Introduction

Throughout natural adult ageing, muscle mass, strength, and power start to decline around the fifth decade of life (Larsson et al. 1979; Janssen et al. 2000) and these declines accelerate into very old age (Borkan et al. 1983; Ditroilo et al. 2010; Kallman et al. 1990). While these factors are undoubtedly important for maintaining health and function (Metter et al. 2002; Newman et al. 2006), the rate at which a muscle can generate force may be more important for optimizing neuromechanical function, such as recovering from a slip, trip, or fall (Bento et al. 2010; Pijnappels et al. 2005, 2008; Lomborg et al. 2022).

Rate of force development (RFD), or the ability to produce force quickly, is strongly related to an older individual’s ability to rise from a chair (Fleming et al. 1991), ascend a flight of stairs (Altubasi 2015), and maintain mobility (Hester et al. 2021). Rapid RFD is suggested to be more important than maximal strength in terms of reducing fall risk (Bento et al. 2010; Lomborg et al. 2022) as the amount of force generated within the initial 200 ms is critical to catch one’s self from a potential fall (Pijnappels et al. 2005). In old age, RFD is impaired to a greater extent than strength (Thelen et al. 1996), especially for lower body muscle groups (Candow and Chilibeck 2005), but the mechanisms behind this preferential reduction in RFD capacity are not fully understood. There are several neuromuscular factors that have been associated with the age-related reduction in RFD such as reduced motor neuron firing rates (Klass et al. 2008), asynchronous firing of motor units (Piasecki et al. 2016), a lower rate of activation (Reid et al. 2012; Clark et al. 2011), increased antagonist coactivation (Izquierdo et al. 1999; Klein et al. 2001), neuromuscular junction degradation (Hourigan et al. 2015; Oda 1984), and impairments in force transmission (Hughes et al. 2015). Although these age-associated impairments in neuromuscular function clearly play a substantial role in the diminished ability of older individuals to produce force rapidly, they are not the only factors that affect RFD, as intrinsic properties of skeletal muscle, such as muscle quality (Gerstner et al. 2017), the relative abundance of Type I and Type II fibres (Harridge et al. 1996; Korhonen et al. 2006), and musculotendinous stiffness (Bojsen-Møller et al. 2005), among others, also contribute to rapid force production.

Rate of force development can be partitioned into various time epochs (e.g. early and late), in which peak performance is dependent on specific physiological determinants (Andersen and Aagaard 2006; Maffiuletti et al. 2016). The earliest phase (< 50 ms) is often said to be associated with ‘intrinsic contractile properties of the muscle’ while the late phase (> 50 ms) appears to be more reliant on mechanisms related to maximal strength (Andersen and Aagaard 2006) (i.e. muscle mass (Suchomel and Stone 2017), muscle quality (Metter et al. 1999), and neuromuscular activation (Moritani 1979). These relationships are based on associations between the RFD of electrically evoked twitches and voluntary RFD of the knee extensors (Andersen and Aagaard 2006). While electrically evoked twitch contractions provide important insight into skeletal muscle function, there are numerous confounding factors when attempting to relate evoked joint-level measures to intrinsic contractile properties of skeletal muscle. These factors include, but are not limited to, musculotendinous stiffness, fascicle length (Lieber and Ward 2011), sarcolemmal excitability (Bigland-Ritchie et al. 1979), and differences in moment arm length (Krevolin et al. 2004). Therefore, direct measures of intrinsic contractile properties are necessary to elucidate any potential relationship between cellular- and joint-level function as related to RFD.

