The Effect of Ageing on the Calcium Release Unit of Rat Skeletal Muscle

Abstract

As life expectancy has increased in developed countries, the aging population has grown exponentially. Population data forecast that by 2040, there will be 1.2 million individuals over 65 years of age in Australia. While European figures predict that 4.2 million of the population will be elderly, corresponding to approximately 4% of the population. Comorbidities of ageing constitute a significant population health risk, and one of the more insidious and costly is sarcopenia. Sarcopenia is the progressive loss of skeletal muscle mass and muscle strength as we age, leading to increasing frailty. Changes in fibre size and muscle atrophy were initially considered causative factors in sarcopenia; however, research suggests that additional pathophysiological changes are involved, specifically related to excitation-contraction coupling (EC coupling). A number of observations have been made regarding changes in human muscle fibre composition, while animal models using rodents have demonstrated age-related impairments of EC coupling. Electron microscopy data of young and old humans have also confirmed EC uncoupling and t-tubule aggregation in skeletal muscles. EC coupling is facilitated by two calcium (Ca2+) channels, the dihydropyridine receptor (DHPR) and the ryanodine receptor (RyR1). RyR1 is the gatekeeper at the sarcoplasmic reticulum for the release of Ca2+ required to convert action potentials into contractions of the muscle. RyR1 is regulated by a complex of proteins forming the Calcium Release Unit (CRU). These major proteins in the CRU are Triadin, Junctin, and Calsequestrin (CSQ). This project also examines, the Junctophillin (JP) proteins 1 and 2 (referred to as anchoring proteins) which keep the EC coupling apparatus intact. Changes in muscle contraction speed also occur during ageing influenced by three main factors: MyHC isoforms; Ca2+ release, and uptake from the sarcoplasmic reticulum (SR) (Schiaffino and Reggiani 2011a). Research suggests that atrophy alone is not the only factor contributing to altered contraction speed, as changes in protein abundance and interactions can also affect contraction speed through the slowed transmission of the EC coupling signal. This study investigated the fibre type distribution and the levels of proteins involved in the formation of CRU: RyR1, Triadin, Junctin, Calsequestrin (also isoforms), and both the attachment proteins JP-1 & 2 in fast and slow muscle from 18 rats (aged 6 wk to 72 wk). The expression of CSQ isoforms was also investigated to understand the effect of ageing in slow CSQ (CSQ2) and fast CSQ (CSQ1). This pilot study confirmed that the extensor digitorum longus (EDL) was predominantly a fast-twitch fibres, whereas soleus had primarily slow-twitch fibres. As anticipated, age significantly influenced fibre type composition in EDL and soleus which was used in correlation data for CRU and attachment proteins. Moreover, this study was conducted to examine CRU and attachment proteins in correlation with fibre type information. Contrary to previous findings, changes in protein expression were observed with ageing rodents, including a decrease in Junctin and an increase in JP 2 in combined data of EDL and soleus muscles. Encouragingly, a highly significant positive correlation was shown between type 1 fibres and CSQs, as well as type 2 fibres with RyR1. In addition, a highly significant negative correlation was observed between type1 fibres with RyR1, and type 2 fibres with CSQs. As expected, fibres type correlation of CSQ fast (CSQ1) and CSQ slow (CSQ2) was observed with slow and fast fibres respectively in combined muscle samples. Show more