Doppler Estimation of Reduced Coronary Flow Reserve in Mice with Pressure Overload Cardiac Hypertrophy

Abstract

Aortic banding produces pressure overload cardiac hypertrophy in mice, leading to decompensated heart failure in four to eight weeks, but the effects on coronary blood flow velocity and reserve are unknown. To determine whether coronary flow reserve (CFR) was reduced, we used noninvasive 20-MHz Doppler ultrasound to measure left main coronary flow velocity at baseline (B) and at hyperemia (H) induced by low (1%) and high (2.5%) concentrations of isoflurane gas anesthesia. Ten mice were studied before (Pre) and at 1 d, 7 d, 14 d and 21 d after constricting the aortic arch to 0.4 mm diameter distal to the innominate artery. We also measured cardiac inflow and outflow velocities at the mitral and aortic valves and velocity at the jet distal to the aortic constriction. The pressure drop as estimated by 4V² at the jet was 51 ± 5.1 (mean ± SE) mm Hg at 1 d, increasing progressively to 74 ± 5.2 mm Hg at 21 d. Aortic and mitral blood velocities were not significantly different after banding (p = NS), but CFR, as estimated by H/B, dropped progressively from 3.2 ± 0.3 before banding to 2.2 ± 0.4, 1.7 ± 0.3, 1.4 ± 0.2 and 1.1 ± 0.1 at 1 d, 7 d, 14 d and 21 d, respectively (all p < 0.01 vs. Pre). There was also a significant and progressive increase the systolic/diastolic velocity ratio (0.17 Pre to 0.92 at 21 d, all p < 0.01 vs. Pre) suggesting a redistribution of perfusion from subendocardium to subepicardium. We show for the first time that CFR, as estimated by the hyperemic response to isoflurane and measured by Doppler ultrasound, can be measured serially in mice and conclude that CFR is virtually eliminated in banded mice after 21 d of remodeling and hypertrophy. These results demonstrate that CFR is reduced in mice as in humans with cardiac disease but before the onset of decompensated heart failure. (E-mail: [email protected])

Introduction

Coronary flow is often normal at rest, even in the presence of significant coronary lesions, but maximum flow during exercise or stress can be decreased (Gould et al. 1974). Coronary flow reserve (CFR) is defined as the ratio of maximal-hyperemic/baseline-resting flow or velocity and is often used as an index of the functional severity of coronary stenoses seen angiographically (Cole and Hartley 1977, White et al 1984). Coronary flow reserve is also reduced in the presence of valvular and other forms of heart and vascular diseases, which increase loading conditions and produce cardiac hypertrophy (Fallen et al 1967, Marcus et al 1982, Marcus 1983, Santagata et al 2005, Neishi et al 2005, Parrish et al 1985). The mechanism for this reduction is thought to be caused by increases in resting baseline coronary flow as a result of increased cardiac work (Marcus et al 1982, Eberli et al 1989, Parrish et al 1985, Bache et al 1987). Because oxygen delivery and cardiac work are closely related, the magnitude of CFR is similar to that of cardiac reserve (the ratio of maximal/baseline cardiac output) in normal individuals, and both CFR and exercise capacity tend to be reduced by similar amounts in the presence of cardiovascular disease (Rushmer 1976). It is thought that decompensated heart failure ensues only after cardiac and coronary reserves are exhausted (Rushmer 1976, Fallen et al 1967, Vatner and Hittinger 1993, Hittinger et al 1989). Now that mice are being used as cardiovascular disease models (Niebauer et al 1999, Rockman et al 1993, Brickson et al 2006, Barrick et al 2007, Tanaka et al 1996, Maslov et al 2007), it is important to determine if the changes in myocardial perfusion and reserve in the face of increased loading conditions are similar in mice, humans and larger mammals.

We have recently developed a noninvasive method to measure coronary flow velocity in mice using Doppler ultrasound and have shown that isoflurane gas can be used as a convenient coronary vasodilator when the concentration is increased from 1 to 2.5% (Hartley et al. 2007). We used the ratio of hyperemic/baseline (H/B) left main coronary velocity as an index of CFR, and showed that H/B was a function of age and was reduced in 2-year-old apolipoprotein-E null (ApoE^-/-) mice when compared with age-matched controls as shown in Fig. 1. The increased flow velocity at baseline and during hyperemia in the ApoE^-/- mice is consistent with the presence of coronary lesions in some of the mice. Others have shown that cardiac functional reserve as measured by exercise capacity is reduced in ApoE^-/- mice (Niebauer et al. 1999), and we have shown that ApoE^-/- mice have increased aortic and mitral flow velocity and increased heart-weight/body-weight ratio (HW/BW) consistent with volume overload hypertrophy (Hartley et al. 2000). It is unclear whether the observed decrease in H/B in ApoE^-/- mice was a result of the presence of coronary lesions, restricting hyperemic flow or the increased baseline cardiac work, increasing resting flow.

Transverse aortic banding induces pressure overload cardiac hypertrophy in mice and is used to stress normal, old and genetically altered mice to study cardiac function and remodeling (Rockman et al 1993, Li et al 2003, Brickson et al 2006, Barrick et al 2007). Banded mice have significantly elevated HW/BW after one to five weeks, some show signs of heart failure and many die within three to five weeks after banding (Barrick et al. 2007). In dogs with pressure overload cardiac hypertrophy, it has been shown that CFR is reduced and that the subendocardium becomes more seriously underperfused and dysfunctional (Hittinger et al 1989, Vatner and Hittinger 1993, Duncker et al 1998, Hittinger et al 1995, Bache et al 1987). We hypothesized that mice would have lower CFR as estimated by H/B after banding, but before the development of symptomatic heart failure. We report here results from 10 mice showing that H/B is progressively decreased and essentially eliminated 21 d after transverse aortic banding. We also observed a significant and progressive increase in the systolic component of coronary flow following aortic banding, suggesting a redistribution of flow away from the subendocardium.