Automated cross-sectional analysis of trained, severely atrophied, and recovering rat skeletal muscles using MyoVision 2.0

Abstract

The number of myonuclei within a muscle fiber is an important factor in muscle growth, but its regulation during muscle adaptation is not well understood. We aimed to elucidate the timecourse of myonuclear dynamics during endurance training, loaded and concentric resistance training, and nerve silencing-induced disuse atrophy with subsequent recovery. We modified tibialis anterior muscle activity in free-living rats with electrical stimulation from implantable pulse generators, or with implantable osmotic pumps delivering tetrodotoxin (TTX) to silence the motor nerve without transection. We used the updated, automated software MyoVision to measure fiber type-specific responses in whole tibialis anterior cross-sections (~8000 fibers each). Seven days of continuous low frequency stimulation (CLFS) reduced muscle mass (-12%), increased slower myosin isoforms and reduced IIX/IIB fibers (-32%) and substantially increased myonuclei especially in IIX/IIB fibers (55.5%). High load resistance training (Spillover), produced greater hypertrophy (~16%) in muscle mass and fiber cross-sectional area (CSA) than low load resistance training (concentric, ~6%) and was associated with myonuclear addition in all fiber types (35-46%). TTX-induced nerve silencing resulted in progressive loss in muscle mass, fiber CSA, and myonuclei per fiber cross-section (-50.7%, -53.7%, -40.7%, respectively at 14 days). Myonuclear loss occurred in a fiber type-independent manner, but subsequent recovery during voluntary habitual activity suggested that type IIX/IIB fibers contained more new myonuclei during recovery from severe atrophy. This study demonstrates the power and accuracy provided by the updated MyoVision software and introduces new models for studying myonuclear dynamics in training, detraining, retraining, repeated disuse, and recovery.

Muscle-Specific Cellular and Molecular Adaptations to Late-Life Voluntary Concurrent Exercise

“…For all immunohistochemistry (IHC) analyses, entire muscle cross-sections were used (622 ± 35 fibers for soleus, 607 ± 27 fibers for plantaris, and 4616 ± 202 fibers for gastrocnemius), and most parameters were measured using semi-automated analysis software. 54 , 55 In soleus muscles of PoWeR mice, there were significantly more Type 1 (oxidative slow-twitch) fibers (+17%, P = 0.001, ES = 1.29) with a concomitant decrease in Type 2a (oxidative fast-twitch) fibers (−31%, P = 0.01, ES = −1.25) and no change in the relative frequency of 2x + 2b (glycolytic fast-twitch) fibers ( Figure 2A - 100 ) relative to sedentary controls. PoWeR resulted in elevated mean fiber CSA in the soleus (+12%, P = 0.01, ES = 1.04, Figure 2D ), corresponding with larger CSA in Type 1 (+9%, P = 0.04, ES = 0.85) and 2a (+16%, P = 0.01, ES = 1.07), but not relatively scarce 2x + 2b fibers ( Figure 2E ).…”

Cross Talk proposal: Myonuclei are lost with ageing and atrophy

“…2020; Viggars et al . 2022). However, recent evidence demonstrates that myonuclear transcriptional output can be modulated to accommodate variable myonuclear domains (Kirby et al .…”

Skeletal muscle DNA methylation and mRNA responses to a bout of higher versus lower load resistance exercise in previously trained men

“…Immediately following mounting, digital images were captured using a fluorescent microscope (Nikon Instruments, Melville, NY, USA) and a 10x objective. Standardized measurements of type I and type II fiber cross-sectional areas (fCSA) were performed using open-sourced software (MyoVision 2.0) [32,33]. A pixel conversion ratio value of 0.964 µm/pixel was applied to account for the size and bit-depth of images, and a detection range of detection from 500 to 12,000 μm 2 was used to ensure artifact was removed (i.e., large myofibers which may have not been in transverse orientation, or structures between dystrophin stains which were likely small vessels).…”