Effects of long‐term ethanol storage on muscle architecture

Abstract

Muscle excursion and force potential can be estimated from architectural variables, including mass, volume, fascicle length, and density. These have been collected from fresh specimens, preserved specimens, and sometimes mixed samples of both. However, preservation alters the gross morphology of muscles. This study aims to quantify the effects of long-term storage on myological properties across a sample of fresh and ethanol preserved Mus musculus specimens ranging in storage time from 16 to 130 years. Masses, volumes, and densities of biceps femoris, quadriceps femoris, and triceps surae were measured, and histological cross-sections of some specimens were used to evaluate the microscale effects of long-term fluid preservation. For the remainder of the sample, chemically dissected fascicle lengths were measured to evaluate the fixation effects on the linear dimensions of muscle architecture. Relative muscle mass, volume, fascicle length, average fiber area, and density, and percent fiber area were regressed against years stored in ethanol. Muscle size dropped steeply between fresh and stored samples, ultimately decreasing by 62 and 60%, respectively. These losses correlate with histologically measured shrinking of average muscle fiber area. Density of stored specimens plateaued 5% below that of fresh ones. Although muscles lost mass and volume during ethanol storage, fascicle lengths did not shorten significantly (presumably because they were preserved attached on either end to bone). This study demonstrates that muscle mass, volume, and density of specimens stored long-term in ethanol should be corrected by factors of 2.64, 2.49, and 1.054 respectively for comparability to fresh specimens.

Lizards as models to explore the ecological and neuroanatomical correlates of miniaturization

“…Specimen preparation for scanning such as formalin fixation and exposure to iodine-based solutions results in tissue shrinkage, the latter in a concentration-dependent manner (Vickerton et al, 2013;Buytaert et al, 2014). Long-term storage in ethanol, typical of museum specimens, also contributes to tissue deformation, hindering comparability with freshly collected specimens (Hedrick et al, 2018;Leonard et al, 2021). There is evidence that variable fixation times, on the other hand, do not significantly affect the degree of shrinkage (Powell & Leal, 2012).…”

A comparison of diceCT and histology for determination of nasal epithelial type

“…Before discussing the extent of shrinkage to which diceCT or histology may induce on nasal structures, we should acknowledge uncertainty regarding the extent of shrinkage caused by fixation and long-term ethanol storage. Multiple studies have documented that ethanol storage, especially long-term storage, causes marked shrinkage in soft tissue structures ( e.g ., Hedrick et al, 2018 ; Leonard et al, 2021 ). Some structures appear more susceptible than others ( e.g ., eyes), but it is also demonstrable that intact, whole animal specimens shrink less than isolated organs or tissue blocks ( Fox et al, 1985 ).…”

Scaling of fibre area and fibre glycogen concentration in the hindlimb musculature of monitor lizards: implications for locomotor performance with increasing body size

“…All fCSA values were multiplied by a conversion factor of 1.63212 to account for the reduction in fibre cross-sectional area that occurs as a result of 1 month of ethanol storage according to published decay equations (Leonard et al, 2021a,b). Storage in ethanol for 1 month after fixation in formalin causes a decrease in muscle mass of 41% compared with fresh tissue (Leonard et al, 2021a) and reductions in fCSA relate to reductions in muscle mass over longer periods by a reduced major axis regression [relative fCSA=(relative muscle mass×10,113.5)+5.9272; R 2 =0.9648] (Leonard et al, 2021b) such that fCSA can be expected to decrease by 61.27%. To control for differences in specimen preparation, fCSA was calculated among all samples, and also among a subset of the data where specimens were never frozen prior to analysis.…”