Eur J Appl Physiol

DOI 10.1007/s00421-011-1919-y

ORIGINAL ARTICLE

Maximal voluntary contraction force, SR function and glycogen resynthesis during the Wrst 72 h after a high-level competitive soccer game

Peter Krustrup · Niels Ørtenblad · Joachim Nielsen · Lars Nybo · Thomas P. Gunnarsson · F. Marcello Iaia · Klavs Madsen · Francis Stephens · Paul GreenhaV · Jens Bangsbo

Received: 21 December 2010 / Accepted: 8 March 2011© Springer-Verlag 2011

Abstract The aim of this study was to examine maximalvoluntary knee-extensor contraction force (MVC force),sarcoplasmic reticulum (SR) function and muscle glycogenlevels in the days after a high-level soccer game when play-ers ingested an optimised diet. Seven high-level male soc-cer players had a vastus lateralis muscle biopsy and a bloodsample collected in a control situation and at 0, 24, 48 and72 h after a competitive soccer game. MVC force, SR func-tion, muscle glycogen, muscle soreness and plasma myo-globin were measured. MVC force sustained over 1 s was11 and 10% lower (P < 0.05) after 0 and 24 h, respectively,compared with control. The rate of SR Ca2+ uptake at

800 nM [Ca2+]free was lower (P < 0.05) after 0 h (2.5 �molCa2+ g prot¡1 min¡1) than for all other time points (24 h:5.1 �mol Ca2+ g prot¡1 min¡1). However, SR Ca2+ releaserate was not aVected. Plasma myoglobin was sixfold higher(P < 0.05) immediately after the game, but normalised 24 hafter the game. Quadriceps muscle soreness (0–10 VAS-scale) was higher (P < 0.05) after 0 h (3.6), 24 h (1.8), 48 h(1.1) and 72 h (1.4) compared with control (0.1). Muscleglycogen was 57 and 27% lower (P < 0.001) 0 and 24 hafter the game compared with control (193 and 328 vs.449 mmol kg d w¡1). In conclusion, maximal voluntarycontraction force and SR Ca2+ uptake were impaired andmuscle soreness was elevated after a high-level soccergame, with faster recovery of SR function in comparisonwith MVC force, soreness and muscle glycogen.

Keywords Recovery · Performance · Peak force · SR Ca2+ uptake · Muscle glycogen · Myoglobin · Creatine kinase · Association football

Introduction

The amount of high-intensity running and sprinting is low-ered towards the end of high-level soccer games (Krustrupet al. 2006a, 2010; Mohr et al. 2003a), and performancetesting immediately after friendly and competitive soccergames have shown that repeated jump performance(Andersson et al. 2008; Mohr et al. 2010) and sprint perfor-mance (Krustrup et al. 2006a, 2010; Mohr et al. 2010) isdeteriorated. It has also been revealed that the functionalityof the muscles is reduced after a soccer game. Thus, maxi-mal isometric knee extensor and Xexor torque in young soc-cer players was lowered by »10% and the rate of kneeextensor force development by 5–10% following a soccer

Communicated by Håkan Westerblad.

P. Krustrup · L. Nybo · T. P. Gunnarsson · F. M. Iaia · J. BangsboDepartment of Exercise and Sport Sciences, Section of Human Physiology, University of Copenhagen, Copenhagen, Denmark

N. Ørtenblad · J. NielsenInstitute of Sports Science and Clinical Biomechanics, University of Southern Denmark, Odense, Denmark

K. MadsenDepartment of Sport Science, Aarhus University, Aarhus, Denmark

F. Stephens · P. GreenhaVSchool of Biomedical Sciences, University of Nottingham Medical School, Queen’s Medical Centre, Nottingham, UK

J. Bangsbo (&)August Krogh Building, Section of Human Physiology, Universitetsparken 13, 2100 Copenhagen Ø, Denmarke-mail: jbangsbo@ifi.ku.dk

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match (Thorlund et al. 2009). Furthermore, selective reduc-tions were observed in several EMG parameters during thestrength tests. The impairment of the muscle function is ingeneral caused by changes in both central (spinal andsupraspinal processes) and peripheral (neuromuscular prop-agation, excitation–contraction coupling, myoWbrillaractivity) mechanisms (Abbiss and Laursen 2005; Millet andLepers 2004). However, to our knowledge, no study hasexamined the recovery pattern of muscle function following ahigh-level soccer game for senior players.

