Rat hindlimb unloading: soleus histochemistry, ultrastructure, and electromyography

Abstract

Soleus muscle atrophy was induced by hindlimb unloading of male Sprague-Dawley rats (305 +/- 15 g) for 4, 7, and 10-14 days. Controls (291 +/- 14 g) were housed in vivarium cages. Soleus electromyogram (EMG) activity was recorded before and during tail suspension. Unloading caused progressive reduction in the muscle-to-body weight ratio. After 14 days, type I and IIa fibers decreased in area 63 and 47%, respectively. Subsarcolemmal mitochondria and myofibrils were degraded more rapidly than intermyofibrillar mitochondria and the cell membrane. After 10 days, 3% of the fibers exhibited segmental necrosis; affected fibers were all high-oxidative type IIa fibers. This suggested ischemic injury. By 13 days, 30% of the fibers possessed central corelike lesions involving primarily type I fibers. Video monitoring revealed abnormal plantar flexion of the hindfeet by 4 days; this posture shortened the soleus working range. Corelike lesions indicated adaptation to the shortened length. No morphological signs of denervation were detected. EMG activity shifted from tonic to phasic, and aggregate activity was 13% of normal after 7 days. These findings indicate that the atrophy and pathological changes result from unloaded contractions, reduced use, compromised blood flow, and shortened working length.

Passive stretch inhibits central corelike lesion formation in the soleus muscles of hindlimb-suspended unloaded rats

“…HSU was performed for 12 days as described previously (16,29). Each rat was fitted with a tail harness and suspended by a wire just high enough to prevent the hindlimbs from bearing weight on the grate floor.…”

“…Control and treated muscles were stored in liquid nitrogen and cross-sectioned serially at 10 m in a cryostat at Ϫ20°C. The sections were collected on glass slides (Colorfrost*/Plus, Fisher Scientific) and histochemically reacted for myofibrillar ATPase activity after acid (pH 4.35) and alkaline (pH 9.4) preincubation as previously performed (29). The splinted and nonsplinted muscles of an animal as well as a control soleus were always collected on the same slide and histochemically reacted together.…”

“…Rats were housed in an isolated and environmentally controlled room at 25 Ϯ 1°C and on a 12:12-h light-dark cycle. Animal treatment and husbandry procedures were approved by the Medical College of Wisconsin's Animal Care Committee and complied with the Laboratory Animal Welfare Act.HSU was performed for 12 days as described previously (16,29). Each rat was fitted with a tail harness and suspended by a wire just high enough to prevent the hindlimbs from bearing weight on the grate floor.…”

“…Preventing rats from touching the ground with their hindlimbs causes the antigravity soleus muscles to atrophy due to decreased use, unloading, and shortened working range (29). Central corelike lesions (CCLs), which are areas of focal myofibril dissolution, appear within soleus fibers by day 12 of HSU.…”

“…Central corelike lesions (CCLs), which are areas of focal myofibril dissolution, appear within soleus fibers by day 12 of HSU. For rat solei, the CCLs were postulated to result from the ϳ20% shortened contraction range, which is a consequence of suspensioninduced foot drop posture (plantar flexion) (22,29). Foot drop posture of humans floating in microgravity was first documented in their neutral body position during Skylab missions (26).…”