# Effects of aging on transmission from Ia afferents to α-motoneurons and presynaptic mechanisms across muscle lengths and contraction types

Yohan Betus, Julian Colard, Marc Jubeau, Anne-Sophie Boureau, Guillaume Chapelet, Christophe Cornu, Thomas Cattagni

<figure><img src="image_1.png" /></figure>

## To cite this version:

Yohan Betus, Julian Colard, Marc Jubeau, Anne-Sophie Boureau, Guillaume Chapelet, et al., Effects of aging on transmission from Ia afferents to α-motoneurons and presynaptic mechanisms across muscle lengths and contraction types. GeroScience, 2025, Online ahead of print. 10.1007/s11357-025-01958-2. [https://hal.science/hal-05381213](https://hal.science/hal-05381213)

HAL Id: hal-05381213

https://hal.science/hal-05381213v1

Submitted on 25 Nov 2025

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

<figure><img src="image_2.png" /></figure>

Distributed under a Creative Commons Attribution 4.0 International License
1 **TITLE**

2 Effects of aging on transmission from Ia afferents to α-motoneurons and presynaptic

3 mechanisms across muscle lengths and contraction types.

4 **AUTHORS**

5 Yohan Betus¹, Julian Colard¹, Marc Jubeau¹, Anne-Sophie Boureau², Guillaume Chapelet²,

6 Christophe Cornu¹ & Thomas Cattagni¹

7 **AFFILIATION**

8 1. Nantes Université, Movement - Interactions - Performance, MIP, UR 4334, F-44000

9 Nantes, France

10 2. Pôle de Gérontologie Clinique, Nantes Université, CHU Nantes, Nantes, France

11 **RUNING TITLE**

12 **CORRESPONDING AUTHOR**

13 Thomas Cattagni

14 Nantes Université, Movement - Interactions - Performance, MIP, UR 4334, F-44000 Nantes,

15 France

16 25 bis Boulevard Guy Mollet - BP 72206

17 44 322 Nantes cedex 3, France

18 Email: thomas.cattagni@univ-nantes.fr

19 ORCID: 0000-0002-6650-8339

20

21
22

ABSTRACT

23

24

25

26

27

28

29

30

31

32

33

34

35

36

37

38

39

Presynaptic inhibition of Ia afferents, particularly via primary afferent depolarization (PAD), plays a crucial role in modulating transmission from Ia afferents to α-motoneurons according to task demands. In young adults, PAD is enhanced during eccentric contractions and at longer muscle lengths, supporting adaptable motor control. However, aging is associated with reduced muscle spindle sensitivity, which may impair the regulation of presynaptic inhibition via PAD.

This study investigated whether aging alters the modulation of PAD according to contraction type and muscle length. Nineteen older adults (69 ± 4 years) and sixteen young adults (23 ± 2 years) participated in three experimental sessions. The maximal H-reflex (Hmax) and maximal M-wave (Mmax) were recorded in the soleus muscle to assess Ia-alpha motoneuron transmission (Hmax/Mmax). PAD was evaluated using the D1 inhibition method (HDI/Htest), while ongoing presynaptic inhibition was assessed via heteronymous facilitation (Hfac/Htest). Measurements were performed during isometric and eccentric contractions at intermediate (0°) and long (-15° dorsiflexion) muscle lengths. Young adults showed significant modulation of both HD1/Htest and Hfac/Htest across conditions, while older adults did not (p > 0.05). Similarly, Hmax/Mmax was reduced during eccentric contractions at intermediate muscle length in young but not in older adults. These results indicate that aging impairs regulation of PAD according to muscle mechanics, potentially contributing to deficits in sensorimotor control.

40

KEYWORDS

41

Eccentric, long muscle length, Ia afferent, H-reflex, presynaptic inhibition

42

43

44
45

# INTRODUCTION

Regulation in neural transmission from Ia afferent fibres to alpha motoneurons (Ia-motoneuron transmission) is essential for fine motor control, balance, posture, and coordinated movement (1–4). This regulation is influenced by various motor task constraints and characteristics, including muscle length, and contraction type, i.e. isometric, concentric or eccentric contraction (5–7). Aging is often associated with an impaired capacity to regulate Ia-motoneuron transmission in response to motor tasks (8–11), potentially leading to a decline in motor function (12,13). A deeper understanding of the mechanisms underlying age-related impairments in the regulation of Ia-motoneuron transmission according to contraction type and muscle length is essential for advancing our knowledge of motor dysfunction in older adults.

Ia afferent fibers are fast-conducting sensory axons from muscle spindles that detect changes in muscle stretch rate (14). Their discharge rate increases during muscle lengthening and sustained static muscle length (15). Within the spinal cord, Ia afferents form excitatory synaptic connections with alpha motoneurons (16). The spinal regulation of Ia afferent input involves presynaptic inhibition through primary afferent depolarization (PAD), mediated by GABA<sub>ergic</sub> interneurons. These interneurons, modulated by supraspinal and spinal inputs, enable fine-tuned control of Ia afferent transmission. In healthy young adults, PAD is modulated by various motor task constraints and is particularly high during eccentric contractions and at longer muscle lengths, where the firing rate of Ia afferent fibers increases significantly (5,17–20). This modulation leads to reduced Ia-motoneuron transmission (5–7,19).

Aging impacts muscle spindles through various morphological and functional changes, including capsule thickening (21) and a reduction in the number of intrafusal fibres (22). Muscle spindle sensitivity also declines, due to diminished responsiveness to stretching and changes in muscle length (23,24). As PAD regulation adapts to changes in Ia afferent discharge, it is
69 plausible that aging may attenuate the modulation of PAD regulation during muscle
70 lengthening, especially during eccentric contractions with changes in muscle length. Some
71 evidence suggests that aging impairs the capacity to modulate PAD according to motor tasks,
72 which are assumed to have different Ia afferent activities. For example, while young adults
73 adjust PAD based on contraction intensity, postural control, or specific motor tasks, older adults
74 often exhibit diminished PAD regulation (8,9,11,25). However, further direct evidence is
75 needed to clarify how aging influences PAD regulation across different contraction types and
76 muscle lengths, and how these changes ultimately impact Ia-motoneuron transmission.

77 This study investigated whether aging alters PAD regulation based on contraction type and
78 muscle length. Using the Hoffmann reflex, we assessed PAD and Ia-motoneuron transmission
79 in the soleus muscle of young and older adults. Measurements were done at two muscle lengths
80 (intermediate and long) and during two contraction types (isometric and eccentric). We
81 hypothesized that, unlike young adults who modulate PAD based on contraction type and
82 muscle length, older adults exhibit diminished or no PAD modulation, which is associated with
83 impaired presynaptic regulation of Ia-motoneuron transmission.

84
87 A total of 35 participants participated in the study :16 young adults (8 males and 8 females; age:
88 23 ± 2 years; height: 175 ± 4 cm; weight: 70 ± 13 kg; BMI: 23.0 kg·m⁻²) and 19 older adults (8
89 males and 11 females; age: 69 ± 4 years; height: 168 ± 7 cm; weight: 64 ± 10 kg; BMI: 22.8
90 kg·m⁻²). Individuals were classified as part of the “young” group if they were between 18 and
91 35 years old, and as part of the "older" group if they were between 60 and 80 years old (26).

92 Exclusion criteria included any neurological or neuromuscular disorders, musculoskeletal
93 injuries that could impair torque production, and the use of medications known to affect the
94 nervous system. During the medical screening, we ensured that participants were 1) physically
95 active according to WHO guidelines (i.e., at least 150 minutes per week of moderate-intensity
96 aerobic activity or 75 minutes per week of vigorous-intensity aerobic activity) and 2) not
97 engaged in any resistance training. These inclusion criteria were applied equally to both cohorts
98 to minimize potential bias from resistance training that may influence neuromuscular function.

99 In addition, all older participants underwent a medical screening with a geriatrician, including
100 the WHO ICOPE (Integrated Care for Older People) assessment, to ensure that only healthy
101 older adults were included. Participants were instructed to refrain from engaging in strenuous
102 exercise for 48 hours prior to the testing sessions. Ethical approval for the project was obtained
103 from the Ethics Committee on Non-Interventional Research of Nantes University (No.
104 21022024-3). All procedures conducted in this study adhered to the requirements of the
105 Declaration of Helsinki (last revised in 2011). Data from the young adult group were previously
106 reported in an earlier study (18).
108

All 35 participants were engaged in two experimental sessions. All younger participants and 11 older participants (among the 19) also participated in a third session. Each experiment was separated by a minimum of 24 hours. During each experiment, assessments of contraction type (isometric and eccentric contractions) and muscle length (intermediate and long muscle lengths) were randomly assigned to avoid potential biases stemming from the order of testing procedures.

109

110

111

112

113

114

In the first experimental session (Experiment A, duration: 2 h 30), we assessed soleus fascicle length and the recruitment curves for the soleus H-reflex and M-wave for each contraction type and soleus length. The second experimental session (Experiment B, duration: 2 h) examined post-activation depression by PAD using the D1 method. In the third experimental session (Experiment C, duration: 2 h), heteronymous Ia facilitation was assessed. The order of Experiments B and C was randomized across participants. All data were collected from the right leg of the participants.

115

116

117

118

119

120

121

At the end of each experimental session, two maximal voluntary concentric and isometric dorsiflexions were performed to assess maximal dorsiflexor activity (3 seconds of duration, interspaced by 1-minute rest).

