ECSS Paris 2023: CP-PN16
INTRODUCTION: Titin connects the myofilaments in skeletal muscle and plays a pivotal role in modulating passive and active muscle tension. Residual force enhancement (RFE) describes the phenomenon where force remains elevated following an active stretch compared to a purely isometric contraction at the same final length. Despite its relevance to near-instantaneous muscle force control, the structural underpinnings of RFE remain elusive. This study investigates the contribution of titin to RFE by using 2 genetically modified mouse models: the titin cleavage (TC) and the muscular dystrophy with myositis (mdm) mouse lines,. Small-angle X-ray diffraction was used to elucidate structural changes underlying titins mechanical role during muscle contractions. METHODS: Fast-twitch, skinned skeletal muscle fibers were isolated from TC (heterozygous) and mdm (wild type and homozygous) mice. In TC mice, a splice site was engineered into the I-band region of titin, enabling rapid and specific cleavage, thereby reducing titin-based force and stiffness. In mdm mice, a small deletion in the I-band of titin nearly ablates RFE. Mechanical properties and sarcomeric structures were assessed during: (1) isometric contractions at short sarcomere lengths (IsoShort), (2) isometric contractions at long sarcomere lengths (IsoLong), and (3) after active lengthen-hold from IsoShort to IsoLong (IsoRFE). X-ray diffraction captured the sarcomeric structural signature in each condition. Stats - full-factorial ANOVA design with individual random effect. Box-Cox transformation ensured normality and variance homogeneity. RESULTS: Cleaving 50% of I-band titin in TC fibers significantly reduced RFE from 30.69 ± 3.35% to 17.44 ± 2.24% of the reference force (IsoLong; P < 0.01). Structural analyses revealed that IsoRFE contractions exhibited increased thick filament strain and reduced lattice spacing compared to IsoLong (P < 0.01). Notably, these RFE-specific structural changes were absent in mdm fibers but were prominent in wild-type controls (P < 0.01). We estimate that titin-mediated reductions in lattice spacing and thick filament elongation account for approximately 40-70% of RFE. The remaining 30-60% is likely attributable to an activation-dependent, 2-3-fold increase in titin stiffness preceding eccentric stretch, which relatively increases titin-based forces during and after the stretch, compared to a passive stretch-hold. CONCLUSION: These results underscore titin’s integral role in modulating sarcomeric tension and its essential contribution to RFE. The structural signatures associated with RFE—decreased lattice spacing and increased thick filament strain—are directly influenced by titin-based forces. The absence of the mechanical and structural signatures of RFE in mdm mutants highlights the necessity of I-band titin for proper RFE. Our data suggest that RFE involves both structural adjustments and an activation-dependent modulation of titin stiffness, though the precise molecular imechanisms remain to be elucidated.
Read CV Michel KuehnECSS Paris 2023: CP-PN16
INTRODUCTION: Age-related declines in skeletal muscle mass and function lead to impairments in mobility and decreased quality of life in older adults. At the single muscle fiber level, many studies show preserved function of slow myosin heavy chain (MyHC) I fibers but decrements in size and contractile function of fast MyHC II fibers(1). Studies of fast fibers from old rats suggest impairments in myofibrillar ATPase efficiency with age(2,3), however it is unknown if these impairments also occur in aging human skeletal muscle. METHODS: Muscle biopsies were obtained from the vastus lateralis of 6 young (4F; 26.5±4.8 yrs) and 5 older (3F; 71.1±6.1 yrs) adults to assess single fiber myofibrillar ATPase activity via a micro epifluorescence system that measures NADH fluorescence during ATP hydrolysis. Activity was measured in relaxing conditions (basal ATPase) and in saturating Ca2+ activating conditions (pCa 4.5) during isometric contractions of chemically permeabilized single muscle fibers. Simultaneous measures of force and NADH fluorescence were made at 20°C, and the MyHC isoform for each fiber determined by 5% SDS-PAGE. RESULTS: A total of 63 (33 old) MyHC I and 82 (44 old) MyHC II fibers were tested. Basal ATPase activity in relaxing solution was higher in young compared with older adults in both MyHC I (754±115 vs 583±87nM NADH/s; p=0.004) and MyHC II fibers (733±129 vs 606±131nM NADH/s; p=0.005); however, there were no differences when the basal