Development and optimization of an in vivo electrocardiogram recording method and analysis program for adult zebrafish.

Clinically pertinent electrocardiogram (ECG) data from model systems, such as zebrafish, are crucial for illuminating factors contributing to human cardiac electrophysiological abnormalities and disease. Current zebrafish ECG collection strategies have not adequately addressed the consistent acquisition of high-quality traces or sources of phenotypic variation that could obscure data interpretation. Thus, we developed a novel platform to ensure high-quality recording of in vivo subdermal adult zebrafish ECGs and zebrafish ECG reading GUI (zERG), a program to acquire measurements from traces that commercial software cannot examine owing to erroneous peak calling. We evaluate normal ECG trait variation, revealing highly reproducible intervals and wave amplitude variation largely driven by recording artifacts, and identify sex and body size as potential confounders to PR, QRS and QT intervals. With this framework, we characterize the effect of the class I anti-arrhythmic drug flecainide acetate on adults, provide support for the impact of a Long QT syndrome model, and establish power calculations for this and other studies. These results highlight our pipeline as a robust approach to evaluate zebrafish models of human cardiac electrophysiological phenotypes.

ROBO4 variants predispose individuals to bicuspid aortic valve and thoracic aortic aneurysm.

Bicuspid aortic valve (BAV) is a common congenital heart defect (population incidence, 1-2%)1-3 that frequently presents with ascending aortic aneurysm (AscAA)4. BAV/AscAA shows autosomal dominant inheritance with incomplete penetrance and male predominance. Causative gene mutations (for example, NOTCH1, SMAD6) are known for ≤1% of nonsyndromic BAV cases with and without AscAA5-8, impeding mechanistic insight and development of therapeutic strategies. Here, we report the identification of variants in ROBO4 (which encodes a factor known to contribute to endothelial performance) that segregate with disease in two families. Targeted sequencing of ROBO4 showed enrichment for rare variants in BAV/AscAA probands compared with controls. Targeted silencing of ROBO4 or mutant ROBO4 expression in endothelial cell lines results in impaired barrier function and a synthetic repertoire suggestive of endothelial-to-mesenchymal transition. This is consistent with BAV/AscAA-associated findings in patients and in animal models deficient for ROBO4. These data identify a novel endothelial etiology for this common human disease phenotype.

Effects of aging and hormonal status on bimanual motor coordination in the rhesus monkey.

Studies of age-related changes in motor function in nonhuman primates have been based exclusively on unimanual motor tasks. In this study, we examined whether aging affects bimanual motor coordination in the monkey model. In addition, we compared performance of ovariectomized and intact females on the task, to examine whether estrogen deficiency impairs motor function. The task required 29 rhesus monkeys (6-26 years old) to extract a maximum of 15 raisins from a testing hole-board. While the task could most efficiently be performed with two hands, other motor strategies were possible. The number of raisins extracted per minute was measured in each of eight sessions, the first and last of which were videotaped for analysis of motor patterns. The number of raisins retrieved per minute declined significantly with age. All monkeys improved with practice, but aged monkeys improved more slowly than young ones. The proportion of bimanual actions tended to increase between the first and the last sessions but was not significantly different between young and aged monkeys. Hormonal status did not affect performance. Finally, performance on the bimanual task was significantly correlated with performance on a previously administered unimanual motor test emphasizing speed, suggesting that age-related motor slowing may explain deficits in both tasks.