In studies involving other types of physical activity, suchas marathon running and strength training, maximal voluntarycontraction force (MVC force) seems to be lowered for48 h or more (Martin et al. 2004; Petersen et al. 2007;Raastad and Hallén 2000). A reduction in muscle force pro-duction after a soccer game may also be related to animpaired skeletal muscle sarcoplasmic reticulum (SR)function, as SR Ca2+ release, activation of the contractileapparatus and SR Ca2+ reuptake are important during exer-cise, as evident from experiments with both rodents andhumans (Favero 1999; Li et al. 2002; Schertzer et al. 2004;Ørtenblad et al. 2000a). Most studies investigating SRfunction in human muscle have employed prolonged sub-maximal exercise and relatively untrained subjects, whichmay have an important bearing on the eVect of exercise onSR function. Thus, we have previously demonstrated thathigh-intensity training induced an enhanced peak SR Ca2+

release, due to an enhanced total volume of SR, whereas SRCa2+ sequestration function was not altered (Ørtenbladet al. 2000b). However, SR function is depressed to a simi-lar extent with fatigue in trained and untrained subjects,despite that trained subjects exercise at a higher intensity orlonger (Inashima et al. 2003; Li et al. 2002), indicating thattraining may delay the exercise-induced progressive deteri-oration of SR function. However, little is known about theeVect of a soccer match on SR function and the recovery ofthe SR system in the days after a match.

The development of fatigue in soccer has also beenassociated with lowered muscle glycogen and depletion ofglycogen in some muscle Wbres (Bangsbo et al. 2006;Krustrup et al. 2006a). In addition, it has been demon-strated that elevating muscle glycogen before prolongedintermittent exercise using a carbohydrate diet increasesperformance (Balsom et al. 1999; Bangsbo et al. 1992).Thus, the level of muscle glycogen appears to play a role inperformance during and after a soccer game. Sometimesgames are played within a 48- or 72-h window, and theplayers have to train in the days after a game, and the ques-tion is whether muscle glycogen is fully restored. A studyin the early 1980s with elite players of a Swedish team,Malmø FF, showed that muscle glycogen content was only50% of the pre-match value 2 days after a match (Jacobset al. 1982). These Wndings were conWrmed in a later study,

and in addition it was observed that despite the playersreceiving a high-carbohydrate diet [68 of total energyintake (E%) »9.5 g kg body mass (BM)¡1 carbohydrate]for 72 h after the game, they had only slightly highermuscle glycogen levels compared with when a normal dietwas consumed in the days after a match (Bangsbo et al.2006). It would therefore be of interest to determine thetime-course of muscle glycogen resynthesis after a high-level male soccer game under conditions with optimiseddiet (high in carbohydrates and creatine) and relate thedegree of muscle glycogen replenishment with theobserved alterations in muscle force production, SR func-tion and muscle soreness.

Thus, the present study examined maximal voluntarycontraction force, muscle SR function, glycogen synthesisand muscle soreness in the days after a high-level competitivemale soccer game.

Methods

Subjects

Seven First and Second division Danish soccer players,with a mean age, height, body mass and fat percentage of27 § 1 (§SEM) years, 180 § 3 cm, 79.5 § 2.2 kg and14.1 § 1.5%, respectively, participated in the study. Thesubjects had a VO2max of 4.53 § 0.29 L min¡1 or56.8 § 2.8 mL min¡1 kg¡1, as well as a resting and peakheart rate of 41 § 3 and 185 § 3 bpm, respectively, and apeak ventilation of 148 § 7 L min¡1. The mass of the rightand left quadriceps was estimated to be 2.62 § 0.07 and2.65 § 0.09 kg, respectively (Krustrup et al. 2004). Noneof the subjects had taken creatine over the last 2 years. Allplayers were outWeld players, i.e. two defenders, twocentral midWelders, two lateral midWelders and oneattacker. The players were fully informed of experimentalprocedures and possible discomfort associated with thestudy before giving their written informed consent to par-ticipate. The study was carried out in accordance with theguidelines contained in the Declaration of Helsinki and wasapproved by the local ethical committee of Copenhagen.