122

123

124

## Experimental set-up

125

### Mechanical data

126

The participants were seated with the trunk inclined at 30°, the hip flexed at 85° (0° representing the anatomical position), and the knee joint fully extended at 0° (representing full extension).

127

128

Torque values were collected using an isokinetic dynamometer (Biodex 3, Shirley, NY, USA) (Figure 1). A Biodex ergometer enabled the instantaneous recording of muscle torque at a constant velocity of 20°·s⁻¹. The output signal from the dynamometer was collected at 2 kHz

129

130
131 using a commercial acquisition system (CED Power 1401-3A, Cambridge Electronic Design;
132 Cambridge, UK) and was displayed and stored with Signal 7 software (Cambridge Electronic
133 Design; Cambridge, UK). The right foot was strapped to the footplate, while the trunk and
134 thighs were secured to the seat of the isokinetic dynamometer. The ankle angle was modulated
135 according to experimental conditions. During isometric contractions, the ankle angle was set at
136 0° (neutral position) and -15° (dorsiflexion) for intermediate and long muscle lengths,
137 respectively. During eccentric contractions, the ankle range of motion was set at 30°. The ankle
138 angle moved from 15° to -15° and from 0° to -30° for intermediate and long muscle lengths,
139 respectively. Throughout the three experiments, the dynamometer movement cycles lasted 5
140 seconds, including 1.5 seconds for performing the movement, 1.5 seconds to return to the initial
141 position, and 1 second of rest before the next cycle. Within the latter period, participants
142 performed a 1-second submaximal isometric plantarflexion as a pre-activation contraction to
143 standardize thixotropic effects across conditions (27).

144

145

146

147

148

<figure><img src="image_3.png" /></figure>

149

<caption>**Figure 1 - Participant positioning on the dynamometer for plantar flexor testing.** Trunk reclined at 30°, hip flexed at 85°, knee fully extended (0° = full extension). Torque recorded with an isokinetic dynamometer.</caption>

150

151

152

153
154 Electromyography (EMG)

155 EMGs were recorded from the soleus, tibialis anterior and vastus lateralis muscles using pairs
156 of self-adhesive surface electrodes (Kendall™ H124SG; Covidien, Dublin, Ireland) configured
157 in a bipolar arrangement with a 30-mm inter-electrode distance. The soleus electrodes were
158 placed 2 cm below the muscle-tendon junction of the gastrocnemii. For the tibialis anterior,
159 electrodes were placed on the muscle belly parallel to the longitudinal axis of the muscle,
160 located one-third of the distance between the head of the fibula and the tip of the medial
161 malleolus. The vastus lateralis electrodes were positioned two-thirds along the line from the
162 anterior superior iliac spine to the lateral side of the patella. Prior to electrode application, the
163 skin was shaved, gently abraded, and cleaned with alcohol. The signals were sampled at 5 kHz,
164 amplified 1000x, and bandpass filtered from 5–500 Hz using a bio-amplifier (g.BSamp 0201a;
165 Guger Technologies, Shieldberg, Austria).

166 Percutaneous electrical nerve stimulation

167 The electrophysiological responses (i.e., H reflex and M wave) of the soleus were evoked by
168 percutaneous electrical stimulation of the posterior tibial nerve using a 1-ms single rectangular
169 pulse at high voltage (400 V), delivered by a stimulator (Digitimer, model DS8R Biphasic
170 Constant Current Stimulator, Hertfordshire, UK). A self-adhesive electrode (Kendall™
171 H124SG; Covidien, Dublin, Ireland) was used as a cathode and was placed in the popliteal
172 fossa. The anode electrode (5 × 10 cm, Medicompex SA, Ecublens, Switzerland) was positioned
173 on the anterior surface of the knee, below the patella. Once positioning was confirmed, the
174 cathode electrode was firmly secured to this site with a strap and tape, as recommended (28).
175 To investigate PAD (D1 method), the common peroneal nerve was stimulated using a DS7R
176 Current Stimulator (Digitimer, model DS7R Biphasic Constant Current Stimulator,
177 Hertfordshire, UK), with the cathode electrode placed close to the head of the fibula and the
anode electrode near the medial part of the tibial head. For assessment of heteronymous Ia facilitation, the femoral nerve was stimulated with the cathode positioned over the nerve in the femoral triangle and the anode placed over the greater trochanter. The location of the stimulation electrode was identified to obtain an M wave in the vastus lateralis, which is associated with an upward movement of the patella. The motor threshold for stimulation of the common peroneal and femoral nerves was determined as the lowest intensity that evoked at least three M waves out of five stimulations. To ensure that the electrophysiological recordings reflected neurophysiological modulations rather than methodological artefacts, several controls were implemented. Stimulation was automatically triggered at fixed joint angles (0° and -15°) and verified by ultrasound to correspond to distinct fascicle lengths. The maximal M-wave (Mmax) remained stable across sessions and conditions (p > 0.05), and all responses were normalized to Mmax. In addition, MatHmax/Mmax and Htest/Mmax ratios confirmed consistent recruitment of the α-motoneuron pool across conditions. These procedures ensured that the observed effects were not influenced by electrode displacement, limb movement, or differences in muscle architecture.

## 193 Ultrasound recording

Soleus fascicle length was measured using a Mach30 ultrasound scanner (Hologic-Supersonic Imagine, Aix-en-Provence, France) with B-mode image acquisition and a linear probe (SL18-5, 7.5 MHz central frequency, Hologic-Supersonic Imagine, Aix-en-Provence, France).

Participants were seated in the isokinetic dynamometer with their trunk inclined 20° backward from the vertical and their knee fully extended. The lateral malleolus was aligned with the axis of rotation of the dynamometer. Images were captured from +30° (plantarflexion) to -30° (dorsiflexion) during eccentric submaximal contractions, with 0° representing the anatomical
position. Fascicle length changes were evaluated during submaximal eccentric contractions (50% of maximal EMG<sub>RMS</sub>) at a constant angular velocity of 20 deg·s<sup>-1</sup>.

Proximal and distal insertions of the soleus muscle were identified, positioning the probe just distal to the myotendinous junction of the medial gastrocnemius muscle to ensure clear visibility of the soleus fascicles in this region. Despite the multipennate structure of the soleus muscle, a study on its three-dimensional architecture demonstrated no significant differences in fascicle length across its four compartments: medial-anterior, lateral-anterior, medial-posterior, and lateral-posterior (29). Therefore, the chosen probe position provides a fascicle length measurement representative of the entire soleus muscle. The probe was secured in a foam holder and firmly attached to the leg with tape. To condition the muscle, five ankle angle rotations ranging from 30° (plantarflexion) to -30° (dorsiflexion) were performed back and forth (30). B-mode ultrasound images were continuously recorded through three acquisitions during eccentric contractions at 50% of maximal EMG<sub>RMS</sub>, covering the full range of motion to assess the architecture of the soleus muscle (Fig. 2). The acquired ultrasound images were sampled at 50 Hz and processed using a customized MATLAB script (UltraTrack software; 5-2-2, Digitizing Services, UK) (31).

<figure><img src="image_4.png" /><figcaption>**Figure 2.** Examples of two ultrasound images in the sagittal plane during eccentric contraction at two different ankle angles in the soleus of one elderly participant. This image was used to calculate</figcaption></figure>
219
220

soleus fascicle length (blue dashed line) from the visible insertion of the fibres between the deep and superficial aponeuroses (yellow lines).

221

*Experimental protocol*

222

**Experiment A**

223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240

Experiment A was conducted in two parts. In the first part, three maximal voluntary isometric contractions (MVCs) at 0° were performed to determine the maximal root mean square (RMS) value of soleus EMG. The maximum soleus EMG<sub>RMS</sub> activity recorded during isometric MVCs was used to establish the activation level targeted, i.e., 50% of maximal EMG<sub>RMS</sub> provided to the participants during the study. This submaximal activation level was selected to avoid inducing muscle damage and/or neuromuscular fatigue during and between experimental sessions. Moreover it is functionally relevant because it is quite close to the loads required in activities of daily living (32–34). In this phase, participants were instructed to contract their plantar flexors to move and maintain their soleus EMG<sub>RMS</sub> biofeedback at the target corresponding to 50% of the maximal EMG<sub>RMS</sub>. The biofeedback represented the RMS EMG signal, which was computed in real-time by a digital computing channel. This channel instantaneously calculated the RMS EMG signal with an integration time of 500 ms. This method ensured consistent muscle activation levels across all conditions (5,6,18). In addition, to ensure that the modulation of our neurophysiological variables reflected neural activity rather than peripheral or recording factors, we measured M<sub>max</sub> in each condition. M<sub>max</sub> remained stable across all conditions (p > 0.05). Furthermore, all trials were visually inspected to confirm the absence of artefacts, and any trials showing obvious movement contamination were excluded from analysis. Subsequently, soleus fascicle length was measured.

