In vivo Ca2+dynamics induced by Ca2+injection in individual rat skeletal muscle fibers

Figures

• 

  Figure 1. Representative example of changes in intracellular Ca2+ concentration ([Ca2+]i) induced by 2.0 mmol/L Ca2+ microinjection into a single rat spinotrapezius muscle myocyte. The injection site is shown with an arrow. The specific region of interest (width: 20 μm, height: 80% of muscle fiber diameter) for the Ca2+ measurement was set to a distance of 40 μm from the injection point. Scale bar = 50 μm. Pseudocolor bar indicates the 340/380‐nm ratio value.

• 

  Figure 2. Influence of the different Ca2+ injections (0.02, 0.2 2.0 mmol/L) on intracellular Ca2+ concentration ([Ca2+]i) in vivo. Fluorescence was measured continuously during the 60 sec measurement interval and ratiometrically quantified data were graphed as changes from precondition (i.e. −10 sec) levels (R0). Values shown are means ± SE (N = 10, 0.02 mmol/L: n = 14, 0.2 mmol/L: n = 17, 2.0 mmol/L: n = 17). N = number of animals. n = number of muscle fibers measured. There was a significant interaction effect for treatment condition (0.02, 0.2 and 2.0 mmol/L) and time course of injection (P < 0.001). Significant difference between each conditions for the same time points: *P < 0.05 (vs. 0.02 mmol/L), # P < 0.05 (vs. 2.0 mmol/L).

• 

  Figure 3. (A) The dose–response curves show the pharmacologic function of DAN (100, 500 μmol/L) and CPA (10, 50, 100 μmol/L) in the in vivo environment. (B) Influence of the DAN, top panel and CPA, bottom panel in vivo on intracellular Ca2+ concentration ([Ca2+]i). Values shown are means ± SE (DAN condition: N = 7, 0.02 mmol/L: n = 14, 0.2 mmol/L: n = 18, 2.0 mmol/L: n = 18, CPA condition: N = 7, 0.02 mmol/L: n = 15, 0.2 mmol/L: n = 15, 2.0 mmol/L: n = 15). N = number of animals. n = number of muscle fibers measured. CPA, cyclopiazonic acid; DAN, dantrolene sodium.

• 

  Figure 4. Influence of caffeine injection with/without DAN in vivo on intracellular Ca2+ concentration ([Ca2+]i). Values shown are means ± SE (caffeine condition: N = 5, n = 9, caffeine/DAN condition: N = 5, n = 8). N = number of animals. n = number of muscle fibers measured. There was a significant interaction effect for treatment condition (caffeine injection with/without DAN) and time course of injection (P < 0.001). Significant difference between each condition at the same time point: *P < 0.05 (caffeine injection with vs. without DAN). DAN, dantrolene sodium.

• 

  Figure 5. (A) Changes of muscle fiber length (i.e., extent of shortening) during each injection protocol (Ca2+, Ca2++DAN, Ca2++CPA, and caffeine). Values shown are means ± SE (Ca2+: n = 12, Ca2++DAN: n = 14 Ca2++CPA: n = 15, caffeine: n = 9). n = number of muscle fibers measured. B, C, and D: Shortening dynamics at 2.0 mmol/L Ca2+ and caffeine injection. (B) Bar graph shows peak shortening length. There was no statistically significant difference among conditions. (C) Bar graph shows peak shorting velocity. The Caffeine condition tended to be slower than 2.0 mmol/L Ca2+ control (# P = 0.060). (D) Bar graph shows mean fiber length shortening during 60 sec following injection. Significance compared with Ca2+ condition, *P < 0.05, **P < 0.01. The Caffeine condition tended to be shorter than the Ca2+ condition (# P = 0.059). (E) Bar graph shows peak shortening length for 0.2 mmol/L and 0.02 mmol/L Ca2+ conditions. Significant changes with the inhibitors were not found in either Ca2+ concentration conditions. CPA, cyclopiazonic acid; DAN, dantrolene sodium.