Experimental design

All subjects were followed during the last competitivegame of the season, during which locomotor activities,heart rate, weight loss and Xuid intake were determined.Immediately after the game as well as 24, 48 and 72 h afterthe game a biopsy was obtained from m. vastus lateralisand an arm venous blood sample was collected. In addition,maximum voluntary contraction isometric force (MVCforce) was determined at 0, 24, 48 and 72 h and a Yo–Yo

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intermittent recovery level 2 test (Yo–Yo IR2), and a3 £ 30 m sprint test was performed after 72 h of recovery.

Under control conditions in the last 3 weeks of theseason, 64 h after the latest training session, an anticubitalvein blood sample, resting heart rate measurements andperformance testing were carried out and for two of sevenplayers also a muscle biopsy from m. vastus lateralis. ForWve of the seven players, the control biopsy was collected5 days (120 h) after the analysed match. It was proposedthat biopsies were collected immediately prior to the high-level competitive match but this was not accepted by theclubs. Throughout the 120-h recovery period the playersconsumed a controlled diet (see below).

Dietary intake

The players received a controlled diet over the Wrst 5 daysafter the match, consisting of 9.5, 2.7 and 0.8 g kg bodymass¡1 day¡1 of carbohydrates, proteins and fats, respec-tively, corresponding to 67, 20 and 13 E%. The playersconsumed 450 § 39 g of carbohydrates, 130 § 9 g ofprotein and 32 § 10 g of fat on the match-day after thegame; 750 § 48 g of CHO, 210 § 8 g of protein and55 § 3 g of fat on the Wrst day after the match; 725 § 48 gof CHO, 220 § 9 g of protein and 70 § 14 g of fat on thesecond day after the match; and 265 § 34 g of CHO,80 § 7 g of protein and 21 § 2 g of fat prior to the 72-htesting on the third day after the match. The diet was sup-plemented with 0.20, 0.25 and 0.20 g creatine (Cr) kg¡1

body mass at the Wrst, second and third 24-h period after thematch, respectively. Portions of creatine (»4 g) werediluted in 250 mL of skimmed milk and 150 mL of hotwater together with 30 g of maltodextrin (Maxim, UK) and15 g of protein powder (12 g casein protein). The bottleswere placed in a refrigerator until consumption. The energycontent of each 0.5 L bottle was 1,130 kJ. The playersdrank two bottles on the match-day, four bottles on the Wrstand second day after the match and two bottles on the thirdday. A combination of creatine and carbohydrate ingestionafter has been shown to augments muscle glycogen storageafter exercise compared with carbohydrate ingestion alone(Robinson et al. 1999). Therefore, creatine was added to thediet in the present study in order to optimise the dietaryconditions for recovery. The players drank water ad libitumafter the game. The Wrst dose of creatine and carbohydrateswas provided 30 min after the match, and the Wrst meal(consisting of 200 g of pasta, 100 g of vegetables and 50 gof ham) was provided »60 min after the match, after termi-nation of the testing procedures.

The players registered their diet 3 days prior to the match.During this period the intake of carbohydrates, proteins andfats was 5.0, 1.6 and 1.1 g kg body mass¡1 day¡1, respec-tively, corresponding to 55, 18 and 26 E%, respectively.

Procedures

Prior to the match, the players refrained from strenuousexercise and intake of alcohol for 48 h as well as fromtobacco and caVeine for 12 h. The game started early in theafternoon (2–4 p.m.). A POLAR heart rate monitor(POLAR Electro, Kempele, Finland) consisting of a chestbelt and wrist receiver weighing »100 g was placed oneach subject »45 min prior to the game. In addition, theplayers were weighed wearing dry shorts »10 min beforethe match using a digital scale (OHAUS 1–10, New Jersey,USA). The players had individual water bottles preparedand were allowed to drink water ad libitum during the gameand the water intake was recorded.