Single muscle fibre preparations offer an experimental design that controls for the previously described factors that may confound measures of intrinsic contractile properties when measured via electrically evoked twitches. Previous studies have provided evidence that single muscle fibre properties are related to joint-level power (Miller et al. 2013) and RFD (Callahan et al. 2015), highlighting an important link between single fibre and joint-level neuromuscular function. Specifically, Callahan et al. (2015) investigated the relationship between intrinsic muscle fibre properties (i.e. fibre size, shortening velocity, and power) and joint-level RFD in healthy older adults, as well as older adults diagnosed with knee osteoarthritis. They found that Type IIA/X fibre cross-sectional area (CSA) in the healthy cohort and shortening velocity of Type I fibres from those with osteoarthritis were related to peak RFD. When group data were pooled, Type I and IIA CSA, as well as Type IIA specific tension, were all significantly related to peak RFD. While results from Callahan et al. (2015) provide important evidence for links between cellular and joint-level muscle function, building upon their work by relating intrinsic contractile properties across several RFD time epochs, rather than solely peak RFD, would clarify the role these properties have during force development. Further, no study has yet examined the relationship between RFD at the joint and single fibre level. The single fibre equivalent of RFD, rate of force redevelopment (ktr), allows for the direct investigation of intrinsic muscle contractile properties as it is a measure of the rapid transition from non-force bearing to force bearing cross-bridge states (i.e. rate of cross-bridge attachment), making it an ideal method for relating cellular mechanisms to joint-level RFD (Brenner and Eisenberg 1986; Power et al. 2016; Mazara et al. 2021).

Thus, investigating the relationship between single fibre and joint-level RFD may further elucidate mechanisms contributing to the age-related reduction in RFD and whether partitioning RFD into time epochs is truly representative of muscle ‘intrinsic contractile properties’ at the cellular level. Therefore, the purpose of this study was to investigate the relationship between single muscle fibre ktr and joint-level rate of torque development (RTD) of the knee extensors in young and older adults. Based off of the proposed relationship between intrinsic contractile properties, measured via electrically evoked twitches (Andersen and Aagaard 2006), and voluntary RTD, we hypothesized that single muscle fibre ktr will be associated with joint-level RTD in both young and older adults and that this relationship will be stronger during the early as compared to late phase of RTD.

Methods

Participants

Data in the present study came from the same thesis used to generate the results in Mazara et al. (2021). Participants were recruited from the University of Guelph and surrounding communities. Eight healthy, young (26.4 ± 1.5 yrs, 179 ± 3.3 cm, 83.9 ± 4.2 kg) and eight healthy, independently living, old (70.1 ± 2.8 yrs, 174 ± 2.0 cm, 74.8 ± 3.4 kg) males underwent neuromuscular assessment of the knee extensors and a muscle biopsy of the vastus lateralis. Anthropometric and physical activity data were reported previously (Mazara et al. 2021). For exclusion criteria, see Mazara et al. (2021). Written, informed consent was obtained from all participants, and all procedures and protocols were approved by the local Research Ethics Board and conformed to the Declaration of Helsinki.

Peripheral nerve stimulation

The femoral nerve was stimulated using a Digitimer DS7AH constant current stimulator (Digitimer Ltd., Welwyn Garden City, UK). The anode (Cleartrace 1700-030 ECG Electrode, ConMed, Utica, New York, USA) was placed over the inguinal triangle and the cathode was placed over the inferior gluteal fold. The cathode was a custom-made aluminium electrode pad (~ 6–8 cm width, ~ 8–10 cm length) wrapped in damp paper towel and covered in conductive gel. First, the level of current needed to deliver a supramaximal twitch was found by delivering increasing amounts of current until twitch torque plateaued, and then the current was increased by 10%. This amplitude was used for all subsequent stimulations.

Neuromuscular assessment

Participants performed maximal voluntary contractions (MVC) of the left knee extensors while seated on a HUMAC Norm dynamometer (CSMi Medical Solutions, MA) with their hip angle set at 110° and knee angle set at 80° of knee extension (180° being full extension). MVCs were performed until the participant achieved ≥ 90% voluntary activation during two MVCs, as assessed by the interpolated twitch technique (Power et al. 2014). During each MVC, participants were given real time visual feedback of torque production and verbal encouragement to ensure a maximal effort was given. At least 3 min of rest was given between each MVC to mitigate fatigue.

To measure RTD, using the same joint configuration as listed above, two brief (< 1 s) explosive isometric contractions (instruction: “kick as fast and as hard as you can”) were performed consecutively followed by a 1 min rest; this was repeated twice for a total of three trials and six explosive contractions (Maffiuletti et al. 2016). Torque data were sampled at 1000 Hz using a 12-bit analogue-to-digital converter (PowerLab System 16/35; ADInstruments, Bella Vista, Australia).