241
242

The second part of the experiment aimed to obtain maximal H reflex (H<sub>max</sub>) and maximal M
wave (M<sub>max</sub>) for both contraction types (eccentric and isometric) at each muscle length
(intermediate and long) in soleus. Accordingly, we determined the soleus H reflex and M wave
recruitment curves during plantarflexion at 50% of maximal EMG<sub>RMS</sub> for each contraction type
and soleus length. This resulted in a total of four recruitment curves per participant (Fig. 3). For
each recruitment curve, the stimulation intensity was gradually increased in 2-mA and 3-mA
increments for young and older individuals, respectively. The H reflex threshold to the
stimulation intensity was where no further increase in M wave amplitudes was observed
between two stimulations (i.e., M wave plateau). We chose a greater increment in older adults
compared to younger adults because the relationship between the amplitude of the evoked
potentials and stimulation intensity is less steep in older adults. Even with the different
increments used between young and older subjects, the M<sub>atHmax</sub>/H<sub>max</sub> ratio remained consistent
across conditions and groups, allowing for a comparison of neural mechanisms rather than
recording and stimulation conditions (35). Finally, supramaximal stimulations were delivered
at 120% of this stimulation intensity to ensure the recording of soleus M<sub>max</sub>. Five electrical
nerve stimulations were delivered at each stimulation intensity with an interstimulus interval of
5 seconds (4).
<figure><img src="image_5.png" /><figcaption>Figure 3. Recruitment curves of the soleus H-reflex and M-wave according to muscle length and contraction type in a representative participant from the older group (panel A) and a participant from the younger group (panel B).</figcaption></figure>

Electrical tibial nerve stimulations were delivered at ankle angles of 0° and -15° (representing the midpoint of each range of motion) for intermediate and long muscle lengths, respectively.

Stimulation of the tibial nerve was driven by the CED acquisition system. During eccentric
contractions, stimulations were automatically triggered when the ankle angle passed 0° (intermediate) or -15° (long).

## 267 Experiment B

To investigate presynaptic modulation by PAD, we employed the D1 method as described by Mizuno et al. (1971). This technique involves eliciting an electrical volley on the nerve supplying the α-motoneuron pool of the antagonist muscle, thereby activating the Ia terminals and resulting in depression of the H reflex (36). The H reflex of the soleus was conditioned using a stimulus applied to the fibular nerve, activating GABA<sub>ergic</sub> interneurons responsible for the PAD of Ia afferents from the soleus muscle (37). To evoke the conditioned H reflex of the soleus (H<sub>D1</sub>), a train of three 1-ms stimulations at 300 Hz was applied to the common peroneal nerve, with an intensity set at 120% of the stimulation intensity at the motor threshold of the tibialis anterior. The interval between the first stimulus of the train and the soleus H reflex test (H<sub>test</sub>) was 21 ms (38,39). A total of 20 H<sub>D1</sub> and 20 H<sub>test</sub> responses were randomly evoked across all conditions (eccentric and isometric contractions at intermediate, and long muscle lengths) (5).

## 280 Experiment C

A limitation of the D1 method is that changes in PAD activity may be contaminated by potential occlusion or reflex gain phenomena in the reflex loop. To avoid this misinterpretation, it is recommended to compare the results obtained with the D1 method to those obtained with an independent method that assesses ongoing post-activation depression with heteronymous Ia facilitation (5,16). To investigate heteronymous Ia facilitation, we employed the method of Hultborn et al. (1987), which evaluates the monosynaptic facilitatory effect of femoral nerve stimulation on the amplitude of the soleus H reflex. To minimize contamination from
polysynaptic inputs, femoral nerve stimulation was delivered following tibial nerve stimulation due to the shorter neural pathway of the heteronymous Ia afferent pathway compared to the homonymous pathway (40). Considering the average duration of a synaptic event (0.5 to 1 ms), we systematically assessed the onset of facilitation in 1-ms increments to mitigate the influence of polysynaptic excitatory inputs (41). For each participant, the onset of H reflex facilitation in response to femoral nerve stimulation was determined by adjusting the delay between the test (tibial nerve) and conditioning (femoral nerve) stimuli in 1-ms increments, ranging from 9 ms (test stimulus preceding conditioning stimulus by 9 ms) to 1 ms (test stimulus preceding conditioning stimulus by 1 ms). The conditioning stimulation intensity was set at 130% of the motor threshold, and at least five trials were averaged for each delay (42). A total of 20 H<sub>fac</sub> and 20 H<sub>test</sub> responses were randomly evoked in all conditions (eccentric and isometric contractions at intermediate and long muscle lengths).

To determine non-conditioned H reflexes, the H<sub>test</sub> was based on the smallest H<sub>max</sub> amplitude measured across the four experimental conditions (Fig. 4). This approach ensures that each condition is subjected to the same H<sub>test</sub> size, allowing for accurate comparisons between the different experimental conditions. Before testing each condition, soleus H<sub>test</sub> was adjusted to ensure a consistent amplitude across conditions, as the number and/or origin of the motoneurons recruited may vary with stimulation intensity (43). Each condition was evaluated in a randomized order, with a 5-minute rest period between each tested condition.

307

308

309
310

311

312

313

314

315

316

317

318

319

320

321

322

<figure><img src="image_6.png" /><figcaption>Figure 4. Sigmoidal fittings of the soleus recruitment curve for the H-reflex (blue circles) and M-wave (red squares) to determine optimal amplitudes and stimulation intensities for post-activation depression (PAD) assessment, as well as H<sub>slope</sub> (blue line) and M<sub>slope</sub> (red line) evaluation for a representative participant. H<sub>max</sub>: maximal H-reflex; H<sub>test</sub>: test H-reflex for PAD assessment; M<sub>max</sub>: maximal M-wave; M<sub>atHmax</sub>: M-wave corresponding to H<sub>max</sub>; H<sub>slope</sub>: slope of the ascending portion of the H-reflex curve; M<sub>slope</sub>: slope of the M-wave curve.</figcaption></figure>

323

Data analysis

Evoked potentials

324

325

326

327

For each combination of contraction type and muscle length, the mean peak-to-peak amplitude of the following soleus evoked potentials were calculated over all trials: Hmax, Mmax, MatHmax, HD1, Hfac, and Htest. All ratios of evoked potentials were calculated for each experimental condition.

328

329

330

331

332

333

To estimate the effectiveness of Ia afferents in activating α-motoneurons, we calculated the Hmax/Mmax ratio. Hmax/Mmax is commonly used as an indicator of the maximal proportion of α-motoneurons recruited via the Ia afferent pathway. To control the stability of stimulation conditions in each condition, the M wave elicited concomitantly with Hmax (MatHmax) – a small fraction of Mmax (MatHmax/Mmax) – was measured and analysed (35). In addition, the slope of the ascending limb of the recruitment curve reflects the rate of motoneuron recruitment as a
334

function of increasing Ia input to the motoneuron pool and is generally used to examine H reflex gain (1,44). This reflex gain serves as a complementary measure, more closely linked to transmission efficiency between Ia afferents and α-motoneurons. By employing this approach, the influence of peripheral factors on the results is minimized, ensuring the accuracy and reliability of the findings (45). The sigmoidal slopes for both H reflex (Hslope) and M wave (Mslope) were determined by fitting the relationship between the amplitude of H reflex in the ascending part of the recruitment curve or M wave in entire recruitment curve by stimulation intensity (46). The Hslope/Mslope ratio was then calculated.

335

336

337

338

339

340

341

342

To examine presynaptic modulation by PAD using the D1 method, we calculated the H<sub>D1</sub>/H<sub>test</sub> ratio. A higher H<sub>D1</sub>/H<sub>test</sub> ratio is associated with lower post-activation depression by PAD (36).

343

344

To examine heteronymous Ia facilitation, we calculated the H<sub>Fac</sub>/H<sub>test</sub> ratio. A higher H<sub>Fac</sub>/H<sub>test</sub> ratio is associated with greater heteronymous Ia facilitation (40).

345

346

The H<sub>test</sub>/M<sub>max</sub> ratios obtained in the study of post-activation by PAD and heteronymous Ia facilitation were used to ensure that the amplitude of the non-conditioned H reflex remained consistent across the four experimental conditions. Hence, we controlled such that approximatively the same amplitude of α-motoneuron pool was investigated through experimental conditions.

347

348

349

350

351

*EMG activity during voluntary contractions*

352

The RMS of the soleus EMG signals over a 500-ms period preceding stimulation was normalized to the corresponding M-wave amplitude (Mmax) (EMGRMS/Mmax) for all experimental conditions. This normalization procedure was implemented to ensure that the EMG activity of the soleus (agonist) remained consistent across conditions. Similarly, the RMS of the TA EMG signal over the same time period preceding stimulation was calculated to assess

353

354

355

356
357

coactivation across all conditions. To quantify the level of coactivation in isometric contractions, TA EMG<sub>RMS</sub> during submaximal isometric contractions was divided by the peak TA EMG<sub>RMS</sub> obtained during the dorsiflexors MVC. TA RMS during eccentric plantar flexions was normalized to the maximal concentric dorsiflexion. TA EMG<sub>RMS</sub> was calculated across all trials within each experimental condition.

358

359

360

361

362

*Changes in Fascicle Length*

363

364

365

366

367

368

Ultrasound images were compiled to track the length of the soleus fascicles using a custom MATLAB script that enabled the tracking of fascicle behaviour during eccentric submaximal contractions. Fascicle length was measured at two distinct angles: 0° (intermediate muscle length), and -15° (long muscle length). A single fascicle was tracked at each angle. In cases where fascicles were not fully visible, fascicle length and the deep aponeurosis were extrapolated linearly to estimate the complete fascicle length.

