MATERIALS AND METHODS

Subjects

We used data from subjects who were recruited for a recently published study.¹² Written consent was obtained before their participation. All were fully informed of possible risks or discomfort associated with the experimental protocol. Fifteen sedentary, healthy, young men (mean (SD) age 22 (5) years) were selected. Their good health had been confirmed by physical examination. The study conformed to the code of ethics of the World Medical Association (Declaration of Helsinki), and the ethics committee of the Universidad Complutense de Madrid approved it.

Protocol

The participants were randomly divided into two groups: control and electrostimulated (ES) subjects. The five subjects forming the control group (mean (SD) age 22 (3) years; height 176.2 (8.6) cm; body mass 74.0 (9.4) kg) did not undergo muscle ES but were subjected to the same tests as those in the ES group. The latter (n = 10; age 22 (5) years; height 175.0 (5.5) cm; body mass 71.2 (6.7) kg) underwent ES of both quadriceps muscles over a six week period as described below. All participants continued with their sedentary lifestyle during the six week period of the study. Before and after this period, all subjects performed an exercise test on a cycle ergometer.

ES programme

The quadriceps muscles of both legs of subjects assigned to the ES group were subjected to 30 minutes per session and day of transcutaneous ES, three days a week for six weeks. ES was elicited using bipolar electrodes²⁴ and a portable battery powered neuromuscular electrical stimulator (Stiwell Medical Technologies, Villeneuve, Switzerland). The cathode was positioned proximally—that is, 5 cm below the major trochanter of the femur—and the anode was placed over the motor point of the vastus lateralis muscle—that is, the region 5 cm laterally above the upper edge of the patella.⁴

The stimulation frequency during each session was 45–60 Hz with an impulse of 300 μs and a duty cycle of 12 seconds on/eight seconds off. This frequency was selected because it has been applied in numerous human studies and because low frequencies are used by most physiotherapists. The pulse amplitude was selected by each subject according to their maximum tolerance level. The average stimulation intensity during the six week period increased from about 25 mA (1st week) to about 75 mA (6th week). During each session, subjects lay on a bed in a supine position with legs semiflexed (knee joint at 150–160°).

Cycle ergometer tests

The exercise test consisted of a ramp protocol until exhaustion performed on an electronically braked cycle ergometer (Ergometrics 900; Ergo-line, Barcelona, Spain). Environmental conditions were similar for all the tests (21–24°C, 45–55% relative humidity), and the subjects were cooled with a fan throughout the bouts of exercise. After a two minute rest sitting on the cycle ergometer, the test started at 20 W and the workload was increased by 20 W/min (5 W/15 s). Pedalling cadence was kept between 60 and 80 rev/min depending on the subject’s preferences. A pedal frequency meter was used by the subject to maintain this cadence. Each exercise test was terminated when pedal cadence could not be maintained at 60 rev/min (at least). Verbal encouragement was given to the subjects to continue the test until they were exhausted.

Measurements during exercise tests

Gas exchange data were collected continuously using an automated breath by breath system (CPX; Medical Graphics, St Paul, Minnesota, USA). The ventilatory threshold (VT) was determined as the power output (W) corresponding to (a) the first breakpoint in CO₂ output (Vco₂) with respect to Vo₂ (the so-called V-slope method)²⁵ and (b) the first breakpoint in pulmonary ventilation with respect to Vo₂ but not to Vco₂. Heart rate (in beats/min) was continuously monitored during the tests from modified 12 lead electrocardiogram tracings (EK56; Hellige, Freiburg, Germany).

Vo₂ kinetics

The slope (ΔVo₂/ΔW) of the regression line of the Vo₂-power output (W) relation was calculated in each subject across all workloads above the VT. Previous research has shown that Vo₂ kinetics above the VT in a ramp protocol such as the present one (a) could be used as an index of adaptation to endurance training,²¹ and (b) is related to muscle fibre distribution (percentage of type IIX fibres),¹⁴ on which ES had a significant effect, as mentioned above.

Delta efficiency

DE—that is, the ratio of the change in work accomplished/min to the change in energy expended/min—was calculated for each subject above the VT. DE was measured from linear regression (y = ax + b) of the relation between energy expended/min (y, in kcal/min), and work accomplished/min (x, in kcal/min).¹⁵ DE was equal to the reciprocal of the slope of the aforementioned relation—that is, 1/a.

Data analysis

Once a gaussian distribution of results was established by the Kolmogorov-Smirnov test, an unpaired Student’s t test was applied to examine the possible differences in ΔVo₂/ΔW and DE between control and electrostimulated groups before the onset of the experiment (week = 0). Given that no significant differences (p>0.05) were observed in either variable between the two groups of subjects at week 0, it was decided to consider these two groups as a single group (n = 15 subjects) to increase the statistical power of further analyses. Longitudinal changes (week 0 v week 6) in ΔVo₂/ΔW and DE were analysed by one way analysis of variance. When significant differences were found (p<0.05), a Fisher LSD test was used post hoc to determine which mean values were significantly different from remaining values.

The level of significance was set at 0.05 for all statistical analyses, and results are expressed as mean (SEM).

RESULTS

Figures 1 and 2 show an example of Vo₂ kinetics and DE respectively in one subject before and after the ES programme. The average values of ΔVo₂/ΔW and DE had significantly decreased and increased (p<0.05) respectively after the six week ES programme (table 1).