Match activities

The players were video-Wlmed individually close up duringthe entire match. The VHS movie cameras (NV-M50, Pan-asonic, Germany) were positioned at the side of the pitch, atthe level of the midWeld line, at a height of about 15 m andat a distance of 30–40 m from the touchline. The videotapeswere later replayed on a monitor for computerised codingof the activity pattern. The following locomotor categorieswere used: standing (0 km h¡1), walking (6 km h¡1),jogging (8 km h¡1), low-speed running (12 km h¡1),moderate-speed running (15 km h¡1), high-speed running(18 km h¡1), sprinting (25 km h¡1) and backwards running(10 km h¡1). The locomotor categories were chosen inaccordance with Bangsbo et al. (1991) and Mohr et al.(2003a). The frequency and duration of each activity wererecorded. The distance covered for each activity was deter-mined as the product of the total time and mean speed forthat activity. The total distance covered during a match wascalculated as the sum of the distances covered during eachtype of activity.

Post game measurements

Immediately after the game, the players walked to a dress-ing room within 40 m from the pitch. The players wereweighed in dry shorts after which they were positioned in asupine position on a bed. A catheter (18G, 32 mm) wasplaced in an antecubital vein for collection of blood sam-ples in two 5-mL syringes with and without heparin,respectively. These samples were taken »10 min after thegame. Local anaesthesia (20 mg mL¡1 lidocain withoutadrenalin) was given and a skin incision was made abovem. vastus lateralis. Approximately 15 min after the game, aneedle biopsy with suction was obtained. Approximately45 min after the game the players were transported adistance of 0.5–4 km to the Department of Exercise andSport Sciences, University of Copenhagen, for testing

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procedures. Additional measurements were made at theUniversity facilities after 24, 48 and 72 h.

Muscle analysis

A part of the muscle tissue was immediately frozen inliquid N2 and stored at ¡80°C. Another part (»40 mg) wasdiluted in 10 volumes (weight.volume¡1) ice-cold homog-enising buVer (300 mM Sucrose, 1 mM EDTA, 10 mMNaN3, 40 mM Tris-base, 40 mM L-Histidin, pH 7.8). Then,the muscles sample was mechanically homogenised with aDuall glass–glass hand homogeniser (Kontes Class, Duall20), frozen in liquid N2 and stored at ¡80°C until analysedfor crude homogenate SR vesicle Ca2+ uptake and releaserate.

Muscle SR vesicle Ca2+ uptake and release

The technique for measuring SR vesicle Ca2+ uptake andrelease has been described previously (Nielsen et al. 2005,Ørtenblad et al. 2000a). BrieXy, the SR vesicle, oxalate-mediated Ca2+ uptake and release were measuredXuorometrically (Ratiomaster RCM, Photon TechnologyInternational, Brunswick, NJ, USA) at 37°C using aXuorescent Ca2+ indicator (indo-1). All raw-data of Ca2+

release and uptake were mathematically Wtted using monoexponential equations as previously described (Nielsenet al. 2005). The ATP-supported SR Ca2+ uptake rate wasestimated as both the tau (�) constant, which is the timeconstant (inverse rate constant) for Ca2+ uptake and repre-sents the time to reduce free [Ca2+] by 63%, and as theuptake rates at the physiological concentrations of 600 and800 nM free Ca2+ ([Ca2+]free). Following the plateauing ofthe [Ca2+]free, the SR vesicle Ca2+ release was induced byaddition of 1 mM 4-chloro-M-Cresol. The values obtainedfor SR Ca2+ uptake and -release rates are relative and there-fore expressed as arbitrary units Ca2+ min¡1 g¡1 protein.

Muscle glycogen

One portion of the frozen sample was freeze-drieddissected free of blood, fat and connective tissue. Muscleglycogen was determined using a modiWed version of thespectrophotometric method of Harris et al. (1974).

Quadriceps muscle soreness

Perceived muscle soreness was assessed using a 10-cm linearanalogue scale with labels that corresponded to “no pain”and “extreme pain” on either end. After 30 m of walking ata speed of 1.5 m s¡1, the subjects marked their level ofsubjective pain in the quadriceps muscle of the non-biopsiedleg. The pain was marked along the continuum and the

distance was reported as the raw score. The visual analoguescale has been established as a reliable method for assess-ing pain (Mohr et al. 2003b).

Plasma CK and myoglobin

Within 10 s of sampling the blood was centrifuged for 30 s.From this, plasma was collected and stored at ¡20°C untilanalysed for creatine kinase (CK) and myoglobin on aCobas Mira AutoAnalyser as previously described(Schumann et al. 2002).

