Long-term treatment with ivabradine in post-myocardial infarcted rats counteracts f-channel overexpression

Abstract

Background and purpose: Recent clinical data suggest beneficial effects of ivabradine, a specific heart rate (HR)-lowering drug, in heart failure patients. However, the mechanisms responsible for these effects have not been completely clarified. Thus, we investigated functional/molecular changes in I(f), the specific target of ivabradine, in the failing atrial and ventricular myocytes where this current is up-regulated as a consequence of maladaptive remodelling.

Experimental approach: We investigated the effects of ivabradine (IVA; 10 mg·kg(-1) ·day(-1) for 90 days) on electrophysiological remodelling in left atrial (LA), left ventricular (LV) and right ventricular (RV) myocytes from post-mycardial infarcted (MI) rats, with sham-operated (sham or sham + IVA) rats as controls. I(f) current was measured by patch-clamp; hyperpolarization-activated cyclic nucleotide-gated (HCN) channel isoforms and microRNA (miRNA-1 and miR-133) expression were evaluated by reverse transcription quantitative PCR.

Key results: Maximal specific conductance of I(f) was increased in MI, versus sham, in LV (P < 0.01) and LA myocytes (P < 0.05). Ivabradine reduced HR in both MI and sham rats (P < 0.05). In MI + IVA, I(f) overexpression was attenuated and HCN4 transcription reduced by 66% and 54% in LV and RV tissue, respectively, versus MI rats (all P < 0.05). miR-1 and miR-133, which modulate post-transcriptional expression of HCN2 and HCN4 genes, were significantly increased in myocytes from MI + IVA.

Conclusion and implication: The beneficial effects of ivabradine may be due to the reversal of electrophysiological cardiac remodelling in post-MI rats by reduction of functional overexpression of HCN channels. This is attributable to transcriptional and post-transcriptional mechanisms.

Figure 1

Individual heart rate measurements at the beginning (HR7) and end (HR83) of treatment with vehicle (left panels) or 10 mg·kg⁻¹·day⁻¹ ivabradine in drinking water (n = 12 sham; n = 12 sham + IVA; n = 15 MI; n = 15 MI + IVA; P < 0.05 or P < 0.01, Student's t-test for paired data). MI, myocardial infarction.

Figure 2

I_f activation curves for left ventricular (LV, A) and left atrial (LA, B) myocytes isolated from post-myocardial infarcted rats receiving ivabradine (10 mg·kg⁻¹· day⁻¹ in drinking water) for 3 months (MI + IVA), post-myocardial infarcted (MI) rats and sham-operated animals receiving vehicle for 3 months. (C) Typical I_f recordings obtained in LV myocytes from sham, MI and MI + IVA rats. Number of animals and cells (in parenthesis) are shown in the figure. **P < 0.01, two-way anova for activation curves (sham, MI and MI + IVA).

Figure 3

Occurrence of I_f in right ventricular (RV) myocytes isolated from post-myocardial infarcted rats receiving ivabradine (10 mg·kg⁻¹·day⁻¹ in drinking water) (MI + IVA), post-myocardial infarcted rats (MI), and sham-operated animals receiving vehicle for 3 months. Each column represents the number of cells; 6 sham, 8 MI and 8 MI + IVA rats were used for experiments. **P < 0.01 MI versus sham, ^†P < 0.05 MI + IVA versus MI, for expression of I_f, χ² test.

Figure 4

Relative expression of HCN2 and HCN4 normalized to GAPDH in left ventricular (LV), left atrial (LA) and right ventricular (RV) tissue samples from post-myocardial infarcted rats receiving ivabradine (10 mg·kg⁻¹·day⁻¹ in drinking water) for 3 months (MI + IVA, n = 5 or 6 samples), post-myocardial infarcted rats (MI, n = 4–7 samples) and sham-operated animals (sham, n = 5–7 samples) receiving vehicle for 3 months. RV data are averages of four to six samples. *P < 0.05, **P < 0.01 versus sham rats; ^†P < 0.05 versus MI rats. MI, myocardial infarction.

Figure 5

Relative expression of miR-1 and miR-133a/b in left ventricular (A) and atrial (B) tissue from post-myocardial infarcted rats (MI), and post-myocardial infarcted rats treated with ivabradine (MI + IVA), and sham-operated animals (Sham) by TaqMan-qRT-PCR. Each column represents the mean of six different samples in triplicate ± SEM. *P < 0.05, **P < 0.01 versus sham and MI rats. MI, myocardial infarction; LA, left atrial tissue; LV, left ventricular tissue.