Nitric oxide and CaMKII: critical steps in the inotropic response to IGF-1

BURGOS JI; YEVES AM; ENNIS IL; VILA-PETROFF MG

Buenos Aires

Congreso; International Society for Heart Research XXII World Congress (ISHR); 2016

International Society for Heart Research

Cardiac adaptation to aerobic exercise training includes improved cardiomyocyte contractility, by a non-yet clarified mechanism in which nitric oxide (NO) and CaMKII have been implicated. At the cellular level, IGF-1 is the main mediator of the adaptive response to exercise. Our purpose was to explore the effect of IGF-1 on mice cardiomyocyte contractility and the underlying signaling pathway. IGF-1 (10nmol/L) increased cardiomyocyte shortening (128.12±4.62%, n=8 vs basal; p˂0.05), effect abrogated by inhibition of NO production with the non-selective nitric oxide synthase inhibitor L-NAME (2.5 mmol/L; 103.2±3.02%, n=5) or nitroguanidine (NG, 240 nmol/L), specific inhibitor for the neuronal isoform (nNOS, 97.4±1.21%, n=5) and by CaMKII inhibition with KN93 (101.50±2.04%, n=6). In agreement, a significant increase in NO production in response to IGF-1 (133.75±2.17%, n=16) was detected by epifluorescence with DAF-FM. Again, this was prevented by L-NAME (110.36±3.20%, n=11) and NG (114.44±1.83%, n=9), confirming the involvement of nNOS but not altered by KN93 (135.22±1.36%, n=9) suggesting that CaMKII activation was downstream NO production. We explored the pathway involved in nNOS activation by measuring AKT phosphorylation. As expected, IGF-1 increased P-AKT (185.90±10.18%, n=3; p˂0.05). Since NO-dependent CaMKII activation has been proposed, we next determined CaMKII activity (P-CaMKII) and the phosphorylation of its downstream target Thr17-phospholamban, detecting a significant increase in both in the presence of IGF-1 (227.19±29.43% and 143.34±5.44%, n=3 respectively) but not when NO production was prevented by NG (126.61±5.48 and 65.76±15.04, n=3 respectively). Interestingly, similar results showing nNOS and CaMKII activation were obtained in the hypertrophied myocardium of mice subjected to swimming training. In conclusion, our results support a critical role of CaMKII in the positive inotropic effect of IGF-1. Our findings suggest that IGF-1 through the IGF-1R triggers the phosphorylation of AKT which in turn activates nNOS and increases NO production which would be responsible for CaMKII activation.