Performance testing

MVC measurements

Maximal voluntary contraction force was evaluated aspreviously reported (Nielsen et al. 2005). BrieXy, players wereplaced in a custom-built chair, with an adjustable support forthe back adjusted for a knee angle of 90°. Trunk and hip wereWrmly Wxed using three belts: one around the chest, one justbelow spina ilica antero-superior and the third going as a Ybetween the legs to above the shoulders. The experimental legwas Wxed (2 cm proximal the malleoli) in a metal cuV con-nected to a strain-gauge (Darcus 1953). Data were sampled at1 kHz using Spike 2 version 4.18 (Cambridge ElectronicDesign, Cambridge, UK) and through a CED Micron 1401 II16 bit AD-converter (CED, Cambridge, UK). After 10 min ofwarm-up (80 W on a cycle ergometer), each player performedthree knee extensions at maximal voluntary eVort separated by45 s. Each MVC was sustained for »3 s and strong verbalencouragement was given during every contraction to promotemaximal voluntary eVort. Peak force and 1 s sustainable forcewere measured using Spike 2.

Yo–Yo intermittent recovery level 2 test

The Yo–Yo intermittent recovery level 2 (Yo–Yo IR2) testconsists of repeated 20-m shuttle runs at progressivelyincreasing speeds dictated by an audio bleep emitted from aCD player (Krustrup et al. 2006b). Between each shuttle theplayers had a 10-s period of active recovery, during whichthe players jogged around a marker placed 5 m behind thestarting line. Failure to achieve the shuttle run on two suc-cessive occasions resulted in termination of the test andthe distance covered was recorded and represented thetest result. All testing sessions were performed indoors on2-m £ 20-m running lanes marked by cones.

Repeated sprint test

All players performed a repeated sprint test in recoveredstate as well as 72 h after the game. The sprint test

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consisted of three 30-m sprints, separated by a 25-s periodof active recovery, during which the players jogged back tothe starting line (Krustrup et al. 2006a, 2010). The sprinttimes were recorded by infra red light sensors, having aprecision of 0.01 s (Time It, Eleiko Sport, Halmstad,Sweden).

Statistics

Changes in muscle and blood variables as well as perfor-mance measures were evaluated by the one-way analysis ofvariance (ANOVA) on repeated measurement. When asigniWcant interaction was detected, data were subsequentlyanalysed using a Newman-Keuls post-hoc test. A signiWcance

level of 0.05 was chosen. Data are presented asmean § standard error of the mean (SEM).

Results

Activity proWle, heart rate and Xuid loss during game

The total distance covered during the match was11.31 § 0.22 km of which 1.68 § 0.09 km was covered withhigh-intensity running (>15 km h¡1) and 0.44 § 0.14 km wascovered with backwards running. The number of high-inten-sity and backwards running bouts were 170 § 18 and 66 § 16,respectively. The players were standing, walking and joggingfor 9.3 § 0.9, 49.3 § 2.6 and 21.4 § 2.5% of the total time,respectively, and performed low-, moderate- and high-speedrunning for 10.6 § 0.8, 4.2 § 0.4 and 1.8 § 0.2% of the totaltime, respectively, with sprint and backwards running account-ing for 0.5 § 0.1 and 2.9 § 0.9%, respectively, of the totaltime.

The heart rate during the Wrst, middle and last 15 min ofWrst half was 159 § 4, 159 § 5 and 158 § 3 bpm, respec-tively, with corresponding values for second half being157 § 3, 154 § 3 and 157 § 3 bpm, respectively. The val-ues represented 83–86% of HRmax (185 § 3 bpm).

The weight loss during the game was 1.9 § 0.4 kg, cor-responding to 2.3 § 0.4% of body mass. Estimated Xuidloss during the game was 2.9 § 0.6 L during the game, and1.1 § 0.3 L of water was consumed during the game.

Performance after game

MVC force sustained over 1 s was lowered (P < 0.05) by11% after the game and by 10% (P < 0.05) 24 h after thegame compared with control (Table 2). Peak MVC forcewas 9 and 8% lower (P < 0.05) 0 and 24 h after the game,respectively, compared with control. Yo–Yo IR2 perfor-mance 72 h after match was 731 § 53 m and not diVerentfrom the control value (800 § 68 m). Similarly, after 72 hthe mean time of the 30-m sprints was 4.49 § 0.07 s andthe same as in the control situation (4.44 § 0.03 s;Table 2).

