bioRxiv2020
Zebrafish (Danio rerio) swim within days of fertilization, powered by muscles of the axial myotomes. Forces generated by these muscles can be measured rapidly in whole, intact larval tails by adapting protocols developed for ex vivo muscle mechanics. But it is not known how well these measurements reflect the function of the underlying muscle fibers and sarcomeres. Here we consider the anatomy of the 5-day-old, wild-type larval tail, and implement technical modifications to measuring muscle physiology in intact tails. Specifically, we quantify fundamental relationships between force, length, and shortening velocity, and capture the extreme contractile speeds required to swim with tail-beat frequencies of 80-100 Hz. Therefore, we analyze 1000 frames/second movies to track the movement of structures, visible in the transparent tail, which correlate with sarcomere length. We also characterize the passive viscoelastic properties of the preparation to isolate forces contributed by non-muscle structures within the tail. Myotomal muscles generate more than 95% of their maximum isometric stress (76{+/-}3 mN/mm2) over the range of muscle lengths used in vivo. They have rapid twitch kinetics (full width at half-maximum stress: 11{+/-}1 msec) and a high twitch to tetanus ratio (0.91{+/-}0.05), indicating adaptations for fast excitation-contraction coupling. Although contractile stress is relatively low, myotomal muscles develop high net power (134{+/-}20 W/kg at 80 Hz) in cyclical work loop experiments designed to simulate the in vivo dynamics of muscle fibers during swimming. When shortening at a constant speed of 7{+/-}1 muscle lengths/second, muscles develop 86{+/-}2% of isometric stress, while peak instantaneous power during 100Hz work loops occurs at 18{+/-}2 muscle lengths/second. These approaches can improve the usefulness of zebrafish as a model system for muscle research by providing a rapid and sensitive functional readout for experimental interventions. Statement of significanceThe zebrafish (Danio rerio) may prove a uniquely efficient model system for characterizing vertebrate muscle physiology. Transparent, drug-permeable larva - each, in essence, a fully functional muscle - can be generated rapidly, inexpensively, and in large numbers. Critically, the zebrafish genome contains homologs of major muscle genes and is highly amenable to manipulation. To reach its potential, reliable (and preferably rapid) means are needed to observe the effects of experimental interventions on larval muscle function. In the present study we show how mechanical measurements made on whole, intact larval tails can provide a readout of fundamental muscle-mechanical properties. Additionally, we show that these muscles are among the fastest ever measured, and therefore worthy of study in their own right.