Neuromuscular assessment analysis

Peak MVC torque was taken at the highest point along the torque–time curve during the MVC with the highest torque and voluntary activation. To calculate the RTD of the isometric explosive contractions, the average slope of the torque–time trace was calculated during four different time epochs: 0–30 ms, 0–50 ms, 0–100 ms, and 0–200 ms. (Fig. 1). The greatest slope along the torque–time curve was deemed peak RTD. Onset occurred when the torque trace exceeded 3 standard deviations above baseline (De Ruiter et al. 2006). An average of the three trials was used for each participant. Peak twitch torque and twitch RTD were defined as the greatest twitch torque amplitude and slope, respectively. Voluntary peak RTD and twitch RTD were normalized to peak MVC torque and twitch torque, respectively, to account for differences in muscle strength between participants (Callahan et al. 2015; Korhonen et al. 2006).

Fig. 1
figure 1

Representative traces of A an electrically evoked twitch with determination of rate of torque development (RTD), B A joint-level voluntary contraction with RTD determination, and C a single fibre activation with a shorten–restretch manoeuvre to assess rate of force redevelopment (ktr). Electrically evoked twitch and voluntary peak RTD were defined as the greatest slope along the torque–time curve. Voluntary RTD was also calculated across four time epochs: 0–30 ms, 0–50 ms, 0–100 ms, and 0–200 ms. Single fibre ktr was determined by fitting force redevelopment data following a rapid shortening restretch manoeuvre using a mono-exponential equation: y = a (1 − e−kt) + b. Inset D shows the force and length recordings during the ktr test

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Biopsy and muscle fibre preparation

Following the neuromuscular assessment, a percutaneous muscle biopsy using the modified suction Bergstrom technique (Tarnopolsky et al. 2007) was taken from each participant for single muscle fibre experiments. The biopsy site was approximately midway between the lateral femoral epicondyle and the greater trochanter of the femur on the left vastus lateralis, and the needle was placed at an angle close to parallel with fascicle direction to obtain the longest possible fibres. Biopsies were taken from the vastus lateralis owing to its contribution to knee extension torque, and safety of the participant (Farahmand et al. 1998). The muscle sample was placed into storage solution (relaxing-glycerol 50:50 with protease inhibitors) to permeabilize at 4 °C for 24 h. The muscle sample was then placed in fresh storage solution at −20 °C for at least 1 week and up to 4 weeks (Teigen et al. 2020). On testing days, the sample was placed in a petri dish with fresh storage solution, and a strip was cut from which single fibres could be dissected from. Single fibres were dissected and placed into relaxing solution for ~ 1 min. Fibres were transferred into a temperature-controlled chamber filled with relaxing solution and tied with nylon suture knots between a force transducer (model 403A; Aurora Scientific, Toronto, ON, Canada) and a length controller (model 322C; Aurora Scientific).

Single fibre experiments

As reported previously (Mazara et al. 2021), the average sarcomere length was first measured using a highspeed camera (Aurora Scientific, Aurora, ON) and set at ~ 2.8 μm to adjust for shortening upon ‘isometric’ activation to ~ 2.7 μm, which is within the known optimal length range for force production of human quadriceps muscle (Walker and Schrodt 1974). Fibre length (Lo) was recorded as the distance between the innermost portion of the suture knots on either side, and fibre diameters were measured at three points along the fibre using a reticule on the microscope, and CSA was calculated assuming circularity. Chemical activation of single fibres was induced by transferring from the relaxing to pre-activating bath (reduced Ca2+-buffering capacity), and then to an activating solution (for all solutions, please see Mazara et al. (2021)). A ‘fitness’ contraction was first performed in pCa 4.2 to ensure the suture knots were not loose and to ensure fibre viability both visually and mechanically. After the fitness test, sarcomere length was rechecked and, if necessary, re-adjusted to 2.8 μm. The fibres were then transferred to an activating solution of pCa 5.0 and allowed to develop force for ~ 30 s. The highest force (Po) reached during the 30 s window was taken (Fig. 1C). Specific force (SF) was then defined by dividing Po by the fibre's CSA.