369

*Statistical analysis*

370

371

372

373

374

375

376

377

378

379

All descriptive statistics presented in the text, tables and figures are expressed as mean values ± standard deviation. The significance level for all analyses was set at P < 0.05. Normality of the data and homogeneity of variances were verified and validated using the Shapiro-Wilks W test and Levene's test, respectively. Two-factor ANOVAs [muscle length (intermediate vs. long) × age (young vs. old)] were conducted to assess changes in fascicle length at different ankle angles (0° and -15°) during eccentric submaximal contractions. Three-factor ANOVAs [contraction type (eccentric vs. isometric) × muscle length (intermediate vs. long) × age (young vs. old)] with repeated measures on contraction type and muscle length were performed to compare soleus $M_{atHmax}/M_{max}$, $H_{max}/M_{max}$, $H_{slope}/M_{slope}$, $H_{D1}/H_{test}$, and $H_{Fac}/H_{test}$ across conditions. Three-factor ANOVAs [contraction type (eccentric vs. isometric) × muscle length
(intermediate vs. long) × experiment (experiment B vs. experiment C)] with repeated measures on contraction type, muscle length, and experiment were conducted to compare H<sub>test</sub>/M<sub>max</sub>. Similarly, four-factor ANOVAs [contraction type (eccentric vs. isometric) × muscle length (intermediate vs. long) × experiment (experiment A vs. experiment B vs. experiment C) × age (young vs. old)] with repeated measures were used to compare soleus M<sub>max</sub> and EMG<sub>RMS</sub>/M<sub>max</sub> for soleus, as well as tibialis anterior coactivation. When significant interactions were found, a post-hoc Holm test was performed. Statistical analyses were performed using JASP (Version 0.17.2.1, Amsterdam, NED) and GraphPad Prism software (Version 10.2.1; GraphPad Software Inc., San Diego, CA, USA).

## 389 RESULTS

### 390 Changes in fascicle length with ankle angle

Changes in fascicle length were calculated to ensure that the muscle lengths associated with the angle of electrical nerve stimulation were distinct. As shown in figure 5, muscle length at -15° of ankle angle was 15.1% greater than at 0° (P < 0.001), with no significant difference between young and older participants (p = 0.718).

395

396

397

398

399

400

<figure><img src="image_7.png" /></figure>
**Figure 5.** Changes in soleus muscle fascicle length during 50% submaximal eccentric contraction across ankle angles. Data are presented as mean ± standard deviation with individual values shown. Young adults are represented in blue with filled circles, while older adults are represented in orange with open circles. * Significant difference for ankle angle effect at P < 0.001.

EMG activity

We compared soleus EMG<sub>RMS</sub>/M<sub>max</sub> between eccentric and isometric contractions at intermediate and long muscle lengths to ensure consistent agonist muscle activation across experimental conditions. No significant effect was found (all P values > 0.05) (Table 1). However, an age effect was observed (P < 0.001), with young participants displaying a 33% higher soleus 50% EMG<sub>RMS</sub>/M<sub>max</sub> ratio than older participants, regardless of muscle length or contraction type.

Tibialis anterior coactivation during plantar flexion was assessed to determine if antagonist activation remained consistent across all experimental conditions. No main effects or interactions were found for tibialis anterior coactivation levels (all P values > 0.05) (Table 1). However, an age effect was observed (P < 0.001), with older participants showing approximately 1.18% lower coactivation than younger participants. The mean tibialis anterior coactivation was around 10.10% in younger participants and 8.83% in older participants during all plantar flexor conditions.

<table><thead><tr><th rowspan="2"></th><th colspan="2">Young</th><th colspan="2">Old</th></tr><tr><th>Intermediate</th><th>Long</th><th>Intermediate</th><th>Long</th></tr></thead><tbody><tr><td><strong>Eccentric</strong></td><td></td><td></td><td></td><td></td></tr><tr><td>SOL</td><td></td><td></td><td></td><td></td></tr><tr><td>EMG<sub>RMS</sub>/M<sub>max</sub></td><td>0.025 ± 0.006</td><td>0.026 ± 0.006</td><td>0.016 ± 0.006</td><td>0.017 ± 0.006</td></tr><tr><td>Tibialis anterior</td><td></td><td></td><td></td><td></td></tr><tr><td>Coactivation (%)</td><td>10.269 ± 2.571</td><td>10.124 ± 1.510</td><td>8.818 ± 0.937</td><td>8.850 ± 1.071</td></tr><tr><td><strong>Isometric</strong></td><td></td><td></td><td></td><td></td></tr><tr><td>SOL</td><td></td><td></td><td></td><td></td></tr><tr><td>EMG<sub>RMS</sub>/M<sub>max</sub></td><td>0.024 ± 0.005</td><td>0.025 ± 0.008</td><td>0.018 ± 0.007</td><td>0.018 ± 0.006</td></tr><tr><td>Tibialis anterior</td><td></td><td></td><td></td><td></td></tr><tr><td>Coactivation (%)</td><td>10.139 ± 1.492</td><td>9.891 ± 1.909</td><td>8.782 ± 1.058</td><td>8.838 ± 1.510</td></tr></tbody></table>
420

421

422

423

424

425

426

427

428

429

430

431

432

433

434

435

436

437

438

439

440

441

442

443

444

445

446

447

**Table 1. Effect of muscle length, contraction type and age on soleus and tibialis anterior EMG-RMS activity.** Data are pooled across the four experimental conditions and presented as mean ± standard deviation. EMG, electromyography; SOL, soleus; $M_{max}$, maximal M wave; RMS, root mean square.

Presynaptic modulation by PAD (D1 method)

SOL $H_{D1}/H_{test}$ was assessed to estimate the level of PAD. Figure 6A shows raw traces of the conditioned ($H_{D1}$) and non-conditioned ($H_{test}$) H reflexes measured during isometric and eccentric contractions at intermediate and long muscle lengths in a representative older participant.

A significant three-way interaction was observed between contraction type, muscle length, and age ($F(1, 33) = 4.314, p = 0.046, \eta p^2 = 0.116$), suggesting differential modulation of $H_{D1}/H_{test}$ across age groups. In older participants, $H_{D1}/H_{test}$ was not significantly influenced by muscle length or contraction type ($p > 0.05$), indicating an impaired capacity for modulation of PAD in response to these factors. This lack of adaptation contrasts with younger participants, where both muscle length and contraction type had a significant impact.

In younger participants, $H_{D1}/H_{test}$ decreased by 13.4% during isometric contractions at long compared to intermediate muscle lengths ($p < 0.001$). This reduction was more pronounced during eccentric contractions (29.3%) ($p < 0.001$). Additionally, contraction type influenced PAD, with a 25.3% lower $H_{D1}/H_{test}$ during eccentric compared to isometric contractions at long muscle lengths ($p < 0.001$). Age-related differences were particularly evident during eccentric contractions at long muscle lengths, where $H_{D1}/H_{test}$ was 28.9% higher in older compared to younger participants.

444

445

446

447
<figure><img src="image_8.png" /><figcaption>**Figure 6.** Modulation of post-activation depression via primary afferent depolarization in relation to muscle length, contraction type, and age. (A) Representative traces of the test H reflex (H<sub>test</sub>) and conditioned soleus H reflex (H<sub>D1</sub>) evoked by fibular nerve stimulation preceding posterior tibial nerve stimulation at a 21 ms conditioning-test interval during isometric and eccentric contractions at intermediate and long muscle lengths. (B) Mean absolute data and individual values are shown as mean ± standard deviation. H<sub>D1</sub> and H<sub>test</sub> are presented as a ratio during isometric and eccentric contractions at intermediate and long muscle lengths. Young adults are represented in blue with filled circles, while older adults are represented in orange with open circles. The electrical intensity for H<sub>test</sub> was normalized across all contractions for each muscle length. Abbreviations: H<sub>D1</sub>, conditioned soleus H reflex; H<sub>test</sub>, non-conditioned soleus H reflex; Ecc, eccentric; Iso, isometric. *Significantly different from eccentric contraction type.* # *Significantly different by age.*</figcaption></figure>

448

449

450

451

452

453

454

455

456

457

458

459

460

461

462

463

464

465

466

467

468

469

470

471

472

473

474

475

476

477

478

479

480

481

482

483

484

485

486

487

488

489

490

491

492

493

494

495
497 SOL $H_{\text{Fac}}/H_{\text{test}}$ was assessed to estimate the level of heteronymous Ia facilitation. Figure 7A
498 illustrates raw traces showing the conditioned ($H_{\text{Fac}}$) and non-conditioned ($H_{\text{test}}$) H reflex
499 measured during isometric and eccentric contractions at intermediate and long muscle lengths
500 in a representative older participant.

501 A significant three-way interaction was found between age, contraction type and muscle length
502 for $H_{\text{fac}}/H_{\text{test}}$ ($F(1, 21) = 18.557, p < 0.001$, $\eta p^2 = 0.469$), indicating differential modulation of
503 heteronymous facilitation across age groups. In older participants, $H_{\text{fac}}/H_{\text{test}}$ remained relatively
504 stable across contraction types and muscle lengths, with no significant differences observed
505 between eccentric and isometric contractions or between intermediate and long muscle lengths.
506 However, age-related differences emerged when comparing younger and older participants.
507 During isometric contractions at intermediate muscle lengths, younger participants exhibited a
508 27.0 % higher $H_{\text{fac}}/H_{\text{test}}$ than older adults ($p < 0.001$), while this difference remained significant
509 but slightly lower at long muscle lengths (+18.7% in younger adults). Similarly, during
510 eccentric contractions at intermediate lengths, $H_{\text{fac}}/H_{\text{test}}$ was 24.0 % higher in younger compared
511 to older participants ($p < 0.001$). However, at long muscle lengths during eccentric contractions,
512 no significant age-related differences were observed ($p > 0.05$).

