"Avatar", a Modified Ex vivo Work Loop Experiments Using In vivo Strain and Activation

Summary

This article details the methodology for emulating in vivo muscle force production during ex vivo work loop experiments using an "avatar" muscle from a laboratory rodent to assess the contributions of strain transients and activation to the muscle force response.

Abstract

Movement behaviors are emergent features of dynamic systems that result from muscle force production and work output. The interplay between neural and mechanical systems occurs at all levels of biological organization concurrently, from the tuning of leg muscle properties while running to the dynamics of the limbs interacting with the ground. Understanding the conditions under which animals shift their neural control strategies toward intrinsic muscle mechanics ('preflexes') in the control hierarchy would allow muscle models to predict in vivo muscle force and work more accurately. To understand in vivo muscle mechanics, ex vivo investigation of muscle force and work under dynamically varying strain and loading conditions similar to in vivo locomotion is required. In vivo strain trajectories typically exhibit abrupt changes (i.e., strain and velocity transients) that arise from interactions among neural activation, musculoskeletal kinematics, and loads applied by the environment. The principal goal of our "avatar" technique is to investigate how muscles function during abrupt changes in strain rate and loading when the contribution of intrinsic mechanical properties to muscle force production may be highest. In the "avatar" technique, the traditional work-loop approach is modified using measured in vivo strain trajectories and electromyographic (EMG) signals from animals during dynamic movements to drive ex vivo muscles through multiple stretch-shortening cycles. This approach is similar to the work-loop technique, except that in vivo strain trajectories are scaled appropriately and imposed on ex vivo mouse muscles attached to a servo motor. This technique allows one to: (1) emulate in vivo strain, activation, stride frequency, and work-loop patterns; (2) vary these patterns to match in vivo force responses most accurately; and (3) vary specific features of strain and/or activation in controlled combinations to test mechanistic hypotheses.

Introduction

Moving animals achieve impressive athletic feats of endurance, speed, and agility in complex environments. Animal locomotion is particularly impressive in contrast to human-engineered machines-the stability and agility of current-legged robots, prostheses, and exoskeletons remain poor compared to animals. Legged locomotion in natural terrain requires precise control and rapid adjustments to alter the speed and maneuver environmental features that act as unexpected perturbations^1,2,3,4. Yet, understanding non-steady locomotion is inherently c....

Protocol

All animal studies were approved by the Institutional Animal Care and Use Committee at Northern Arizona University. Extensor digitorum longus (EDL) muscles from male and female wild-type mice (strain B6C3Fe a/a-Ttn^mdm/J), aged 60-280 days, were used for the present study. The animals were obtained from a commercial source (see Table of Materials), and established in a colony at Northern Arizona University.

1. Selecting in vivo strain trajectory and p.......

Discussion

While organisms move seamlessly across landscapes, the underlying loads and strains that the muscles experience vary drastically^1,6,23. During both in vivo locomotion^1,24 and in "avatar" experiments, muscles are stimulated submaximally under cyclical, non-steady conditions. The isometric force-length and isotonic force-velocity relationships are not well.......

Materials

Name	Company	Catalog Number	Comments
Braided Non-Absorbable Silk Suture 4-0	Mersilk	734H
Calcium Chloride Dihydrate (CaCl2)	Sigma-Aldrich	1086436	Krebs-Henseleit solution
Dextrose	Sigma-Aldrich	D9434	Krebs-Henseleit solution
HEPES	Sigma-Aldrich	PHR1428	Krebs-Henseleit solution
Hydorchloric Acid (HCl)	Sigma-Aldrich	1.37055	Krebs-Henseleit solution
LabView Data Collection	Lab-View
Magnesium Sulfate (MgSO4)	Sigma-Aldrich	M7506	Krebs-Henseleit solution
Potassium Chloride (KCl)	Sigma-Aldrich	P3911	Krebs-Henseleit solution
Potassium Phosphate Monobasic (KH2PO4)	Sigma-Aldrich	5.43841	Krebs-Henseleit solution
S88 Stimulator	Grass	M643H05	Available for purchase on Ebay
Series 300B Lever System	Aurora	1200A	includes water-jacket tissue bath
Sodium Bicarbonate (NaHCO3)	Sigma-Aldrich	S5761	Krebs-Henseleit solution
Sodium Chloride (NaCl)	Sigma-Aldrich	S9888	Krebs-Henseleit solution
Sodium Hydroxide (NaOH)	Sigma-Aldrich	S5881	Krebs-Henseleit solution
Wild Type Mice	Jackson Laboratory	B6C3Fe a/a Ttn mdm/J