Following the Po assessment, a length step was induced to measure rate of force redevelopment (Brenner and Eisenberg 1986). This was done by rapidly shortening the fibre with a ramp of 10 Lo/s by 15% of Lo and then a rapid (500 Lo/s) restretch back to Lo (Fig. 1C, D). At saturating Ca2+ levels, the rapid shortening causes all cross-bridges to break, and then the restretch allows further dissociation of any remaining cross-bridges and redevelopment of force independent of Ca2+-dependent regulatory proteins at Lo. A mono-exponential equation, y = a (1 − e−kt) + b, was fit to the redevelopment curve to determine ktr.

Unloaded shortening velocity (Vo) was assessed using the slack-test method (Edman 1979). Three separate slack tests were performed on each fibre, shortening by 5%, 10%, and 15% Lo. After the fibre’s force plateaued, a rapid length step was performed, causing the force to drop to zero. Force then redeveloped over time proportional to the amount of shortening. The fibre was then transferred back to the relaxing bath where length was returned to Lo and SL was rechecked and reset to ~ 2.8 μm if necessary. The data resulting from the three slack tests were plotted and a linear regression was performed with the slope of the resulting line taken as Vo. At the end of all tests, if a single fibre’s Po fell by > 10%, it was excluded from analysis. All the above-mentioned experiments were completed at 16 °C.

Following mechanical testing, fibres were removed from the testing apparatus and were fibre typed as reported previously (Mazara et al. 2021).

Data and statistical analysis

Of the 182 single muscle fibres (Young: n = 94, Old: n = 88) that were tested, a total of 122 fibres (Young: n = 67, Old: n = 55) remained intact and viable during mechanical testing and fibre typing thus were included in the analysis. After fibre typing via SDS-PAGE, 27 Type I and 40 Type II fibres in the young group, and 42 Type I and 13 Type II fibres in the old group were identified. To not violate independence in the statistical analysis, single fibre properties (i.e. CSA, Po, Vo, ktr, and SF) were averaged within each fibre type (Young: Type I ~ 3 ± 1 fibres, Type II ~ 5 ± 1 fibres; Old: Type I ~ 5 ± 2 fibres, Type II: ~ 2 ± 2 fibres) for every participant.

Normality of data was assessed within each group using the Shapiro–Wilk test. Independent samples t tests were performed to compare group means and effect sizes are reported as Cohen’s d. Linear regressions were performed to examine relationships between the variables of interest. Repeated measures ANOVA was used to assess differences in joint-level RTD across time epochs and effect sizes are reported as partial eta squared (\({\eta }_{p}^{2}\)). Spearman’s rank correlation (rs) and Mann–Whitney tests were performed for non-normally distributed data. Basic single fibre data (CSA, Po, Vo, ktr, and SF) were analysed and reported previously by our lab (Mazara et al. 2021). All statistical analyses were performed using SPSS version 28 (SPSS Inc., Chicago, Illinois) and an ɑ level of p ≤ 0.05 was used to determine statistical significance. All data are represented as mean ± SEM.

Results

Anthropometrics and physical activity

Height, mass, and physical activity levels did not differ significantly (p > 0.05) between young and old.

