MU-induced recovery of depressed L-type Ca²⁺ current (A) and t-tubule density (B).

<p>Maintenance of L-type Ca²⁺ current recovery by combined MetCl therapy, and antagonism of such recovery by Cl or Met mono-therapy (A) is shown, along with antagonism of t-tubule density recovery by all treatments (B). *P<0.05, **P<0.01 and ***P<0.001. Representative di-8-Anepps stained cells from sham (C), HF (D), MUHF (E), MUHF+Cl (F), MUHF+Met (G) and MUHF+MetCl (H) groups are shown. Data for the Met group has been previously published <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092909#pone.0092909-Navaratnarajah1" target="_blank">[22]</a> and added here for comparison.</p

Full recovery of HF-induced depression of L-type Ca²⁺ current (A) and t-tubule density (B) caused by Cl, and lack of improvement in these parameters following Met and MetCl therapy are shown.

<p>***P<0.001. Representative di-8-Anepps stained cells from sham (C), HF (D), HF+Cl (E), HF+Met (F) and HF+MetCl (G) groups. Data for the Met group has been previously published <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092909#pone.0092909-Navaratnarajah1" target="_blank">[22]</a> and added here for comparison.</p

Effect of Cl, Met and combined MetCl treatment on heart weight (HW) (A) and cardiomyocyte volume measured using confocal microscopy (B) during unloading (MUHF).

<p>Prevention of MU-induced cardiac and cardiomyocyte atrophy is achieved by Met and not Cl, with combined MetCl therapy increasing atrophy. *P<0.05, **P<0.01 and ***P<0.001, (HW and cardiomyocyte volume data acquired from 8 and 4 hearts per group, respectively). Effect of Cl, Met and combined MetCl therapy on heart weight∶body weight ratio (HW∶BW) (C) and cardiomyocyte volume (D). HF-induced cardiac hypertrophy was enhanced by Cl therapy but this effect disappeared during combined MetCl therapy. HF-induced myocyte hypertrophy was partially attenuated by Met, but this effect was lost during combination MetCl therapy (HW and cardiomyocyte volume data acquired from 8 and 4 hearts per group, respectively).</p

Effects of Cl, Met and MetCl therapy on cardiomyocyte contractility (A–C) and Ca²⁺ handling (D–F) from non-transplanted failing hearts, measured using Indo-1 and Ionoptix system are shown.

<p>Full recovery of speed of sarcomeric contraction (B), relaxation (C), Ca²⁺ transient amplitude (D) and speed of Ca²⁺ release (E) caused by Met therapy, either alone or in combination with Cl, and lack of improvement following Cl mono-therapy is shown. Superiority of Met mono-therapy in recovering depressed SR Ca²⁺ content is also shown (F). *P<0.05, **P<0.01 and ***P<0.001.</p

Effect of Cl, Met and combined MetCl therapy on contractile function in non-transplanted failing hearts, EF (A) and FS (B): No difference in baseline values in treatment groups was seen (light grey bars).

<p>Average values at end of treatment period are shown (black bars): Cl-treated group showed improved EF and FS compared to untreated HF group, and improvement in EF was further enhanced in MetCl group. Met-induced improvement in EF and FS was not statistically significant. ***P<0.001 (n = 8 per group).</p

Anatomical and cell volume data (16 weeks after LCA ligation).

<p>**P<0.01,</p><p>***P<0.001 vs. sham.</p><p>Anatomical and cell volume data (16 weeks after LCA ligation).</p

Effects of Cl, Met and MetCl therapy on cardiomyocyte contractility (A–C) and Ca²⁺ handling (D–F) during mechanical unloading (MUHF), measured using Indo-1 and Ionoptix system.

<p>Cl's enhancement of MU-induced recovery of speed of sarcomeric contraction (B) and Met's antagonism of MU-induced improvement in speed of relaxation (C) are shown. Cl and Met's enhancement of MU-induced recovery of Ca²⁺ transient amplitude (D), speed of Ca²⁺ release (E) and SR Ca²⁺ content (F), and lack of enhancement during combined MetCl therapy is shown. *P<0.05, **P<0.01 and ***P<0.001. Representative traces of sarcomeric contractions (G) and Ca²⁺ transients (H) are shown.</p

Average HR of mechanically unloaded failing hearts (MUHF) (A) and non-transplanted failing hearts (B) measured using telemetry devices (n = 4 per group).

<p>Cl increased HR significantly at all time-points during MU, whereas Met caused equal HR reduction either alone, or in combination with Cl at all time-points during MU. ***P<0.001 vs. MUHF and ^&&&P<0.001 vs. MUHF+Cl. Cl increased HR in-non-transplanted failing hearts significantly but this effect was lost at week 4. Met caused equal HR reduction in-non-transplanted failing hearts either alone, or in combination with Cl, at all time-points. ***P<0.001 vs. HF and ^&&&P<0.001 vs. HF+Cl.</p