ATPase activity was normalized to fiber size for either the MyHC I (0.15±0.07 vs 0.12±0.04nM NADH/s/µm2; p=0.638) or MyHC II fibers (0.13±0.08 vs 0.18±0.14nM NADH/s/µm2; p=0.852). Maximal Ca2+ ATPase activity was greater in young compared with older adults in both MyHC I (1,320±410 vs 910±227nM NADH/s; p=0.010) and MyHC II (3,628±1,022 vs 2,408±923nM NADH/s; p=0.031) fibers. When normalized to absolute and size-specific force, ATPase activity of MyHC I fibers was higher in young compared to older adults (2,173±824 vs 1,631±510nM NADH/s/mN; p=0.035 and 11.4±3.9 vs 8.2±2.6nM NADH/s/kN/m2; p=0.044), suggesting a greater myofibrillar ATPase efficiency with aging. In contrast, there were no differences in the MyHC II fibers when normalized to absolute (4,322±2,516 vs 3,919±1,431nM NADH/s/mN; p=0.428) or size-specific force (23.9±7.3 vs 16.8±7.2nM NADH/s/kN/m2; p=0.254). CONCLUSION: These preliminary results suggest that during isometric contractions, older adults have more efficient MyHC I skeletal muscle fibers but that contractile efficiency of MyHC II fibers is unaltered by aging. The more efficient MyHC I fibers may explain, in part, the fatigue resistance and improved metabolic economy found in older adults during voluntary isometric contractions(4). References: 1. Grosicki GJ. et al. J Physiol. 600. (2022) 2. Lowe DA. et al. Am J Physiol Cell Physiol. 283. (2002) 3. Lowe DA. et al. Mech Ageing Dev. 125. (2004) 4. Lanza IR. et al. J Physiol. 583. (2007) Funding: Supported by NIA R01 grant (AG048262) to CWS, SKH, and RHF.
Read CV Laura TeigenECSS Paris 2023: CP-PN16
INTRODUCTION: Environmental enrichment (EE) enhances brain function through neuroprotection and neuroplasticity. Our previous study demonstrated that EE induces soleus muscle hypertrophy and increases spontaneous physical activity (Sudo et al., 2023). However, the relationship between soleus muscle hypertrophy, locomotive behavior, and anxiety-like behavior under EE conditions remains unclear. This study aimed to clarify the relationship between soleus muscle hypertrophy and EE exposure under multiple conditions. METHODS: Wistar rats were divided into four groups: EE (wheel + objects), EE-R (wheel only), EE-O (objects only), and CONT (standard environment) (N=6 per group). Physical activity (PA) was continuously recorded via a three-axis accelerometer. Wheel running duration was measured during the 12-hour dark phase in the final week. After four weeks, spatial memory was assessed using the Morris Water Maze (MWM) test, and anxiety-like behavior was evaluated using the Elevated Plus Maze (EPM) test. Soleus and plantaris muscle cross-sectional area (CSA) was measured from hematoxylin-eosin (HE) stained sections to assess muscle hypertrophy. RESULTS: Total PA was higher in EE and EE-R than in CONT (P<0.001), with no difference between EE and EE-R (P=0.63). Wheel running duration was longer in EE (56.3 ± 14.7 min) than in EE-R (37.2 ± 9.4 min, P=0.03). The EPM test showed a higher open-to-closed arm time ratio (O/C ratio) in EE (203.5 ± 27.2), EE-R (98.1 ± 25.7), and EE-O (78.8 ± 33.7) than in CONT (11.4 ± 7.6, P<0.01). Soleus muscle CSA increased in all EE groups (EE: 2889 ± 162, EE-R: 2575 ± 81, EE-O: 2572 ± 132 μm², P<0.01 vs. CONT: 2184 ± 203 μm²). Soleus muscle CSA was also greater in EE than in EE-R (P=0.01). In contrast, plantaris muscle CSA did not differ among groups (EE: 2973 ± 297, EE-R: 2898 ± 514, EE-O: 2695 ± 397, CONT: 2590 ± 399 μm²). Furthermore, soleus muscle CSA correlated positively with PA (r²=0.38, P=0.002) and the O/C ratio in EPM (r²=0.65, P<0.001). In EE and EE-R, soleus muscle CSA also correlated positively with wheel running duration (r²=0.48, P=0.01), whereas plantaris muscle CSA showed no correlations. CONCLUSION: This study revealed that soleus muscle hypertrophy and reduction in anxiety-like behavior are associated with environmental enrichment. Soleus hypertrophy was observed in EE, EE-R, and EE-O groups, with the greatest increase in EE. Since wheel running duration was lower in EE-R than in EE, soleus muscle hypertrophy may be associated with both locomotive activity involving object interaction and spontaneous wheel running in an enriched environment. Furthermore, soleus muscle hypertrophy, in concert with spontaneous intermittent physical activity, may contribute to reductions in anxiety-like behavior. Reference: Sudo M., Kano Y., Ando S. The effects of environmental enrichment on voluntary physical activity and muscle mass gain in growing rats. Front. Physiol. 14, 2023.
Read CV Mizuki SudoECSS Paris 2023: CP-PN16