513 In younger participants, both muscle length and contraction type significantly influenced
514 $H_{\text{fac}}/H_{\text{test}}$. During isometric contractions at long muscle lengths, $H_{\text{fac}}/H_{\text{test}}$ decreased by 11.2%
515 compared to isometric contractions at intermediate length. A similar but more pronounced
516 reduction (19.5%) was observed during eccentric contractions at long muscle lengths compared
517 to intermediate lengths. Furthermore, eccentric contractions led to a 5.8% lower $H_{\text{fac}}/H_{\text{test}}$
518 than isometric contractions at intermediate muscle lengths, with this difference increasing to
519 14.6% at long muscle lengths.
<figure><img src="image_9.png" /><figcaption>**Figure 7.** Effects of age, muscle length, and contraction type on heteronymous Ia facilitation. (A) Representative traces show the non-conditioned soleus H reflex (H<sub>test</sub>) and the conditioned H reflex (H<sub>Fac</sub>) in a representative older participant. H<sub>test</sub> and H<sub>Fac</sub> were evoked by posterior tibial nerve stimulation, with H<sub>Fac</sub> conditioned by femoral nerve stimulation applied after posterior tibial nerve stimulation at conditioning intervals between 9 and 1 ms. Traces are shown for both isometric and eccentric contractions at intermediate and long muscle lengths. (B) Absolute and individual data are presented as mean ± standard deviation. H<sub>Fac</sub> and H<sub>test</sub> ratios are shown for isometric and eccentric contractions at intermediate and long muscle lengths. The electrical intensity to evoke H<sub>test</sub> was normalized across all contractions at each muscle length. Young adults are represented in blue with filled circles, while older adults are represented in orange with open circles. Abbreviations: H<sub>Fac</sub>, conditioned soleus H reflex; H<sub>test</sub>, non-conditioned soleus H reflex; Ecc, eccentric; Con, concentric; Iso,</figcaption></figure>
548
549
550

isometric. * Indicates significant difference from eccentric contraction type. # Indicates significant difference from intermediate muscle length. α Indicates significant difference by age within the same experimental condition.

551

## Effectiveness of activated Ia afferents to discharge α-motoneurons and reflex gain

552
553
554
555
556

In this study, SOL Hmax/Mmax and Hslope/slope were evaluated to estimate the effectiveness of activated Ia afferents in stimulating α-motoneurons and the reflex gain, respectively. Figure 8A presents raw traces of the SOL H-reflexes and M-waves elicited during eccentric and isometric contractions at intermediate and long muscle lengths in representative young and older participants.

557
558
559
560
561
562
563
564
565

A significant three-way interaction was observed between contraction type, muscle length, and age for Hmax/Mmax (F(1, 33) = 31.716, P < 0.001, ηp² = 0.490). Regardless of age, muscle length and contraction type influenced Hmax/Mmax, but the effects differed between young and older participants. During isometric contractions, a significant decrease in Hmax/Mmax at long muscle length compared to intermediate length was observed in young adults (-24%, P < 0.001), while this effect was not significant in older adults (P = 0.064). In contrast, during eccentric contractions, the opposite pattern was found: older adults exhibited a significant 17.8% decrease in Hmax/Mmax at long compared to intermediate muscle length (P < 0.001), whereas no difference was observed in young adults (P > 0.05).

566
567
568
569
570
571

Contraction type also had a differential impact depending on age. At an intermediate muscle length, young adults showed an 18.6% lower Hmax/Mmax during eccentric compared to isometric contractions (P < 0.001), while no difference was observed in older adults (P = 0.271). Conversely, at a long muscle length, the effect was reversed: older adults exhibited a 17.2% lower Hmax/Mmax during eccentric compared to isometric contractions (P < 0.001), while no significant difference was found in young adults (P > 0.05).

572
<figure><img src="image_10.png" /><figcaption>Figure 8. Changes in maximal soleus H-reflex amplitude normalized to the maximal M-wave amplitude across muscle length, contraction type, and age. (A) Representative traces of the maximal soleus H-reflex evoked by posterior tibial nerve stimulation at intermediate and long muscle lengths during isometric and eccentric contractions in a representative young (red line) and older (black line) participant. (B) Absolute and individual data for elderly (n = 19) and young participants (n = 16) are expressed as mean ± standard deviation, showing changes in the maximal H-reflex normalized to the maximal M-wave at intermediate and long muscle lengths during isometric and eccentric contractions. Young adults are represented in blue with filled lines, while older adults are represented in orange with open lines. Abbreviations: Hmax, maximal soleus H-reflex; Mmax, maximal M-wave; Ecc, eccentric; Iso, isometric. * Significantly different from other conditions (p < 0.05).</figcaption></figure>
Figure 9 shows the soleus H-reflex slope to M-wave slope ratio according to muscle length and age. For the $H_{slope}/M_{slope}$, a significant main effect of contraction type was observed ($F(1,29) = 6.853$, $p < 0.05$, $ηp^2 = 0.191$), indicating that this ratio differs between isometric and eccentric contractions, regardless of age and muscle length. Post-hoc analysis revealed an 8.1% reduction in $H_{slope}/M_{slope}$ during eccentric compared to isometric contractions ($p = 0.014$).

Additionally, a significant interaction between age and muscle length was found ($F(1,29) = 26.161$, $p < 0.001$, $ηp^2 = 0.474$), suggesting differential modulation of $H_{slope}/M_{slope}$ by muscle length in young versus older participants. Specifically, age influenced $H_{slope}/M_{slope}$, with a 61.5% reduction at intermediate muscle lengths in older adults compared to younger adults ($p < 0.001$), and a 47.8% reduction at long muscle lengths ($p < 0.001$). Muscle length also affected $H_{slope}/M_{slope}$. In young participants, the ratio decreased by 14.6% at long muscle lengths compared to intermediate lengths ($p < 0.001$); however, this modulation of the $H_{slope}/M_{slope}$ across muscle lengths was not observed in older adults ($p > 0.05$).

<figure><img src="image_11.png" /><figcaption>**Figure 9.** Soleus H-reflex slope to M-wave slope ratio according to muscle length and age. Data are presented as mean ± standard deviation, with individual values shown. Young adults are represented in blue with filled circles, while older adults are represented in orange with open circles. $H_{slope}$, slope of the soleus H-reflex; $M_{slope}$, slope of the M-wave; Inter, intermediate muscle length; Long, long muscle length. * Significantly different from other conditions ($p < 0.05$).</figcaption></figure>
We measured SOL $M_{max}$ to ensure stability of EMG recording conditions across experimental conditions. For $M_{max}$, no interaction effect between conditions was observed (all p values > 0.117, Table 2).

To verify that similar numbers of efferent and potentially afferent nerve fibers were stimulated in each condition, we measured SOL $M_{atHmax}$. Results show that both $M_{max}$ and $M_{atHmax}$ amplitudes remained stable across all conditions, indicating that equivalent proportions of α-motoneurons were activated (Table 2). This consistency suggests that $H_{max}/M_{max}$ modulations are attributable to neural mechanisms rather than recording conditions (35).

<table><thead><tr><th rowspan="2"></th><th colspan="2">Young</th><th colspan="2">Old</th></tr><tr><th>Intermediate</th><th>Long</th><th>Intermediate</th><th>Long</th></tr></thead><tbody><tr><td><strong>Eccentric</strong></td><td></td><td></td><td></td><td></td></tr><tr><td>$M_{max}$ (mV)</td><td>5.968 ± 1.158</td><td>5.747 ± 1.060</td><td>4.399 ± 1.777</td><td>4.306 ± 1.732</td></tr><tr><td>$H_{max}/M_{max}$ (u.a)</td><td>0.549 ± 0.122</td><td>0.417 ± 0.101</td><td>0.459 ± 0.136</td><td>0.413 ± 0.148</td></tr><tr><td>$M_{atHmax}/M_{max}$ (a.u)</td><td>0.155 ± 0.069</td><td>0.141 ± 0.019</td><td>0.171 ± 0.113</td><td>0.206 ± 0.117</td></tr><tr><td>$H_{test}/M_{max}$ (a.u)</td><td>0.398 ± 0.111</td><td>0.392 ± 0.102</td><td>0.347 ± 0.093</td><td>0.327 ± 0.099</td></tr><tr><td><strong>Isometric</strong></td><td></td><td></td><td></td><td></td></tr><tr><td>$M_{max}$ (mV)</td><td>5.883 ± 1.019</td><td>5.992 ± 1.058</td><td>4.459 ± 1.822</td><td>4.286 ± 1.698</td></tr><tr><td>$H_{max}/M_{max}$ (u.a)</td><td>0.447 ± 0.117</td><td>0.427 ± 0.101</td><td>0.416 ± 0.136</td><td>0.342 ± 0.134</td></tr><tr><td>$M_{atHmax}/M_{max}$ (a.u)</td><td>0.158 ± 0.050</td><td>0.164 ± 0.102</td><td>0.170 ± 0.075</td><td>0.178 ± 0.082</td></tr><tr><td>$H_{test}/M_{max}$ (a.u)</td><td>0.399 ± 0.119</td><td>0.396 ± 0.109</td><td>0.339 ± 0.092</td><td>0.340 ± 0.095</td></tr></tbody></table>

**Table 2. Effect of contraction type, muscle length and age on soleus evoked potential.** Absolute data are presented as mean ± standard deviation. $M_{max}$, maximal M wave; $H_{max}$, maximal soleus H reflex; $M_{atHmax}$, M wave corresponding to maximal soleus H reflex; $H_{test}$, non-conditioned soleus H reflex corresponding to its test value for D1 and heteronymous Ia facilitation methods.

