ABSTRACT
Title
Anatomical and Morphological Characterization Of Murine Left Atrium: A Tool To Model Human Disease
Authors
L. Castiglioni
Dept of Pharmacological Sciences, University of Milan, Milano, Italy.
Dottorato in Scienze Farmacotossicologiche, Farmacognostiche e Biotecnologie Farmacologiche
Dept of Pharmacological Sciences, University of Milan, Milano, Italy.
Dottorato in Scienze Farmacotossicologiche, Farmacognostiche e Biotecnologie Farmacologiche
Abstract
Introduction:Murine models to study human cardiovascular disease are extremely attractive; therefore it is crucial to have detailed knowledge of their heart structures. There is evidence that the left atrium (LA) plays a central role in early detection of cardiac disease and prognosis, so this study aims to describe, for the first time, the anatomical, morphological and functional characteristics of the LA in adult mice with immuno-histological analyses and with the detail allowed by high frame rate echocardiography.
MethodsWe studied 30 C57BL6 mice (male:female = 15:15; mean body weight= 21±3 g) with a Vevo 2100 echocardiography system and a 30 MHz transducer, during anaesthesia with isoflurane 0.8-1%. Standard apical 4-chamber and modified views were used to optimize imaging of the LA, mitral valve, LA appendage (LAA) and pulmonary venous (PV) flows with Doppler echocardiography. After the imaging analysis the mice were anesthetized and abdominal aorta cannulated. The heart, arrested in diastole, was perfused with either a silicone elastomer (n=10) to build a cast for macroscopic inspection, or with 4% formalin (n=20). The latest were embedded in paraffin and sectioned for immuno-histological analyses.
Results:Murine LA in mice is disproportionately smaller with respect to the left ventricle than in humans and exhibits an average transmural wall thickness of 0.21 ± 0.03 mm. The atrium is connected to the LA appendage (LAA), a trabeculated crescent-like structure, by a 0.16-0.4 mmdiameterduct. The major branches of the mouse aorta and pulmonary artery are similar to humans; in contrast the venous system has a different disposition with three pulmonary veins tributary of a common pulmonary vein that drains into the LA. These structures were optimally imaged with echocardiography in 28/30 mice. The microscopic evaluation of myocardium revealed the presence of two thin layers of cardiomyocyte crossing each other. Cells spanning LA are cardiomyocyte (60% ± 6 a-sarcomeric actin+ in red) and cardiac fibroblast (9% ± 2.3 vimentine+) exhibiting a rich vascular supply with a dense capillary network (24% ± 2.3 Isolectin-B4+in green). The presence of alpha-smooth muscle actin (7% ± 4.6) identified the presence of arterioles.With echocardiography system we observed mean heart rate 484±43 bpm, duration of systole 76±9 ms and diastole 49±6 ms. There was a three-fold increase in LA volume (LAV) during systolic filling and both LA and LAA were smaller in females (Table). The transmitral flow pattern was dependent on heart rate: monophasic (atrial systolic wave) in 24 mice and biphasic in 4 mice. The PV inflow was characterized by 3 forward waves, 2 systolic (velocity-time integrals, early: 6±4, late: 5±2 mm) and 1 diastolic (17±5 mm), and the LAA by 2 systolic inflow (early: 9±3, late: 6±2 mm) and 1 diastolic (7±3 mm) outflow waves. As shown in other species, variables related to LA compliance correlated with left ventricular stroke volume (LVSV): the LAA long axis was linearly related to LVSV (r= .6, p= .01) and at regression analysis both the systolic PV inflow and LAA diastolic outflow velocity/time integrals determined LVSV (r= .56, p= .006).
Conclusions: This is the first detailed description of murine LA anatomy, morphology and function, highly feasible with high frame rate echocardiography.
These findings might help elucidating human disease patho-physiology, possibly leading to new therapeutical approaches for heart disease treatment.
Table
MethodsWe studied 30 C57BL6 mice (male:female = 15:15; mean body weight= 21±3 g) with a Vevo 2100 echocardiography system and a 30 MHz transducer, during anaesthesia with isoflurane 0.8-1%. Standard apical 4-chamber and modified views were used to optimize imaging of the LA, mitral valve, LA appendage (LAA) and pulmonary venous (PV) flows with Doppler echocardiography. After the imaging analysis the mice were anesthetized and abdominal aorta cannulated. The heart, arrested in diastole, was perfused with either a silicone elastomer (n=10) to build a cast for macroscopic inspection, or with 4% formalin (n=20). The latest were embedded in paraffin and sectioned for immuno-histological analyses.
Results:Murine LA in mice is disproportionately smaller with respect to the left ventricle than in humans and exhibits an average transmural wall thickness of 0.21 ± 0.03 mm. The atrium is connected to the LA appendage (LAA), a trabeculated crescent-like structure, by a 0.16-0.4 mmdiameterduct. The major branches of the mouse aorta and pulmonary artery are similar to humans; in contrast the venous system has a different disposition with three pulmonary veins tributary of a common pulmonary vein that drains into the LA. These structures were optimally imaged with echocardiography in 28/30 mice. The microscopic evaluation of myocardium revealed the presence of two thin layers of cardiomyocyte crossing each other. Cells spanning LA are cardiomyocyte (60% ± 6 a-sarcomeric actin+ in red) and cardiac fibroblast (9% ± 2.3 vimentine+) exhibiting a rich vascular supply with a dense capillary network (24% ± 2.3 Isolectin-B4+in green). The presence of alpha-smooth muscle actin (7% ± 4.6) identified the presence of arterioles.With echocardiography system we observed mean heart rate 484±43 bpm, duration of systole 76±9 ms and diastole 49±6 ms. There was a three-fold increase in LA volume (LAV) during systolic filling and both LA and LAA were smaller in females (Table). The transmitral flow pattern was dependent on heart rate: monophasic (atrial systolic wave) in 24 mice and biphasic in 4 mice. The PV inflow was characterized by 3 forward waves, 2 systolic (velocity-time integrals, early: 6±4, late: 5±2 mm) and 1 diastolic (17±5 mm), and the LAA by 2 systolic inflow (early: 9±3, late: 6±2 mm) and 1 diastolic (7±3 mm) outflow waves. As shown in other species, variables related to LA compliance correlated with left ventricular stroke volume (LVSV): the LAA long axis was linearly related to LVSV (r= .6, p= .01) and at regression analysis both the systolic PV inflow and LAA diastolic outflow velocity/time integrals determined LVSV (r= .56, p= .006).
Conclusions: This is the first detailed description of murine LA anatomy, morphology and function, highly feasible with high frame rate echocardiography.
These findings might help elucidating human disease patho-physiology, possibly leading to new therapeutical approaches for heart disease treatment.
Table
|
LAV end-systole μL |
LA long axis mm |
LAA long axis mm |
LV long axis mm |
LVSV μL |
|
| Male | 6.3±1.7 | 2.2±0.2 | 4±.3 | 7.3±0.8 | 42±9 |
| Female | 4.7±1.3 | 2±0.2 | 3.4±0.2 | 7.1±0.6 | 37±9 |
| p | 0.007 | 0.02 | <0.001 | 0.1 |