Abstract 1065: Outflow Tract Regurgitation and Arrhythmias Contribute to In Utero Demise in Early P0-Alk3 Knockout Mouse Embryos as Defined in vivo by 40 MHz Ultrasound Biomicroscopy-Doppler
Background. We know little about cardiovascular systems pathophysiology underlying embryonic death in mouse models of abnormal development. To know the function of BMP signaling in cardiac neural crest, we deleted BMP type IA receptor (Bmpr1a) in a neural crest-specific manner using floxed mice for Bmpr1a in combination with P0-Cre transgenic mice. These knockout (P0-Alk3 KO) mice exhibit hypoplasia of the sympathetic ganglia, bloodless yolk sacs, and outflow tract (OFT) and endocardial cushion hypoplasia, and die at ~E12.5. We determined hemodynamics to gain insights into pathophysiology underlying death.
Methods. In isoflurane-anesthetized pregnant mice, we non-invasively imaged 104 E11.5–12.5 embryos in vivo, using 40 MHz ultrasound biomicroscopy (UBM)-Doppler to acquire data on aortic, umbilical, and vitelline blood flow, cardiac function and rhythm, and AV conduction (mechanical PR interval). PCR-genotyped embryos were compared to littermate controls.
Results. Heart rate, acceleration and ejection times, and systolic biventricular function were normal in P0-Alk3 KO. The P0-Alk3 KO hallmark was diastolic aortic retrograde flow from OFT regurgitation, leading to reduced net VTI (p<0.01). P0-Alk3 KO embryos also failed to increase their peak aortic velocities and VTI between E11.5 and 12.5 normally. In serial imaging of a subset of embryos, 4 of 5 embryos with retrograde aortic flow at E11.5 were dead at E12.5. Umbilical vein pulsatility was normal, suggesting normal placental vascular development and diastolic cardiac function; PECAM staining confirmed normal vascular development in both placenta and embryo. Arrhythmia was noted in 15% of E11.5 P0-Alk3 KO embryos (none in controls), with a normal mechanical PR interval. Vitelline blood flow to the yolk sac was normal in P0-Alk3 KO.
Conclusions. Severe OFT regurgitation and arrhythmia contribute to cardiovascular insufficiency and acute death between E11.5 and 12.5 in P0-Alk3 KO. The presence of OFT regurgitation early, before true semilunar valve formation at E13.5, attests to the dynamic valve-like role of the endocardial cushions, never before shown in vivo. Non-invasive serial UBM-Doppler imaging can elucidate functional consequences of gene targeting in in vivo mouse embryos.