SR vesicle Ca2+ uptake and release

SR Ca2+ uptake, determined as � and rates at 600 and800 nM [Ca2+]free, was (P < 0.05) lowered immediatelyafter the match (Fig. 1a; Table 1). In the Wrst 24 h after thematch � decreased (P < 0.05) from 53.5 § 8.7 to32.3 § 3.5 s and did not change in the following 48 h(Fig. 1a). Similarly, the rate of SR Ca2+ uptake at both 600and 800 nM [Ca2+]free increased in the Wrst 24 h from 2.5and 1.8 �mol Ca2+ g prot¡1 min¡1, respectively, to 5.1 and

Fig. 1 Sarcoplasmic reticulum vesicle Ca2+ uptake rate after a high-level competitive soccer game, expressed as a the time constant Tau(inverse rate constant) for Ca2+ uptake representing the time to reducefree [Ca2+] by 63% and b SR vesicle Ca2+ uptake rate at 800 nM[Ca2+]free. Values represent mean § SEM (n = 7). $ denotes diVerencefrom control, 24, 48 and 72 h

Control 0 h 24 h 48 h 72 h

SR

rat

e of

Ca2+

upt

ake

(um

ol·g

pro

t-1·m

in-1 )

0

1

2

3

4

5

6

7

$

Control 0 h 24 h 48 h 72 h

SR

rat

e of

Ca2+

upt

ake

(tim

e co

nsta

nt; s

)

0

10

20

30

40

50

60

70

$

A

B

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3.8 �mol Ca2+.g¡1 prot¡1 min¡1 and remained constant inthe following days (Fig. 1b, Table 1). There were no diVer-ences in nadir [Ca2+]free, i.e. the [Ca2+] at which uptake pla-teau, averaging 60 § 17 nmol [Ca2+]free. SR vesicle Ca2+

release rates following the match and in the recovery periodafter the match were the same, averaging 4.4 § 0.4 �molCa2+ g prot¡1 min¡1 (Table 1).

Muscle glycogen after game

Muscle glycogen content immediately after the game was193 § 22 mmol kg dw¡1, which was 43% of the controllevel (449 § 34 mmol kg dw¡1, P < 0.001; Fig. 2). After24 h, muscle glycogen was 27% lower (P < 0.001) thancontrol, whereas the glycogen level was not signiWcantlydiVerent from control after 48 h (9%, P = 0.096; Fig. 2).

Muscle soreness and plasma levels of enzymes

Muscle soreness score was 3.6 § 0.9 on the 0–10 VASscale immediately after the match and higher (P < 0.05)than the control during the entire 72-h period after thematch (Table 2). The plasma level of the skeletal muscleenzyme creatine kinase (CK) and myoglobin (Mb) wereused as a marker of the level of muscle injury. The plasma

level of Mb was elevated (P < 0.05) by more than sixfoldimmediately after the game (Table 2), but normalised 24 hafter the game. Plasma CK was not signiWcantly elevatedafter the game and in the recovery period after the game;however, when expressed as relative to control value a 2.5-fold increase (P < 0.05) in CK was observed after 24 hrecovery (Table 2).

Table 1 SR vesicle Ca2+ uptake and release after a high-level competitive soccer game

Data are mean § SEM (n = 7, except for 24 h; n = 5)$ DiVerent (P < 0.05) from control, 24, 48 and 72 h

Recovery (h) Control 0 24 48 72

Rate of Ca2+ uptake at 600 nM Ca2+ (�mol Ca2+ mg prot¡1 min¡1) 2.6 § 0.3 1.8 § 0.3$ 3.8 § 0.7 3.0 § 0.3 2.5 § 0.1

Rate of Ca2+ uptake at 600 nM Ca2+ (%con) 100 71 § 12$ 145 § 25 114 § 13 96 § 5

Rate of Ca2+ uptake at 800 nM Ca2+ (%con) 100 77 § 5$ 154 § 26 122 § 14 103 § 5

SR Ca2+ release rate (�mol Ca2+ mg prot¡1 min¡1) 3.8 § 0.6 3.8 § 0.5 5.2 § 0.6 4.3 § 0.7 4.1 § 0.5

