Dantrolene improves left ventricular diastolic property in mineralcorticoid-salt-induced hypertensive rats

Figures

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  Fig. 1. Research design, blood pressure, cardiac and lung weights, echocardiograms. (A) Study design of DOCA-salt induced hypertensive rat. (B) Blood pressure of the rats in each group (n = 5–8). (C) Body weight, heart weight and lung weight in each group (n = 4–8). (D) Representative echocardiograms, and the summarized data (n = 6–15). ***P < 0.001 vs UNX control (two-way ANOVA with a post-hoc Bonferroni test) for B. *P < 0.05, **P < 0.01, ***P < 0.001 (one-way ANOVA with a post-hoc Tukey's test). #P < 0.05, ##P < 0.01, ###P < 0.001 (KW with a post-hoc Dunn's test). LV, left ventricular; LVDd, left ventricular diastolic dimension; LVDs, left ventricular systolic dimension; IVS, intra-ventricular septum; PW, posterior wall.

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  Fig. 2. DAN inhibited the development of LV hypertrophy and decreased LV chamber stiffness in UNX + DOCA-salt induced hypertensive rats. (A) Representative images of long-axis sections of HE- or Serius red-stained hearts and the RV tissues. Fibrotic areas of the LV wall obtained from Serius red-stained LV tissue are shown in the lower panel (n = 7–8) (B) Static LV pressure-volume relationship as a parameter of LV chamber stiffness (n = 4–5) *P < 0.05, **P < 0.01, ***P < 0.001 (one-way ANOVA with a post-hoc Tukey's test) for A. *P < 0.05, ***P < 0.001 vs UNX control (two-way ANOVA with a post-hoc Bonferroni test) for B. ††P < 0.05 vs UNX + DOCA-salt (two-way ANOVA with a post-hoc Bonferroni test) for B. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

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  Fig. 3. Morphology, Ca2+ transient and cell shortening in intact cardiomyocytes. (A) Representative images of isolated cardiomyocytes, and the summarized data of cell width, cell length and cell area (n = 89–159 cells from 3 to 4 hearts). (B) Representative recordings of fluo-4 AM fluorescence signal at a pacing rate of 1 Hz, and the summarized data (n = 4–15 cells from 3 to 4 hearts). (C) Representative recordings of sarcomere length at a pacing rate of 2 Hz, and the summarized data (n = 9–15 cells from 3 to 4 hearts). *P < 0.05, **P < 0.01, ***P < 0.001 (one-way ANOVA with a post-hoc Tukey's test).

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  Fig. 4. Ca2+ spark, sarcoplasmic reticulum (SR) Ca2+ content, and spontaneous Ca2+ transient (SCaT) in intact cardiomyocytes. (A) Representative recordings of spontaneous Ca2+ sparks. On the left, recordings of line-scan image of fluo-4 AM fluorescence in cardiomyocytes; on the right, the summarized data (n = 10–15 cells from 3 to 4 hearts). The SR Ca2+ content was measured as follows: caffeine-induced Ca2+ transients were measured by first applying a stimulation train at 2 Hz, and then by rapidly switching from the superfusing solution to a solution containing 20 mM caffeine for 5–6 s. The summarized data is indicated at right lower panel (n = 41–114 cells from 3 to 4 hearts). (B) On the left, representative traces of spontaneous Ca2+ transients (SCaTs); on the right, the summarized data as the incidence of SCaTs (n = 45–84 cells from isolated cardiomyocytes of 3–4 hearts as indicated). Arrows indicate SCaTs. The SCaTs were induced after cessation of pacing at 1, 2, 3, 4, and 5 Hz of stimulation. Values for individual rats are plotted with mean ± standard error of mean. *P < 0.05, **P < 0.01, ***P < 0.001(ANOVA with a post-hoc Tukey's test) for A.

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  Fig. 5. Localization and association with RyR2 of endogenous CaM in cardiomyocytes. (A) Representative images of endogenous CaM, which is co-localized with RyR2 in isolated cardiomyocytes; immunostaining of CaM (shown in green); immunostaining of RyR2 (red). (B) Summarized data of Z line-bound and nuclear CaM. The immunofluorescence signal of Z line-bound CaM was divided by that of RyR2, normalized to control (baseline of Sham), and expressed as ratios. The immunofluorescence signal of nuclear CaM was divided by that of DAPI for nuclear staining, normalized to control (baseline of Sham), and expressed as ratios. Data are presented as mean ± standard error of mean of 3–4 hearts (each plot is the average of 4–5 cells). *P < 0.05, **P < 0.01, ***P < 0.001(ANOVA with a post-hoc Tukey's multiple comparison procedure. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)