Joint-level knee extensor function

Voluntary activation was > 95% for all participants and was similar between groups (p = 0.33). Old were ~ 30% weaker than young (Fig. 2; p ≤ 0.05, d = 1.07). Old had lower RTD values than young during the 0–100 ms (~ 43%, p = 0.013, \({\eta }_{p}^{2}\) = 0.37) and 0–200 ms (~ 40%, p = 0.014, \({\eta }_{p}^{2}\) = 0.36) time epochs (Fig. 3), as well as peak RTD (young: 994.9 ± 88.1 Nm s−1, old: 670.8 ± 67.9 Nm s−1; p < 0.01, d = 1.51). Age related differences in peak RTD were abolished following normalization to peak MVC torque (young: 10.4 ± 1.7 s−1, old: 8.3 ± 0.4 s−1; p = 0.20). There was a strong, positive relationship between peak MVC torque and every RTD time epoch in young males (Fig. 4; r2 ≥ 0.90, p ≤ 0.001), such that the strongest individuals had the greatest RTD, while in old, this relationship was only present during the 0–200 ms time epoch (Fig. 4; r2 = 0.76, p < 0.01). Peak RTD was significantly related to peak MVC torque in old (r2 = 0.84, p < 0.001). When groups were collapsed, all RTD epochs (Fig. 4; r2 > 0.40, p < 0.01) and peak RTD (r2 = 0.42, p = 0.01) were related to peak MVC torque.

Fig. 2
figure 2

Peak MVC torque for young and old males. Open circles represent individual subject data. *Significant difference between groups (p ≤ 0.05). Data are displayed as mean ± SEM

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Fig. 3
figure 3

Joint-level voluntary RTD for young and old males across several time epochs: 0–30, 0–50, 0–100, and 0–200 ms. *Significant difference between groups at the same epoch (p ≤ 0.05). Circles represent individual data points. Data are displayed as mean ± SEM

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Fig. 4
figure 4

Explained variance between peak MVC torque and joint-level voluntary RTD across several time epochs: 0–30, 0–50, 0–100, and 0–200 ms for young and old males. Young: solid black line, old: solid light grey line, grouped: dashed dark grey line. * and ‡significant relationship between variables for young and grouped, respectively (p ≤ 0.001), †significant relationship between variables for old (p ≤ 0.001). See supplemental Fig. 1 for individual scatter plots

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Peak twitch torque (young: 33.9 ± 3.7 Nm, old: 22.7 ± 2.5 Nm) and twitch RTD (young: 601.7 ± 64.2 Nm s−1, old: 411.1 ± 44.3 Nm s−1) were both ~ 32% lower in old compared to young males (p < 0.03, d = 1.25 and 1.22, respectively). Twitch RTD normalized to peak twitch torque was similar between groups (young: 17.8 ± 0.5 s−1, old: 18.2 ± 0.5 s−1; p = 0.5). In young males, there was a strong, positive relationship between twitch RTD and voluntary RTD during the 0–30, 0–50, and 0–100 ms (r2 ≥ 0.70, p ≤ 0.01) time epochs, while also displaying trends towards significance during the 0–200 ms (p = 0.053) time epoch (Fig. 5). There were no relationships between twitch and voluntary RTD in the old group (p > 0.20). When groups were collapsed, twitch RTD was related to all voluntary RTD epochs (Fig. 5; r2 > 0.20, p < 0.02) and peak RTD (r2 = 0.42, p = 0.01).

Fig. 5
figure 5

Explained variance between twitch RTD and joint-level voluntary RTD across several time epochs: 0–30, 0–50, 0–100, and 0–200 ms for young and old males. Young: solid black line, old: solid light grey line, grouped: dashed dark grey line. *Significant relationship between variables for young (p ≤ 0.001). ‡Significant relationship between variables for grouped (p ≤ 0.02). See supplemental Fig. 1 for individual scatter plots

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Single muscle fibre and joint-level relationships’

The results for basic single muscle fibre characteristics were previously reported by our laboratory (Mazara et al. 2021); please see Table 1. There were no relationships between any of the single muscle fibre properties (i.e. CSA, Po, Vo, ktr, and SF) and electrically evoked twitch RTD (p > 0.05) in either the young or old group (p > 0.05), when groups were collapsed Type II CSA was related to twitch RTD (r2 = 0.36, p = 0.02). Also, there were no relationships between any of the single muscle fibre mechanical properties and any voluntary absolute RTD epochs or peak normalized RTD in young (p > 0.05). However, in old, Type II fibre Vo displayed a moderate, negative relationship with joint-level voluntary RTD at the 0–100 (p = 0.033) and 0–200 ms (p = 0.041) time epochs (Fig. 6). Further, there was a moderate, positive relationship between Type I SF and normalized peak RTD in the old group (rs = 0.81, p = 0.02). When groups were collapsed, Type II CSA was related to the 0–100 and 0–200 ms RTD epochs (r2 = 0.32, p = 0.03 and r2 = 0.37, p = 0.02, respectively), and peak MVC torque (r2 = 0.33, p = 0.03).