SR Ca2+ release rate (%con) 100 95 § 11 139 § 18 111 § 20 103 § 13

Fig. 2 Muscle glycogen under control conditions and in recovery aftera high-level competitive soccer game. Values represent mean § SEM(n = 7). * denotes diVerent (P < 0.001) from control. # denotes diVer-ent (P < 0.05) from 72 h

Control 0 h 24 h 48 h 72 h

Mus

cle

glyc

ogen

(m

mol

·kg

dw-1

)

0

100

200

300

400

500

600

#*

#*

#

Table 2 M. Quadriceps maximal voluntary contraction force (MVC force), muscle soreness as well as plasma CK and myoglobin before and aftera high-level competitive soccer game

Data are mean § SEM (n = 7)

* DiVerent (P < 0.05) from control# DiVerent (P < 0.05) from 72 h

Recovery (h) Control 0 24 48 72

MVC peak force (N) 596 § 32 542 § 37* 551 § 31* 562 § 38 574 § 31

MVC force 1-s sustain (N) 571 § 33 509 § 41* 516 § 37* 534 § 37 546 § 31

Muscle soreness (0–10 VAS scale) 0.1 § 0.1 3.6 § 0.8* 1.8 § 0.6* 1.1 § 0.8* 1.4 § 0.4*

CK (U l¡1) 195 § 26 425 § 111 491 § 152 361 § 100 288 § 59

CK (%con) 100 205 § 43 244§ 50*# 179 § 35 107 § 7

Myoglobin (ug l¡1) 36 § 3 228 § 47* 40 § 6 41 § 5 37 § 3

Mb (%con) 100 665 § 188*# 124 § 22 118 § 12 103 § 3

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Discussion

The major Wndings of the present study were that muscleforce and SR function are deteriorated after a high-levelcompetitive male soccer game and that the time-course ofrecovery varies for muscle force, SR function and muscleglycogen. Thus, the rate of SR Ca2+ uptake was recovered24 h after the game, whereas muscle force and muscle gly-cogen were still attenuated. In addition, the present studyprovided further evidence that glycogen resynthesis is slowafter a high-level soccer game even when players digest anoptimised diet rich in carbohydrates and creatine.

In the present study, seven high-level soccer playerswith a representative soccer-speciWc Wtness level (Yo–YoIR2 scores of »800 m; Krustrup et al. 2006b) participatedin competitive matches where the total distance coveredwas 10–12 km, average heart rate was 85% HRmax and netXuid loss was »2 l, which is similar to previous reports onDanish high-level and top-class players (Mohr et al. 2003a,2004; Krustrup et al. 2006a; Randers et al. 2009). The aver-age number of high-intensity running bouts and backwardsrunning bouts was »170 and »65, respectively, which ismore than reported for fourth division players (Mohr et al.2004; Krustrup et al. 2006a) and slightly less than for Dan-ish Premier League players (Mohr et al. 2003a; Randerset al. 2009). Moreover, muscle glycogen reached post-game values of 198 (range: 129–299) mmol kg d w¡1 cor-responding to 43 (36–51)% of control levels, which aresimilar to previously reported data for Danish Wrst division(Bangsbo et al. 2006) and fourth division matches (Krust-rup et al. 2006a).

For the high-level players participating in the competi-tive soccer game, the SR vesicle Ca2+ uptake was impairedimmediately after the game, but was fully recovered within24 h recovery. On the other hand, the game had no eVect onthe SR vesicle Ca2+ release rate. This is in line with moststudies using SR vesicle from in vivo exercise protocols(Duhamel et al. 2006a, b; Li et al. 2002). In skeletal musclethere is generally a slowing of relaxation with increasingdevelopment of fatigue and it has been suggested that thiscan be explained by altered crossbridge kinetics andimpaired Ca2+ reuptake (Westerblad and Allen 1994;Westerblad et al. 1997). Thus, the functional consequenceof an impaired SR Ca-ATPase would be a slowing of relax-ation, i.e. impaired muscle function during high-intensitydynamic contractile activity, with repeated contractions andrelaxations. In line with this, it has previously been demon-strated that repeated jump and sprint performances are low-ered immediately after a soccer game (Andersson et al.2008; Krustrup et al. 2006a, 2010; Mohr et al. 2010).Therefore, it is possible that muscle Ca2+ handling mayhave negatively inXuenced performance during and afterthe game.

