Time-dependent Modulation of Cardiomyocyte Contractility by Alamandine

Alamandine (ALA), a cardioprotective heptapeptide, has recently been described as a modulator of cardiomyocyte (CM) contractility. Whether this modulation can be influenced by temporal variations remains to be seen. Our aim in this project was to determine whether the effect of ALA on CM contractility is affected by the light-dark cycle and, if so, which signaling pathway would be responsible for this event. CMs from 10-14-week-old male mice were isolated in zeitgebers (ZTs) 02 and 08 (light phase) and ZTs 14 and 20 (dark phase) and contractile function was evaluated. Once the EC 50 of ALA was defined, experiments with antagonists and inhibitors of nitric oxide (NO) production were carried out. The concentration-response curve showed that in ZT02 ALA promoted an increase in the CM fractional shortening (FS) (CTR: 5,4±0,4%, ALA: EC 50 : 134nM; Emax at 10µM: 9,7±0,6%), while in ZT14 ALA promoted a reduction in FS (~40%-EC 50 =undefined). At ZTs 08 and 20, ALA had no significant effect on the contractile function of CMs. We have thus shown for the first time that ALA regulates CM contractility in a time-dependent manner, promoting both an increase and a decrease in opposite ZTs during the 24h. Given that ZTs 02 and 14 were the points where ALA significantly altered the FS of CMs, the next experiments were carried out in these phases of the cycle. Interestingly, at ZT02 the pro-contractile effect of ALA was abolished by the alamandine receptor antagonist (MrgD), D-Pro 7 (1μmol/L), as well as by A779 (1μmol/L) and Losartan (1μmol/L), classic Mas and AT1 receptor antagonists, respectively. These data illustrate the complexity of the regulation of contractile function by ALA in the light phase. Conversely, at ZT14, only D-Pro7 (1μmol/L) was able to antagonize the reduction in FS induced by ALA (CTR: 8,5±0,5%; ALA: 5,2±0,3%; ALA+D-Pro 7 : 7,3±0,5%), while A779 and Losartan had no effect, suggesting that in this phase of the cycle the effect of ALA on contractility is mediated by MrgD. This effect contrasts with that observed in the light phase. These data highlight the importance of understanding the effect of time of day on the mechanisms involved in the modulation of contraction by ALA. Our findings could lead to future studies aimed at developing new drugs and therapies that take into account the influence of the circadian cycle on the efficiency of heart disease therapy. INCT Nanobiofarmacêutica, FAPEMIG, CAPES, and CNPq This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.