Table 1 Type I and Type II muscle fibre properties
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Fig. 6
figure 6

Explained variance between Type II fibre Vo and joint-level voluntary RTD across several time epochs: 0–30, 0–50, 0–100, and 0–200 ms for young (n = 40 fibres), old (n = 13 fibres) and grouped (n = 53 fibres). Young: solid black line, old: solid light grey line, grouped: dashed dark grey line. †Significant relationship between variables for old (p ≤ 0.05). See supplemental Fig. 4 for individual scatter plots

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With respect to single fibre and joint-level RTD, there was a strong trend in the old group for ktr of Type I fibres to be associated with joint-level voluntary RTD during the earliest time epochs (0–30 ms: r2 = 0.48; p = 0.055 and 0–50 ms: r2 = 0.49; p = 0.052) (Fig. 7).

Fig. 7
figure 7

Explained variance between Type I ktr and joint-level voluntary RTD across several time epochs: 0–30, 0–50, 0–100, and 0–200 ms for young (Type I: n = 27 fibres, Type II: n = 40 fibres), old (Type I: n = 42 fibres, Type II: n = 13 fibres), and grouped (Type I: n = 69 fibres, Type II: n = 53 fibres). Young: black lines and triangles, old: light grey lines and squares¸ grouped: dark grey lines and circles. Older adults trended (p = 0.052–0.055) towards significant relationships between joint-level RTD and Type I single fibre rate of force development at the earliest time epochs. Inset: Representations of individual data for Type I fibres at the 30 ms and 50 ms time epoch. See supplemental Fig. 5 for individual scatter plots

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Note: please see supplemental figures for all individual linear relationships between variables.

Discussion

The purpose of the present study was to build on past work from our laboratory (Mazara et al. 2021) and identify any relationships between single muscle fibre rate of force development and joint-level RTD. In line with our hypothesis, in older adults, specifically at the earliest time epochs, there appeared to be a relationship emerging (Fig. 7; p = 0.052–0.055) such that those individuals with faster Type I single fibre rate of force development also had the highest joint-level voluntary RTD. However, we did not observe this relationship between single muscle fibre and joint-level RTD for electrically stimulated contractions in young or old adults, which indeed brings into question the basic mechanisms underlying partitioning of RTD into various time epochs (Andersen and Aagaard 2006). While electrically evoked twitch contractions may be a good predictor of early RTD in young, this was not supported for old adults, and joint-level electrically evoked twitch contractions were not truly representative of ‘intrinsic contractile properties of the muscle’.

Joint-level function

As expected, older males were weaker and had a reduced capacity to produce torque rapidly during voluntary and electrically evoked contractions when compared to their younger counterparts (Callahan et al. 2015; Wu et al. 2021, 2016). These age-related impairments in rapid torque production are likely due to differences in strength between groups (Thompson et al. 2013; Thelen et al. 1996), especially when considering that group differences disappeared once peak voluntary RTD and twitch RTD were normalized (i.e. when controlling for strength). The differences in strength between groups is likely mediated by age-related reductions in muscle mass, and more specifically, reduction in overall Type II contractile content as observed in this study and others (Lexell and Downham 1992; Lexell et al. 1988; Grimby and Saltin 1983).