DISCUSSION

This study represents the first attempt to determine the effects on Vo₂ kinetics and DE of an ES programme of practical applicability—that is, short duration (30 minutes/session/day, three days/week for six weeks) and phasic, medium amount (45–60 Hz), intermittent (12 seconds on, 8 seconds off) stimulus pattern. The results indicate that the significant modifications induced by this ES programme on the histochemical and metabolic properties of human skeletal muscle fibres¹² are also reflected in an improvement in Vo₂ kinetics and DE across the moderate to high workloads (>VT) of a ramp test. Although further research is needed, this finding suggests that ES could be used as a supplementary tool to improve one of the factors involved in human adaptation to endurance training—that is, work efficiency at high exercise intensities. Furthermore, in these experiments the physiological changes were obtained with a relatively low number of ES sessions (18 in total) that were of short duration (30 minutes a day). This, together with the minimal discomfort to the patient/athlete, is essential for successful application in clinical and sport medicine situations.

Ramp protocols using an electronically braked cycle ergometer are suitable for evaluation of human endurance performance both in sport²¹ and clinically.²⁶ For instance, in this type of test, the Vo₂ response is linear above the VT (at least in cyclists who are not highly trained), and the VT shows an unequivocal and reproducible response.²⁷ In fact, ramp protocols similar to that used here have been used in research into Vo₂ kinetics and its possible determinants.^14,21,28 Exercise tests with workload increments of 10–30 W/min (20 W/min in this study) have proved invaluable for determination of work efficiency (from the Vo₂-work rate relation), and the short interval (one minute) at each work rate precludes the development of a “Vo₂ slow component”—that is, increase in Vo₂ despite constant power output—within each work rate above the VT.²⁹ In this type of ramp test, ΔVo₂/ΔW typically ranges between 9 and 11 ml O₂/W/min across most workloads, at least in athletes who are not highly trained²⁹ (average of about 9 ml O₂/W/min for our three groups of subjects taken together), which gives a DE of 25–30%³⁰ (average of about 29% for our three groups of subjects taken together). This range of 25–30% fits closely with that calculated on the basis of intracellular chemical-mechanical coupling efficiencies³¹ and only appears to be significantly altered by training status—for example, DE can clearly surpass 30% in highly endurance trained, professional cyclists (based on calculations using data from a previous study²¹).

Although Vo₂max or lactate/ventilatory thresholds are commonly used as markers of endurance performance, endurance training also induces significant changes in Vo₂ kinetics and work efficiency.¹³ The latter adaptation can be easily detected during constant load or more conventional ramp exercise tests such as the one used here. Indeed, endurance training attenuates the rate of Vo₂ rise at the high workloads (>VT) of both constant load^18–20 and gradual tests such as the one used here.²¹ Although the adaptations of central (cardiopulmonary) factors induced by endurance training may partly explain this improvement in Vo₂ kinetics during exercise testing, intramuscular changes appear to be the most important factor.¹³ Indeed, motor unit recruitment patterns—for example, lower reliance on inefficient type IIX fibres after training³²—and percentage distribution of type IIX fibres¹⁴ are thought to be the main determinants of Vo₂ kinetics above the VT. It follows that the adaptations in Vo₂ kinetics observed here after the short term ES programme (a) mimic those produced by endurance training and (b) are largely attributable to the changes in muscle fibre subpopulations (see below) given that motor unit recruitment was not altered after ES (as shown in our previous study using electromyographic data¹²). As detailed elsewhere,¹² ES induced a very significant decrease in the percentage of pure type IIX fibres of our subjects (Δ88%) and, to a much lower degree, of type I fibres (Δ15%), while the percentage of type IIA fibres increased (Δ63%). Furthermore, oxidative capacity and capillarisation of type II fibres improved with ES. The energetics of type II fibres, especially the IIX subtype (which showed the largest decrease here), can explain the findings of the study—that is, attenuation of the Vo₂ rise and improvement in DE above the VT. From both in vitro (animal)²² and in vivo (human)^14–23 studies, it appears that type IIX fibres are considerably less efficient that type I fibres for a given workload. However, it could be argued that, even if the number of inefficient type IIX fibres were decreased in our subjects after ES, the number of the most efficient fibres (type I) was also reduced and that of type IIA fibres increased.¹² This finding must be interpreted with caution. Firstly, it should be kept in mind that the capillarisation and oxidative capacity of type IIA fibres improved after ES, which in turn is expected to improve their efficiency. Secondly, although the pattern of fibre type observed with the present short term ES was IIX → IIA ← I, it would probably be shifted towards a type IIA → type I transition—that is, IIX → IIA → I—if the duration of the programme was prolonged for some more weeks.¹² This notion is also partially supported by previous studies showing a tendency towards a decreased percentage of type I fibres if ES is extended beyond six weeks.^4,10

DE is one of the main determinants of endurance performance—for example, in endurance cycling.^15,16,17 This variable is calculated as the change in energy expended per minute relative only to the change in actual work accomplished per minute, which eliminates the possible influence of metabolic processes that do not contribute to actual work—for example, basal metabolic rate.^15,31,33 Thus, the observed increase in DE over a wide range of work rates—that is, above VT or above 50–60% Vo₂max—supports the notion for a potential improvement in work efficiency after ES. These work rates were selected on the basis of the following: (a) most fibres (including both type I and II) of one of the main muscles involved in pedalling, the vastus lateralis muscle (to which ES was applied), are recruited over this range of work rates^34,35; (b) the most important phases of endurance training and competitions in general are held at these relative intensities.

In conclusion, the significant modifications that a short term (six weeks) ES programme induced in the local properties of the vastus lateralis muscle are accompanied by an improvement in Vo₂ kinetics and DE across the moderate to high workloads (>VT) of a cycle ergometer ramp test. Furthermore, the physiological changes were obtained with a relatively low number of ES sessions (18 in total) of short duration (30 minutes a day). Although further research is needed—for example, with trained athletes—this finding suggests that ES may be used as a supplementary tool to improve one of the main factors involved in human adaptation to endurance training—that is, work efficiency.