## DISCUSSION
636

637

638

639

640

641

642

643

644

This study investigated the impact of aging on presynaptic modulation by PAD with respect to contraction type and muscle length. Our findings revealed that in older adults, PAD remains unchanged across muscle lengths or contraction types. In contrast, younger adults exhibited greater PAD at longer muscle lengths compared to shorter and during eccentric contractions compared to isometric contractions. These results highlight an age-related impairment in the presynaptic control of Ia-motoneuron transmission in response to muscle constraints that alter Ia afferent discharge.

645

646

647

648

649

650

651

652

653

654

655

656

Age-related Modulation of presynaptic inhibition by PAD

Muscle lengthening and muscle length significantly influence PAD regulation in healthy young adults (5,18,19). Specifically, PAD is greater during eccentric contractions compared to isometric ones and at longer muscle lengths compared to shorter ones. This modulation likely results from increased Ia afferent discharge associated with muscle lengthening and long muscle length (18). In contrast to younger participants, older individuals did not exhibit presynaptic modulation by PAD based on contraction type or muscle length. Furthermore, to evaluate presynaptic mechanisms, complementary approaches are necessary, such as assessing ongoing Ia presynaptic inhibition by examining monosynaptic facilitation of the soleus H-reflex via femoral nerve stimulation (40). Interestingly, older adults exhibited limited facilitation (cf. methodological considerations), with no variation across experimental conditions, in contrast to younger participants. This supports the findings of an absence of presynaptic mechanism modulation in older adults based on contraction type and muscle length.

657

658

659

660

The age-related decline in presynaptic modulation by PAD in response to muscle length and contraction type may result from a combination of peripheral and central changes, leading to differential motor control. Peripheral changes may be attributed to morphological changes in muscle spindles (21,22) and decreased sensitivity with aging (23,24), which diminish
responsiveness to stretch and changes in muscle length, thereby affecting the Ia afferent discharge rate. These peripheral changes are accompanied by central alterations at both spinal and supraspinal levels. At the spinal level, Terao et al. (1996) demonstrated a significant reduction in small neurons, i.e., mainly interneurons, distributed within the intermediate zone of the ventral horn of the spinal cord. Furthermore, their findings further indicate that among the spinal ventral horn cells, these small neurons in the intermediate zone, rather than large α-motoneurons and medium-sized γ-motoneurons, are more profoundly affected by aging (47). Consequently, these age-related morphological changes in the spinal cord could impair the integration of afferent inputs, thereby also altering the regulation of PAD.

At the supra-spinal level, aging induces structural and functional changes in brain regions involved in motor control (13,48). Aging affects sensorimotor structures, potentially leading to functional impairments and reduced motor performance. This decline may necessitate compensatory mechanisms to preserve movement execution (49). Indeed, age-related reductions in proprioceptive input from muscle spindles may impair the efficiency of automatic control processes, requiring a greater reliance on controlled processes and consequently increasing cognitive load (12). Neuroimaging and electrophysiological studies show age-related reductions in cortical inhibition and increased activation across multiple cortical areas during motor tasks. These findings suggest a shift toward greater reliance on voluntary, supraspinal control in older adults, potentially due to reduced efficiency of automatic spinal mechanisms (13). Such changes in the supraspinal control of movement may affect the processing and integration of afferent feedback, thereby influencing the regulation of presynaptic inhibition of Ia afferents by PAD, and ultimately modifying its contribution to motor output.

Taken together, these findings suggest that aging significantly impacts PAD regulation and alters motor control strategies. Our results are in line with previous studies examining PAD
686 modulation in different contexts. For instance, Baudry et al. (2010) found that older adults
687 exhibited no variation in PAD between position and force control tasks, relying instead on
688 increased antagonist coactivation for limb stabilization. In contrast, younger adults
689 demonstrated task-specific PAD modulation, with PAD decreasing during position control
690 compared to force control (8). Similarly, Butchart et al. (1993) showed that older participants
691 lacked consistent PAD modulation across torque levels, whereas younger adults exhibited
692 torque-dependent PAD modulation (9). Earles et al. (2001) also observed reduced PAD
693 modulation in older individuals during submaximal efforts, with greater reliance on direct
694 motor activation to maintain motor output (25). Notably, PAD modulation in older adults may
695 depend on task specificity. For example, Baudry and Duchateau (2012) found that older adults
696 showed heightened PAD and greater co-contraction under challenging balance conditions (e.g.,
697 with eyes closed or on compliant surfaces) compared to younger adults (43). Differences
698 between these findings and our results may stem from variations in the motor tasks performed.

699 Although presynaptic mechanisms play a crucial role in regulating synaptic transmission
700 between Ia afferent fibers and motoneurons, the findings on PAD do not consistently align with
701 those on the efficacy of Ia-motoneuron transmission, especially in older adults.

702 Age-related Efficiency of Ia-Motoneuron Transmission

703 In young adults, the capacity of Ia afferents to activate alpha motoneurons is reduced during
704 eccentric contractions at intermediate muscle lengths (6,7), consistent with our findings
705 showing lower Hmax/Mmax and Hslope/slope ratios during eccentric compared to isometric
706 contractions at intermediate lengths. This inhibition likely results from recurrent inhibition via
707 Renshaw cells (18,19,50,51) and post-activation depression by PAD (5,18,19). However,
708 despite greater PAD during eccentric contractions at long muscle lengths, Hmax/Mmax did not
709 differ from isometric contractions at the same length, likely due to differences in recurrent
710 inhibition.
In older adults, $H_{max}/M_{max}$ was modulated by muscle length and contraction type, but differently compared to young individuals. Specifically, $H_{max}/M_{max}$ was similar between isometric and eccentric contractions at intermediate muscle lengths. This highlights a distinct behavior of the $H_{max}/M_{max}$ in older adults compared to young adults, where it is typically reduced during eccentric contractions at intermediate muscle lengths (6,7,18). Furthermore, $H_{max}/M_{max}$ was reduced during eccentric contractions at long muscle lengths in older adults, whereas this was not the case in young adults. These results are surprising, as presynaptic inhibition by PAD does not explain this decrease. It is possible that the loss of neuromuscular spindle sensitivity (23) does not generate sufficient afferent discharge at an intermediate muscle length to regulate $H_{max}/M_{max}$, whereas this regulation may still occur at a longer muscle length. It is therefore plausible that aging also alters other spinal and/or supraspinal mechanisms contributing to the modulation of Ia-alpha motoneuron transmission according to muscle length and lengthening.

At the spinal level there are several contributing age-related changes such as altered recurrent inhibition via Renshaw cells, potential reductions in Group II afferent facilitation (52), and modified Group Ib afferent input due to muscle-tendon adaptations (53). For example, regarding recurrent inhibition, our research team previously demonstrated that recurrent inhibition was more pronounced during eccentric contractions than during isometric contractions at intermediate muscle length. However, at long muscle length, recurrent inhibition was similar between these two contraction types. Similarly, $H_{max}/M_{max}$ was comparable across different contraction types at long muscle length (18). This was not the case in the current study for older participants, suggesting that recurrent inhibition could be a key candidate explaining the observed reduction in Ia-motoneuron alpha transmission in older subjects.

Similar findings were reported by Chen et al. (2014), who observed that older adults exhibited a significantly lower $H_{max}/M_{max}$ during isometric contractions in the dorsiflexion position (-20°) compared to the neutral (0°) and plantar flexion (20°) positions. However, these results should
be interpreted with caution due to differences in joint angle (-15° in our study) and the absence
of $M_{max}$ measurements for each active position in their study, which complicates the
interpretation of $H_{max}$ (54). In our study, the amplitude of $H_{max}/M_{max}$ was not significantly
different between young and older adults, except during eccentric at long muscle length. This
contrasts with studies conducted under static and isometric conditions (55,56) and in dynamic
conditions (57), which indicated lower spinal excitability in older adults. For example, Skarabot
et al. (2019) demonstrated that the amplitudes of the H reflex, normalized to RMS, were lower in
older adults regardless of contraction type (i.e., eccentric and concentric). However, the
differences observed may be related to the muscle group investigated in their study (i.e., tibialis
anterior). Finally, our results showed that the $H_{slope}/M_{slope}$ ratio was significantly lower in older
adults, regardless of contraction type or muscle length. This indicates a reduced reflex gain (45
during muscle contractions in this population, suggesting that the recruitment rate of
motoneurons in response to increased Ia afferent input is reduced with aging. This reduction
may be attributed to morphological changes in muscle spindles (21,22) decreased spindle
sensitivity (23,58), and alterations in the regulation of presynaptic mechanisms (9,25,59).