We observed that twitch RTD was related to voluntary RTD in young males during the earliest time epochs (< 100 ms), which supports previous findings (Andersen and Aagaard 2006). Andersen and Aagaard (2006) also found that twitch RTD explained more of the variance in the early phase of RTD (i.e. < 50 ms) than maximum strength (r2 =  ~ 30% vs ~ 20%, respectively), while the later phases (i.e. > 100 ms) were explained more by maximum strength than twitch RTD (r2=  ~ 50–80% vs 10%, respectively). Although our results from young males are similar to those from Andersen and Aagaard (2006), we failed to observe any relationship between twitch and voluntary RTD in old males. Indeed, only peak MVC torque was related to any RTD epoch (i.e. 0–200 ms) in old, which supports previous literature (Andersen and Aagaard 2006; Mirkov et al. 2004), and is likely due to the fact that this epoch is approaching the time required for maximal force production (i.e. ~ 200 ms) (Aagaard et al. 2002). The failure to observe any relationships between twitch properties and voluntary RTD in old may be explained by differences in motor unit recruitment during electrically evoked and voluntary contractions. Electrically evoked contractions should, in theory, maximally activate all targeted motor units, whereas voluntary contractions are characterized by motor unit recruitment occurring in an orderly fashion from the smallest (primarily Type I fibres) to the largest (primarily Type II fibres) according to Henneman’s size principle (Henneman 1957); ageing has been associated with delays in motor unit recruitment and/or slowing of motor unit firing rate, particularly at low relative force levels (Watanabe et al. 2016; Girts et al. 2020). Therefore, electrically evoked contractions may recruit motor units that are not recruited until much later during voluntary contractions in old males from the current study.

Although we observed a strong relationship between twitch RTD and voluntary RTD in young males, peak MVC torque (i.e. maximum strength) still explained more of the variance in voluntary RTD across all time epochs, regardless of age. The divergent results between the current study and Andersen and Aagaard (2006) may be potentially explained by differences in the participant’s voluntary activation levels during MVCs. Voluntary activation was not assessed in Andersen and Aagaard (2006), so it is not clear whether MVCs were indeed maximal, whereas we were able to confirm near maximum voluntary activation (> 95%) which may provide an explanation as to why voluntary RTD was explained more by twitch contractile properties rather than maximum strength in Andersen and Aagaard (2006), while maximum strength was more related to voluntary RTD in the current study.

Single fibre function

Our findings and comparisons to previous literature for measures of single fibre function have been discussed in detail previously (Mazara et al. 2021); therefore, the current discussion will focus primarily on the limited number of studies that have investigated the effect of age on single fibre ktr and Vo. To summarize, young and older males displayed similar ktr and Vo values across fibre types in the current study which parallels findings from Teigen et al. (2020). Conversely, Power et al. (2016) observed reductions in ktr and Vo in older compared to young males, and these discrepancies may be due to factors such as the temperature at which mechanical testing was performed (10 °C (Power et al. 2016) vs. 15 °C (Teigen et al. 2020) and 16 °C (Mazara et al. 2021)), differences in the methods of fibre typing (categorizing fibres as slow vs. fast based off Vo (Power et al. 2016) vs. determining MHC content via gel electrophoresis (Mazara et al. 2021; Teigen et al. 2020)), and participant training status (master athletes (Power et al. 2016) vs. healthy, independent living adults (Teigen et al. 2020; Mazara et al. 2021)). Although ktr and Vo were not measured, Miller et al. (2013) performed sinusoidal length perturbations of single muscle fibres from physical activity matched individuals to investigate the effects of age and sex on single fibre function. The authors found that ageing was associated with increased myosin attachment time and reduced phosphorylation of MHC II myosin regulatory light chain, but this was primarily driven by females. Although relatively few studies have assessed cross-bridge kinetics across age groups, some indicate an age-related reduction (Power et al. 2016), while others suggest they are preserved in males with similar physical activity levels (Mazara et al. 2021; Miller et al. 2013; Teigen et al. 2020).

Relating single fibre and joint-level function

A novel aspect of the present study was investigating the relationship between single muscle fibre intrinsic contractile properties and joint-level function in young and older males. Despite electrically evoked twitches being often used as indications of intrinsic muscle contractile properties, we failed to observe a relationship between any single fibre mechanical measure with twitch properties. However, Type II CSA and twitch RTD were related, but this was only after collapsing the groups. This observation supports the idea that rapid torque production is, in part, dependent upon Type II fibre CSA (Harridge et al. 1996).

