Age and Exercise Related Changes in Motor Neuron Neuregulin and NT-4; Implications for Sarcopenia

Abstract

Sarcopenia, a syndrome characterized by progressive loss of skeletal muscle mass and strength is a process that begins to affect everyone over the age of forty (Cruz-Jentoft et al., 2010). Its effects are far-reaching; it deprives individuals of their independence and consumes healthcare and societal resources. In hopes of one day finding a tool for combatting the effects of sarcopenia, I have endeavoured to better understand the underlying causes of age-related muscle loss. Age-related muscle loss is associated with age-related denervation, a phenomenon involving the progressive loss of synapses between motor neurons and muscles, which results in muscle fibre atrophy and death that is characteristic of sarcopenia. The causes of denervation are currently unknown, but there are patterns exhibited by muscles undergoing denervation that may lead us to the underlying cause of denervation; highly active muscles and muscles that are proximal to the spinal cord are less affected by age-related denervation than distal/sedentary muscles. The existence of such a pattern implies that the causative factor is not present by the same magnitude in all muscles. Therefore, perhaps the causes of denervation can be found by studying the difference between muscles that are more/less affected by age-related denervation. Two hypothesised causative factors were examined: neuregulin (NRG) and neurotrophin-4 (NT-4). NRG is a Schwann cell trophic factor, produced in motor neurons and transported down the axon to supply myelinating and terminal Schwann cells along the axon and at neuromuscular junctions respectively. Schwann cells are essential for maintaining synapses and aging neuromuscular junctions undergo Schwann cell loss. NT-4 is a neurotrophin that is released by the muscle and binds to its receptor, TrkB, located on the presynaptic terminal. TrkB/NT-4 then undergoes retrograde transport to the motor neuron cell body where it acts as a neuron trophic factor. NT-4 is up regulated in response to muscle activity and is thought to stimulate NRG production in motor neurons. Further, NT-4, TrkB, and NRG knockout models produce mice with precocious sarcopenia. Therefore, in this thesis I proposed that an age-related decline in NT-4, results in a down-regulation of neuregulin (NRG), ultimately leading to Schwann cell loss and denervation seen in age. This study used immunohistochemistry to compare the amount of NT-4 and NRG in the spinal cords of young and old, sedentary and exercised mice to determine if there was an age or activity related effect on NT-4, NRG, or TrkB expression. NRG was also examined in the neuromuscular junctions to determine if there was an age-related decline in neuromuscular NRG. There was an age-related decline in motor neuron NT-4 and NRG; there was a larger difference between the means of young vs. old NRG (0.3 ± 0.12, P = 0.03) compared to NT-4 (0.1 ± 0.03, P < 0.01). Exercise increased NT-4 in old exercised compared to old sedentary animals suggesting that muscle activity increases NT-4 (0.3 ± 0.07, P = 8.5 x 10-4). However, no significant correlation between average motor neuron NT-4 and NRG (P = 0.08) suggests that production of NRG in motor neurons may not be significantly regulated by NT-4. Yet, NRG is correlated with indicators of synaptic health in a distal muscle tibialis anterior (R2 = 0.66, P = 0.01), suggesting that NRG is important in maintaining neuromuscular junction health. Surprisingly, there was no evidence for an age-related decline in neuromuscular NRG (P=0.57), despite a decline in NRG production in spinal cord motor neurons. This indicates that NRG in the neuromuscular junction is predominantly affected by factors other than its production in the motor neurons. This study found that NT-4 was decreasing with age and increased with exercise, which implicates NT-4 as a minor factor that regulates muscle innervation- albeit through unknown mechanisms. NRG continues to be a protein of interest as there was a direct linear correlation between average muscle neuromuscular NRG amount and synapse health within a muscle. Considering the independent role of both of these proteins in age-related fragmentation, this suggests that there isn’t one sole causative factor for sarcopenia, rather, sarcopenia is likely caused by decreased in several, if not all of the pathways that are needed to maintain innervation.