Beyond the neurophysiological mechanisms, these findings may have functional relevance in
older adults. The absence of PAD modulation with respect to contraction type and muscle
length may compromise the capacity to fine-tune reflex pathways during daily motor activities
that demand rapid adjustments, such as maintaining balance during gait or responding to
unexpected perturbations. Such reduced adaptability could contribute to deficits in postural
control, diminished locomotor efficiency, and ultimately a greater risk of falls and fall-related
hospitalizations, which represent major causes of morbidity and loss of independence in older
adults (60). While our study was not designed to directly assess clinical outcomes, it highlights
a potential mechanistic link between altered spinal reflex regulation and age-related functional
760

decline, thereby emphasizing the need to explore whether targeted interventions (e.g., resistance
or balance training) can mitigate these impairments.

761

762

Methodological considerations

763

While this study provides valuable insights into motor control and aging, certain limitations
should be acknowledged. One limitation relates to the measurement of heteronymous Ia
facilitation. Accurately evaluating presynaptic mechanisms requires combining two
complementary methods. The first assesses PAD activity via changes in the H-reflex amplitude
conditioned by fibular nerve stimulation, reflecting its responsiveness to conditioning inputs
(Mizuno et al., 1971). The second evaluates the monosynaptic facilitation of the soleus H-reflex
by femoral nerve stimulation, which indicates the level in ongoing Ia presynaptic inhibition
(Hultborn et al., 1987). This dual approach helps mitigate potential biases related to reflex
recruitment gain (42). However, in older adults, we observed minimal facilitation (<10% of
Htest), which is consistent with previous findings of age-related declines in heteronymous Ia
facilitation (59). This limited facilitation complicates interpretation of PAD amplitude and
raises the possibility of occlusion effects, even though findings were consistent across
conditions and unaffected by muscle length or contraction type. We contend that these
limitations do not affect our main results. Thus, this study offers a solid foundation for
understanding age-related changes in motor control and opens several avenues for future
research.

764

765

766

767

768

769

770

771

772

773

774

775

776

777

778

779

Future directions

780

In this study, presynaptic modulation via PAD differs between older adults and young
participants in this study. However future investigations should explore other spinal
mechanisms that may also contribute to altered Ia-alpha motoneurons transmission. At the
spinal level, age-related changes such as altered recurrent inhibition via Renshaw cells,

781

782

783
potential reductions in Group II afferent facilitation (52), and modified Group Ib afferent input due to muscle-tendon adaptations (53) may all play a role. Additionally, it would be valuable to link the modification of these mechanisms' behaviour with aging to a potential decrease in the sensitivity of muscle spindles. Such age-related reductions in proprioceptive input from muscle spindles may impair the efficiency of automatic control processes, necessitating a greater reliance on controlled processes and consequently increasing cognitive load (12). Therefore, it would be interesting to investigate the contribution of supraspinal changes to the regulation and modulation of PAD. It would be also valuable to examine inter-individual effects to determine which conditions (e.g., sex, level of physical activity, or age) impact PAD regulation. Finally, although PAD appears to be altered with aging, it remains unclear whether older adults can modulate PAD in response to physical interventions such as balance training, resistance training, or aerobic exercise, even though these modalities have been shown to induce other neuromuscular adaptations (44,61). To date, only one study has investigated whether physical intervention is able to modulate PAD (62). This study showed that 8-9 weeks of eccentric resistance training reduce PAD amplitude in young adults.

## Conclusion

This study demonstrates that aging impairs the regulation of presynaptic mechanisms involved in Ia-motoneuron transmission in response to muscle length and contraction type. Unlike young adults, older adults do not adjust PAD according to mechanical constraints, indicating altered spinal reflex control. This dysregulation likely results from age-related peripheral and central changes affecting sensorimotor integration. These findings highlight the need to investigate potential compensatory spinal or supraspinal mechanisms and to determine whether physical interventions can help preserve the adaptability of presynaptic regulation in response to motor task demands and constraints.
809

810 ACKNOWLEDGMENTS

811 We sincerely thank all the participants for their time and commitment. We also acknowledge

812 the technical support provided by the laboratory staff.

813 SOURCES OF FUNDING

814 This study was supported by the French National Research Agency (ANR) under grant number

815 ANR-22-CE17-0002.

816 CONFLICT OF INTEREST

817 None declared.

818 AUTHOR CONTRIBUTIONS

819 Yohan Betus, Julian Colard, Thomas Cattagni, and Marc Jubeau : conception or design of the work.

820 Yohan Betus, Julian Colard, Thomas Cattagni, and Marc Jubeau : acquisition, analysis, or interpretation of data for the work.

821 Anne-Sophie Boureau, Guillaume Chapelet, and Christophe Cornu: participant recruitment, notably through the coordination and conduct of medical consultations.

822 All authors drafted and revised the manuscript, approved the final version, and agree to be accountable for all aspects of the work, ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

823 All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed.

830 REFERENCES

831 1. Baudry S, Penzer F, Duchateau J. Input-output characteristics of soleus homonymous Ia afferents and corticospinal pathways during upright standing differ between young and elderly adults. Acta Physiol. 2014;210(3):667-77.

834 2. Katz R, Meunier S, Pierrot-Deseilligny E. Changes in presynaptic inhibition of Ia fibres in man while standing. Brain. 1988 Apr;111(2):417-437.

836 3. Koceja DM, Trimble MH, Earles DR. Inhibition of the soleus H-reflex in standing man. Brain Res. 26 nov 1993;629(1):155-8.

838 4. Theodosiadou A, Henry M, Duchateau J, Baudry S. Revisiting the use of Hoffmann reflex in motor control research on humans. Eur J Appl Physiol. 1 avr 2023;123(4):695-710.

840 5. Colard J, Jubeau M, Duclay J, Cattagni T. Regulation of primary afferent depolarization and homosynaptic post-activation depression during passive and active lengthening, shortening and isometric conditions. Eur J Appl Physiol. 1 juin 2023;123(6):1257-69.
843

844

845

6. Duclay J, Pasquet B, Martin A, Duchateau J. Specific modulation of spinal and cortical excitabilities during lengthening and shortening submaximal and maximal contractions in plantar flexor muscles. J Appl Physiol. 15 déc 2014;117(12):1440-50.

846

847

7. Duclay J, Martin A. Evoked H-Reflex and V-Wave Responses During Maximal Isometric, Concentric, and Eccentric Muscle Contraction. J Neurophysiol. nov 2005;94(5):3555-62.

848

849

8. Baudry S, Maerz AH, Enoka RM. Presynaptic Modulation of Ia Afferents in Young and Old Adults When Performing Force and Position Control. J Neurophysiol. févr 2010;103(2):623-31.

850

851

852

9. Butchart P, Farquhar R, Part N, Roberts R. The effect of age and voluntary contraction on presynaptic inhibition of soleus muscle Ia afferent terminals in man. Exp Physiol. 1993;78(2):235-42.

853

854

10. Earles DR, Koceja DM, Shively CW. Environmental Changes in Soleus H-Reflex Excitability in Young and Elderly Subjects. Int J Neurosci. 1 janv 2000;105(1-4):1-13.

855

856

11. Koceja DM, Mynark RG. Comparison of Heteronymous Monosynaptic Ia Facilitation in Young and Elderly Subjects in Supine and Standing Positions. Int J Neurosci. 1 janv 2000;104(1):1-15.

857

858

12. Henry M, Baudry S. Age-related changes in leg proprioception: implications for postural control. J Neurophysiol. août 2019;122(2):525-38.

859

860

13. Hunter SK, Pereira HM, Keenan KG. The aging neuromuscular system and motor performance. J Appl Physiol. oct 2016;121(4):982-95.

861

862

14. Matthews PBC, Stein RB. The sensitivity of muscle spindle afferents to small sinusoidal changes of length. J Physiol. 1969;200(3):723-43.

863

864

15. Matthews PBC. Muscle spindles: their messages and their fusimotor supply. In: Comprehensive Physiology. John Wiley & Sons, Ltd; 2011. p. 189-228.

865

866

16. Pierrot-Deseilligny E, Burke D. The Circuitry of the Human Spinal Cord: Spinal and Corticospinal Mechanisms of Movement. Cambridge University Press; 2012. 631 p.

867

868

17. Burke D, Hagbarth KE, Löfstedt L. Muscle spindle activity in man during shortening and lengthening contractions. J Physiol. 1978;277(1):131-42.

869

870

871

18. Colard J, Duclay J, Bétus Y, Cattagni T, Jubeau M. Muscle length modulates recurrent inhibition and presynaptic inhibition of Ia afferents differently depending on type of contraction. Eur J Neurosci. 2025 Jul;62(1):e70172.

872

873

874

19. Papitsa A, Paizis C, Papaiordanidou M, Martin A. Specific modulation of presynaptic and recurrent inhibition of the soleus muscle during lengthening and shortening submaximal and maximal contractions. J Appl Physiol. déc 2022;133(6):1327-40.

875

20. Stein RB. Presynaptic inhibition in humans. Prog Neurobiol. 1 déc 1995;47(6):533-44.

876

877

21. Swash M, Fox KP. The effect of age on human skeletal muscle studies of the morphology and innervation of muscle spindles. J Neurol Sci. 1 août 1972;16(4):417-32.