Our method of testing single fibres (i.e. chemical activation of permeabilized fibres) may be partly responsible for the lack of relationships between single fibre mechanical measures and twitch properties. In the present study, we are able to precisely regulate Ca2+ and ATP concentrations that permeabilized fibres are exposed to, while other single fibre preparations, such as electrical stimulation of intact fibres requires propagation of an electrical signal across the fibre, triggering the release of Ca2+ for force production; however, this technique is very limited in humans due to methodological constraints (Olsson et al. 2020, 2015; Cheng and Westerblad 2017). Regardless, the chemical activation of permeabilized fibres performed in this study provided evidence that twitch properties are not related to myofibrillar contractile function, but rather other factors upstream from basic cross-bridge cycling. These factors may include, but are not limited to, velocity of action potential propagation (Andreassen and Arendt-Nielsen 1987), excitability of the transverse-tubules (Stephenson et al. 1998), rate of calcium release into the myoplasm (Baylor and Hollingworth 2012), and availability of actin binding sites (McKillop and Geeves 1993).

When relating single fibre properties to voluntary joint-level function, the only relationships observed in the current study were in the old group alone or when groups were collapsed. Type I SF was related to peak normalized RTD in old, suggesting that muscle quality of “slow” Type I fibres may become more important for rapid torque production with age. There was also a negative relationship between Type II Vo and joint-level voluntary RTD during the 0–100 and 0–200 time-epochs in the old group (Fig. 6) such that those individuals with the slowest RTD had faster single fibre unloaded shortening velocity—the mechanisms by which we are currently unable to resolve. Additionally, it should be noted that while we did not observe a significant relationship between single fibre ktr and joint-level voluntary RTD in the young group, there was a strong trend for ktr of Type I fibres to be related to early joint-level voluntary RTD (Fig. 7; 0–30 ms: p = 0.055, 0–50 ms: p = 0.052) in old. The lack of statistical significance in these relationships can be explained by our limited sample size and averaging multiple single fibre data points into a single value for the regression analysis—if each single muscle fibre was treated individually and not averaged, the relationships become statistically significant. Further, collapsing the groups revealed that Type II CSA was significantly related to the 0–100 and 0–200 ms RTD epochs, as well as peak MVC torque. The findings from our collapsed groups are similar to those of Callahan et al. (2015), who reported that Type II CSA was related to peak absolute RTD in older individuals with and without knee osteoarthritis. Results from the current study and Callahan et al. (2015) support the widely accepted belief that Type II (“fast”) fibres are vital for powerful movements (Harridge et al. 1996; Thorstensson et al. 1977; Coyle et al. 1979), but there may be an increased reliance on Type I fibre properties for rapid torque production with age.

Limitations and future directions

The primary limitation of the current study was the number of single muscle fibres included in our analyses. While extreme caution was taken during mechanical testing, approximately 33% of fibres tested were not included in analyses due to our criteria of discarding any fibres that peak force dropped by > 10% during testing leaving some interpretations trending but underpowered.

Future studies should also investigate the relationship between intrinsic contractile properties and joint-level muscle function in females. Although males and females display similar RTD when controlling for maximal strength (Folland et al. 2014; Hannah et al. 2012), it is unknown if RTD relies on the same physiological mechanisms (i.e. electrically evoked twitch properties or maximal strength). Cross-bridge kinetics are more affected by age-related impairments in females than males (Miller et al. 2013) so assessing the relationship between intrinsic contractile properties and joint-level RTD in young and older females may elucidate sex-specific mechanisms behind the reduction in RTD capacity.

Conclusion

Although our data support previous research that electrically evoked twitch properties are related to early voluntary RTD in young males, this relationship is not maintained in older males. In line with our hypothesis, in older adults, a relationship emerged such that those individuals with faster Type I single fibre rate of force development also had the highest joint-level voluntary RTD. However, we did not observe this relationship between single muscle fibre and joint-level RTD for electrically stimulated contractions in young or old adults, which indeed brings into question the basic mechanisms underlying partitioning of RTD into various time epochs. Based on these data, joint-level electrically evoked twitch contractions were not truly representative of ‘intrinsic contractile properties’ in terms of rate of force development at the cross-bridge level.