Summary Lifespan is a remarkably diverse trait ranging from a few days to several hundred years in nature, but the mechanisms underlying the evolution of lifespan differences remain elusive. Here we de novo assemble a reference genome for the naturally short-lived African turquoise killifish, providing a unique resource for comparative and experimental genomics. The identification of genes under positive selection in this fish reveals potential candidates to explain its compressed lifespan. Several aging genes are under positive selection in this short-lived fish and long-lived species, raising the intriguing possibility that the same gene could underlie evolution of both compressed and extended lifespans. Comparative genomics and linkage analysis identify candidate genes associated with lifespan differences between various turquoise killifish strains. Remarkably, these genes are clustered on the sex chromosome, suggesting that short lifespan might have co-evolved with sex determination. Our study provides insights into the evolutionary forces that shape lifespan in nature.

Summary Aging is a complex process that affects multiple organs. Modeling aging and age-related diseases in the lab is challenging because classical vertebrate models have relatively long lifespans. Here we develop the first platform for rapid exploration of age-dependent traits and diseases in vertebrates, using the naturally short-lived African turquoise killifish. We provide an integrative genomic and genome-editing toolkit in this organism using our de novo-assembled genome and the CRISPR/Cas9 technology. We mutate many genes encompassing the hallmarks of aging, and for a subset, we produce stable lines within 2–3 months. As a proof-of-principle, we show that fish deficient for the protein subunit of telomerase exhibit the fastest onset of telomere-related pathologies among vertebrates. We further demonstrate the feasibility of creating specific genetic variants. This genome-to-phenotype platform represents a unique resource for studying vertebrate aging and disease in a high throughput manner and for investigating candidates arising from human genome-wide studies.

Crude oil is known to disrupt cardiac function in fish embryos. Large oil spills, such as the Deepwater Horizon (DWH) disaster that occurred in 2010 in the Gulf of Mexico, could severely affect fish at impacted spawning sites. The physiological mechanisms underlying such potential cardiotoxic effects remain unclear. Here, we show that crude oil samples collected from the DWH spill prolonged the action potential of isolated cardiomyocytes from juvenile bluefin and yellowfin tunas, through the blocking of the delayed rectifier potassium current (I(Kr)). Crude oil exposure also decreased calcium current (I(Ca)) and calcium cycling, which disrupted excitation-contraction coupling in cardiomyocytes. Our findings demonstrate a cardiotoxic mechanism by which crude oil affects the regulation of cellular excitability, with implications for life-threatening arrhythmias in vertebrates.

Expression was evaluated 7-10 days later. CHR2 and ARCH were excited with the 488nm and 514nm laser lines from an argon laser. Light power was controlled with an acousto-optic modulator, and focused onto selected areas (10-20mm) of the fibers via optic fibers. TPLSM imaging showed that CHR2 and ARCH were targeted to t-tubules and surface membranes. In fibers expressing CHR2, 1-3ms light pulses of increasing power elicited graded transient depolarizations until an action potential (AP) was triggered. The amplitude and FDHM of light-triggered APs were similar to those in response to current pulses. At all powers used, illuminating non-transfected fibers had no effect. Repetitive (<10Hz) pulsed illuminations elicited trains of APs with no failures. Failures at higher frequencies could be reduced by increasing the muscle fiber's input resistance (e.g. by blocking the chloride conductance with 9-AC), and/or increasing the illumination power. In voltage-clamped fibers, 40ms light pulses elicited slowly decaying inward currents with amplitudes graded with the applied illumination power. When fibers expressing ARCH were illuminated (at 514nm) otherwise suprathreshold current pulses became ineffective to trigger APs. Our results demonstrate that optogenetic tools can be used to excite or depress the excitability of adult skeletal muscle fibers. Since stem cell-derived cardiomyocytes will potentially play an important role in safety evaluations and drug screening in the future, we investigated cells from different stem cell suppliers with a new label-free technique based on impedance analysis. The used platform, called the CardioExcyte 96, allows precise impedance measurements at a time resolution of 1 ms and generates 96 recordings in parallel. Stem cells from GE Healthcare, Axiogenesis and Cellectis were successfully tested and data will be compared. Dose response curves or single-point screening experiments of several drugs (e.g. hERG and Cav1.2 blockers) were collected and will be presented. The capability of the data analysis software of the CardioExcyte 96 platform allows for a time effective analysis of data sets due to it's features and easeof-use. The presented data will show that the CardioExcyte 96 and the corresponding CardioExcyte Control software can be used efficiently to predict drug-induced arrhythmia for cardiac risk assessment. 3703-Pos Board B431The use of focused-ion-beam milling (FIB) to thin vitreously frozen cells for application of cryo-electron tomography is an emerging technology. However, successful application of cryo-FIB milling and tomography to frozen biological tissue has, to our knowledge, not been reported. This is because bulk tissue cannot be effectively frozen by simple plunge freezing of EM grids, but rather must be prepared by high-pressure freezing, which requires extra steps. The cryo-transfers that are required at each step of the process can cause devitrification, excessive frost contamination and mechanical damage, consequently resulting in unacceptably high failure rates. To ad...