The underlying mechanisms for the impaired Ca2+ pumpfunction are not recognised. However, the lowered muscleglycogen level at the end of the game may have aVected theSR function. Thus, Duhamel et al. (2006a, b) have shownthat there is an association between SR vesicle function andmuscle glycogen content, if the latter is below a criticallevel. This notion has recently been supported and furtherextended in a study with highly trained cross country ski-ers, demonstrating that the SR vesicle Ca2+ release rate isassociated with a speciWc pool of glycogen in the intramyo-Wbrillar compartment (Ørtenblad et al. 2011). Furthermore,the existence of a glycogenolytic complex that is physicallyassociated within muscle SR is well established, and thiscomplex persists in the crude homogenate used in the SRfunction assay (Duhamel et al. 2006b; Wanson and Droch-mans 1972). In line, we have recently shown that inmechanically skinned Wbres, the distinct pool of intermyo-Wbrillar glycogen is inversely correlated with muscle half-relaxation time (Nielsen et al. 2009). Thus, the SR Ca2+

pumps are controlled, at least partly, by the glycogen avail-ability in speciWc locations in the myoplasm. However, inthe present study on high-level soccer players, SR vesiclefunction is perturbed at higher glycogen levels as seen dur-ing isometric and concentric exercise (Duhamel et al.2006a, b; Nielsen et al. 2009), which may be explained by ahigh degree of eccentric contractions and multiple intenserunning bouts as determined from the time-motion analy-ses.

The muscle glycogen stores had not reached the pre-game level 24 h, conWrming that glycogen resynthesis isimpaired after a soccer game as earlier observed both with anormal and carbohydrate enriched diet (Bangsbo et al.2006). The rate of glycogen synthesis after the game was5.6 and 3.5 mmol kg d w¡1 h¡1 the Wrst 24 h and next 24 h,respectively, which was not signiWcantly diVerent from therates observed with trained soccer players digesting a car-bohydrate-rich diet in the recovery period after a game (6.9and 2.1 mmol kg d w¡1 h¡1, Bangsbo et al. 2006). Somestudies have shown that creatine intake together with anelevated carbohydrate ingestion does augment glycogenstorage after exercise (Robinson et al. 1999). However,despite an optimal timing of the Wrst CHO intakes (30 and60 min after the game) and a combination of CHO and cre-atine that should have the largest eVect on muscle glycogen(Cribb and Hayes 2006), the glycogen resynthesis was notfully restored after 24 h.

Apparently, other factors than substrate availability arecausing the impaired glycogen resynthesis. It may berelated to the activities in the game since a number ofstudies have shown that muscle glycogen is fullyrestored 24–30 h after prolonged continuous exercise, suchas cycling and running (Kiens and Richter 1998; Zehnderet al. 2004). In these studies, the ingestion of carbohydrate

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(»9 and 10 g carbohydrate kg BM¡1 day¡1 correspondingto »67 and 80 E%, respectively) was similar to that of thepresent study (9.5 g carbohydrate kg BM¡1 day¡1 or »68E%). The game may have caused muscle damage restrict-ing the glycogen synthesis (Asp et al. 1998; Zehnder et al.2004), which is in part supported by the observation thatthe players had elevated Mb values after the game as foundafter eccentric and strenuous exercise, damaging skeletalmuscle structures (Noakes 1987; Zehnder et al. 2004;Ørtenblad et al. 1997). This notion is supported by the pres-ent Wndings of elevated muscle soreness several days afterthe game.

In summary, maximal force production and muscle SRCa2+ reuptake was reduced after the game; the latter and notthe former was recovered 24 h after the game Additionally,muscle glycogen resynthesis was impaired during the Wrst24 h after a soccer game even when ingesting a diet high incarbohydrates and creatine, which may have been causedby the muscle damage that appears to have occurred duringthe game.

Acknowledgments The authors would like to acknowledge the play-ers and their elite soccer clubs for the participation. They would alsolike to thank the Danish Football Association (Dansk Boldspil-Union)for their cooperation. The excellent technical assistance by Jens JungNielsen and Christian Hasson is appreciated. Special thanks to thecamera crew led by Per Hansen and ChristoVer Krustrup. The studywas supported by Arla Foods, Team Denmark and The Danish Minis-try of Culture (Kulturministeriets Udvalg for Idrætsforskning).

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