutes. Effect of removing SO42- on SR [Ca2+] (SO42- is substituted for glutamate at the second data point in this figure). 102804a
Rhod-2 [Ca2+]SR The fastest rise in [Ca2+]SR is perhaps around 500-600 μM s-1? Rhod-2 Rhod-2 [Ca2+]SR [Ca2+]SR 102804c Effect of removing SO42- on SR [Ca2+] (SO42- is substituted for 50mM EGTA at the second data point in this figure). Rhod-2 [Ca2+]SR
SO42- EGTA Effect of loading in 80 mM SO42-, 1 mM EGTA (100 nM Ca2+) and 0.4 mM Mg2+ for ~16 min, then switching to 50 mM EGTA (< 2 nM Ca2+) and 1 mM Mg2+. Total SR Ca is assayed at the end of the expt with caffeine. 30 mM caffeine caffeine 110904d
solution change 110804c Effect of removing SO42- on SR [Ca2+] (SO42- is substituted for 50mM EGTA at the 18-20 s time point in this figure).
Effect of removing 50 mM SO42- (isosmotically substituted with K+-Glut), 1 mM EGTA (100 nM Ca2+) and 0.4 mM Mg2+ on SR [Ca2+] (SO42- is substituted for 50mM EGTA). 50 mM SO42- EGTA 30 mMcaffeine XT’s here aswell 111604a
Effect of loading in 50 mM Pi, 1 mM EGTA (100 nM Ca2+) and 0.4 mM Mg2+ for ~17 min, then switching to 50 mM EGTA (< 2 nM Ca2+) and 1 mM Mg2+. Note: no sparks 110904a
Effect of loading in 50 mM Pi, 1 mM EGTA (100 nM Ca2+) and 0.4 mM Mg2+ for ~17 min, then switching to 50 mM EGTA (< 2 nM Ca2+) and 1 mM Mg2+. Note: no sparks 111604b Open pinhole
Effect of loading in 50 mM Pi, 1 mM EGTA (200 nM Ca2+) and 1 mM Mg2+ for ~17 min, then switching to 50 mM EGTA (< 2 nM Ca2+) and 1 mM Mg2+. Total SR Ca is assayed at the end of the expt with caffeine. 30 mM caffeine 110904c
Glut EGTA Effect of substituting K-Glut, 1 mM EGTA (100 nM Ca2+) and 0.4 mM Mg2+ [after 15 min loading] for 50 mM EGTA (< 2 nM Ca2+) and 1 mM Mg2+. 111004b
50 μM TBQ SO42-, 100 nM Ca2+, 0.4 mM Mg2+ 50 mM EGTA, 1 mM Mg2+ 111804a
50 μM TBQ SO42-, 100 nM Ca2+, 0.4 mM Mg2+ 50 mM EGTA, 1 mM Mg2+ 111804b
Saturated signal 30 mM caff 111604c
