An in vivo rodent model of contraction-induced injury in the quadriceps muscle - PubMed (original) (raw)

An in vivo rodent model of contraction-induced injury in the quadriceps muscle

Stephen J P Pratt et al. Injury. 2012 Jun.

Abstract

Most animal studies of muscle contractile function utilise the anterior or posterior crural muscle (dorsiflexors and plantarflexors, respectively). An advantage to using these muscles is that the common fibular and tibial nerves are readily accessible, while the small size of the crural muscles is a disadvantage. Working with small muscles not only makes some in vivo imaging and the muscle testing techniques more challenging, but also provides limited amounts of tissue to study. The purpose of this study was to describe a new animal muscle injury model in the quadriceps that results in a significant and reproducible loss of force. The thigh of Sprague Dawley rats (N=5) and C57BL/10 mice (N=5) was immobilised and the ankle was attached to a custom-made lever arm. The femoral nerve was stimulated using subcutaneous electrodes and injury was induced using 50 lengthening ("eccentric") contractions through a 70° arc of knee motion. This protocol produces a significant and reproducible injury, with comparable susceptibility to injury in the rats and mice. This novel model shows that the quadriceps muscle provides a means to study whole muscle contractility, injury, and recovery in vivo. In addition to the usual benefits of an in vivo model, the larger size of the quadriceps facilitates in vivo imaging and provides a significant increase in the amount of tissue available for histology and biochemistry studies. A controlled muscle injury in the quadriceps also allows one to study a muscle, with mixed fibre types, which is extremely relevant to gait in humans and quadruped models.

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Figures

Figure 1. Injury model and lever arm specifications

The cartoon describes the injury model for the quadriceps**. A.** Animals are positioned supine on a raised bed (not shown) with a fine needle through the femoral condyles. The needle is secured into the thigh stabilizer, which can be adjusted in multiple planes and then stabilized. The needle/thigh stabilizer complex is aligned with the axis of the torque cell (dotted red line). The lever arm is adjusted so that the protruding arm is positioned over the anterior aspect of the ankle joint. Electrodes (not shown) are used to stimulate the quadriceps muscle. The isometric torque (toward extension) is recorded. For lengthening contractions, a stepper motor superimposes flexion (red arrow) onto a maximally contracting quadriceps muscle. B. Mouse lever arm has a total length of 32.16 mm and weighs 2.3 grams. Protruding ankle rod adjusts from 16 - 24 mm at increments of 0.84 mm. C. Rat lever arm has a total length of 49.71 mm and weighs 3.7 grams. The rat ankle rod adjusts from 28 - 40 mm also at increments of 0.84 mm. Both lever arms connect to the stepper motor and rotate through a pre-determined motion (up to 3600).

Figure 2. Torque data from in vivo apparatus

Representative trace recordings of torque during lengthening contractions in the mouse (A) and rat (B). Specifically, muscles were stimulated maximally for 200 milliseconds to induce a maximal isometric contraction before lengthening by the lever arm through a 70° arc of motion and at an angular velocity of 900°/s. The representative trace recordings show peak isometric (1) and eccentric (2) torque in newton millimeters (Nmm) from a single repetition. C. The histogram shows mean torque production before and after injury from both the mouse and rat quadriceps muscles. The rat showed an approximately 8-fold torque production over that of the mouse. Both animals showed a significant (* = p < 0.05) loss of force (82 ± 1% in the mouse and 73 ± 4.4% in the rat) after 50 repetitions. D. The histogram shows the mean percent loss of torque with each repetition in the mouse and rat over 50 repetitions.

Figure 3. In vivo imaging

We used magnetic resonance imaging (MRI) to examine the quadriceps in the mouse (A-B) and rat (C-D) quadriceps muscles**.** Representative T-2 weighted images of the healthy (A, C) versus injured (B, D) quadriceps are shown. The primary location of increase signal intensity was in the distal quadriceps, close to the muscle-tendon junction (MTJ). Asterisk indicates the vastus intermedius for purposes of orientation. Interestingly, the majority of damage was localized to the vastus medialis and lateralis, while the vastus intermedius was generally spared (belly of the rectus femoris is not seen in these distal cross-sections). The MRI was repeated for 3 mice and 3 rats with similar findings (mild T2 changes confined to the distal aspect of the quadriceps).

Figure 4. Histology

Standard H&E staining of cross-sections of un-injured and injured quadriceps in the mouse and rat. The injured tissue was assessed immediately after injury (Day 0), 4 days after injury (Day 4) and 8 days after injury (Day 8). Uninjured muscle contained peripheral nuclei and limited variation in fiber size, consistent with healthy muscle. Injured muscle at Day 0 showed foci of mild lymphocytic inflammation (arrows) surrounding small perimysial blood vessels, but no evidence of myonecrosis, fibrosis, or other inflammation. Such findings were magnified at a later time point (Day 4); however inflammation was unremarkable by Day 8. Bar = 50 microns

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An in vivo rodent model of contraction-induced injury in the quadriceps muscle - PubMed (original) (raw)