878

879

22. Kararizou E, Manta P, Kalfakis N, Vassilopoulos D. Morphometric study of the human muscle spindle. Anal Quant Cytol Histol. 1 févr 2005;27(1):1-4.

880

881

23. Kim GH, Suzuki S, Kanda K. Age-related physiological and morphological changes of muscle spindles in rats. J Physiol. 2007;582(2):525-38.
882 24. Miwa T, Miwa Y, Kanda K. Dynamic and static sensitivities of muscle spindle primary endings in aged rats to ramp stretch. Neurosci Lett. 8 déc 1995;201(2):179-82.

884 25. Earles D, Vardaxis V, Koceja D. Regulation of motor output between young and elderly subjects. Clin Neurophysiol. 1 juill 2001;112(7):1273-9.

886 26. McNeil CJ, Rice CL. Neuromuscular adaptations to healthy aging. Appl Physiol Nutr Metab. nov 2018;43(11):1158-65.

888 27. Proske U, Morgan DL, Gregory JE. Thixotropy in skeletal muscle and in muscle spindles: A review. Prog Neurobiol. 1 déc 1993;41(6):705-21.

890 28. Cattagni T, Merlet AN, Cornu C, Jubeau M. H-reflex and M-wave recordings: effect of pressure application to the stimulation electrode on the assessment of evoked potentials and subject's discomfort. Clin Physiol Funct Imaging. 2018;38(3):416-24.

893 29. Bolsterlee B, Finni T, D'Souza A, Eguchi J, Clarke EC, Herbert RD. Three-dimensional architecture of the whole human soleus muscle in vivo. PeerJ. 18 avr 2018;6:e4610.

895 30. Nakamura N, Croisier C, Maffiuletti P. Acute effects of static stretching on passive stiffness of the hamstring muscles calculated using different mathematical models. Clin Biomech (Bristol, Avon). 2006 Aug

898 31. Farris DJ, Lichtwark GA. UltraTrack: Software for semi-automated tracking of muscle fascicles in sequences of B-mode ultrasound images. Comput Methods Programs Biomed. mai 2016;128:111-8.

900 32. Carville SF, Perry MC, Rutherford OM, Smith ICH, Newham DJ. Steadiness of quadriceps contractions in young and older adults with and without a history of falling. Eur J Appl Physiol. 1 juill 2007;100(5):527-33.

903 33. Enoka RM, Christou EA, Hunter SK, Kornatz KW, Semmler JG, Taylor AM, et al. Mechanisms that contribute to differences in motor performance between young and old adults. J Electromyogr Kinesiol. 1 févr 2003;13(1):1-12.

906 34. Tracy BL, Enoka RM. Older adults are less steady during submaximal isometric contractions with the knee extensor muscles. J Appl Physiol. mars 2002;92(3):1004-12.

908 35. Schieppati M. The Hoffmann reflex: A means of assessing spinal reflex excitability and its descending control in man. Prog Neurobiol. 1 janv 1987;28(4):345-76.

910 36. Mizuno Y, Tanaka R, Yanagisawa N. Reciprocal group I inhibition on triceps surae motoneurons in man. J Neurophysiol. nov 1971;34(6):1010-7.

912 37. Metz K, Matos IC, Hari K, Bseis O, Afsharipour B, Lin S, et al. Post-activation depression from primary afferent depolarization (PAD) produces extensor H-reflex suppression following flexor afferent conditioning. J Physiol. 2023;601(10):1925-56.

915 38. Aymard C, Katz R, Lafitte C, Lo E, Pénicaud A, Pradat-Diehl P, et al. Presynaptic inhibition and homosynaptic depression: a comparison between lower and upper limbs in normal human subjects and patients with hemiplegia. Brain J Neurol. août 2000;123 (Pt 8):1688-702.

918 39. Magalhães FH, Elias LA, Silva CR da, Lima FF de, Toledo DR de, Kohn AF. D1 and D2 Inhibitions of the Soleus H-Reflex Are Differentially Modulated during Plantarflexion Force and Position Tasks. PLOS ONE. 24 nov 2015;10(11):e0143862.
40. Hultborn H, Meunier S, Morin C, Pierrot-Deseilligny E. Assessing changes in presynaptic inhibition of Ia fibres: a study in man and the cat. J Physiol. août 1987;389:729-56.

41. Baudry S, Enoka RM. Influence of load type on presynaptic modulation of Ia afferent input onto two synergist muscles. Exp Brain Res. 1 oct 2009;199(1):83-8.

42. Johannsson J, Duchateau J, Baudry S. Presynaptic inhibition of soleus Ia afferents does not vary with center of pressure displacements during upright standing. Neuroscience. 9 juill 2015;298:63-73.

43. Baudry S, Duchateau J. Age-related influence of vision and proprioception on Ia presynaptic inhibition in soleus muscle during upright stance. J Physiol. 2012;590(21):5541-54.

44. Penzer F, Duchateau J, Baudry S. Effects of short-term training combining strength and balance exercises on maximal strength and upright standing steadiness in elderly adults. Exp Gerontol. 1 janv 2015;61:38-46.

45. Funase K, Imanaka K, Nishihira Y. Excitability of the soleus motoneuron pool revealed by the developmental slope of the H-reflex as reflex gain. Electromyogr Clin Neurophysiol. 1 déc 1994;34(8):477-89.

46. Klimstra M, Zehr EP. A sigmoid function is the best fit for the ascending limb of the Hoffmann reflex recruitment curve. Exp Brain Res. 1 mars 2008;186(1):93-105.

47. Terao S ichi, Sobue G, Hashizume Y, Li M, Inagaki T, Mitsuma T. Age-related changes in human spinal ventral horn cells with special reference to the loss of small neurons in the intermediate zone: a quantitative analysis. Acta Neuropathol (Berl). 15 juill 1996;92(2):109-14.

48. Borzuola R, Giombini A, Torre G, Campi S, Albo E, Bravi M, et al. Central and Peripheral Neuromuscular Adaptations to Ageing. J Clin Med. mars 2020;9(3):741.

49. Poirier G, Ohayon A, Juranville A, Mourey F, Gaveau J. Deterioration, Compensation and Motor Control Processes in Healthy Aging, Mild Cognitive Impairment and Alzheimer's Disease. Geriatr Basel Switz. 23 mars 2021;6(1):33.

50. Barrué-Belou S, Marque P, Duclay J. Recurrent inhibition is higher in eccentric compared to isometric and concentric maximal voluntary contractions. Acta Physiol. 2018;223(4):e13064.

51. Barrué-Belou S, Marque P, Duclay J. Supraspinal Control of Recurrent Inhibition during Anisometric Contractions. Med Sci Sports Exerc. 21 nov 2019;51(11):2357.

52. Simonetta-Moreau M, Marque P, Marchand-Pauvert V, Pierrot-Deseilligny E. The pattern of excitation of human lower limb motoneurones by probable group II muscle afferents. J Physiol. 1999;517(1):287-300.

53. Lindemann I, Coombes BK, Tucker K, Hug F, Dick TJM. Age-related differences in gastrocnemii muscles and Achilles tendon mechanical properties in vivo. J Biomech. 9 nov 2020;112:110067.

54. Chen YS, Zhou S, Cartwright C. Effects of Ankle Joint Position and Submaximal Muscle Contraction Intensity on Soleus H-reflex Modulation in Young and Older Adults. Motor Control. 1 avr 2014;18(2):112-26.

55. Kido A, Tanaka N, Stein RB. Spinal excitation and inhibition decrease as humans age. Can J Physiol Pharmacol. avr 2004;82(4):238-48.
960
961
962

56. Scaglioni G, Narici MV, Maffiuletti NA, Pensini M, Martin A. Effect of ageing on the electrical and mechanical properties of human soleus motor units activated by the H reflex and M wave. J Physiol. 2003;548(2):649-61.

963
964
965

57. Škarabot J, Ansdell P, Brownstein CG, Hicks KM, Howatson G, Goodall S, et al. Reduced corticospinal responses in older compared with younger adults during submaximal isometric, shortening, and lengthening contractions. J Appl Physiol. 1 avr 2019;126(4):1015-31.

969
970
971

58. Burke JR, Schutten MC, Koceja DM, Kamen G. Age-dependent effects of muscle vibration and the Jendrassik maneuver on the patellar tendon reflex response. Arch Phys Med Rehabil. 1 juin 1996;77(6):600-4.

972
973

59. Morita H, Shindo M, Yanagawa S, Yoshida T, Momoi H, Yanagisawa N. Progressive decrease in heteronymous monosynaptic la facilitation with human ageing. Exp Brain Res. 1 avr 1995;104(1):167-70.

974
975
976

60. Rubenstein LZ. Falls in older people: epidemiology, risk factors and strategies for prevention. Age Ageing. 1 sept 2006;35(suppl_2):ii37-41.

61. Chambers TL, Burnett TR, Raue U, Lee GA, Finch WH, Graham BM, et al. Skeletal muscle size, function, and adiposity with lifelong aerobic exercise. J Appl Physiol Bethesda Md 1985. 1 févr 2020;128(2):368-78.

977
978
979

62. Colard J, Bétus Y, Tallio T, Bizet B, Nordez A, Jubeau M, Cattagni T. Eccentric training at long muscle lengths induces greater corticospinal and spinal reflex plasticity than eccentric training at short muscle lengths. Exp Physiol. 2025 Jun 2

980

981

982

983

984