© 2015 John Wiley & Sons A/S.

Scand J Med Sci Sports 2015: 25 (Suppl. 1): 112–125 doi: 10.1111/sms.12367

Published by John Wiley & Sons Ltd

Hydration and endocrine responses to intravenous fluid and oral glycerol S. P. van Rosendal1, N. A. Strobel1, M. A. Osborne1,2, R. G. Fassett1, J. S. Coombes1 1

Human Performance Laboratory, School of Human Movement Studies, The University of Queensland, Brisbane, Queensland, Australia, 2Queensland Academy of Sport, Brisbane, Queensland, Australia Corresponding author: Jeff S. Coombes, PhD, School of Human Movement Studies, The University of Queensland, St Lucia, Queensland 4072, Australia. Tel: +61-(7)-33656767, Fax: +61-(7)-33656877, E-mail: [email protected] Accepted for publication 6 October 2014

Athletes use intravenous (IV) saline in an attempt to maximize rehydration. The diuresis from IV rehydration may be circumvented through the concomitant use of oral glycerol. We examined the effects of rehydrating with differing regimes of oral and IV fluid, with or without oral glycerol, on hydration, urine, and endocrine indices. Nine endurance-trained men were dehydrated by 4% bodyweight, then rehydrated with 150% of the fluid lost via four protocols: (a) oral = oral fluid only; (b) oral glycerol = oral fluid with added glycerol (1.5 g/kg); (c) IV = 50% IV fluid, 50% oral fluid; and (d) IV with oral glycerol = 50% IV fluid, 50% oral fluid with added glyc-

erol (1.5 g/kg), using a randomized, crossover design. They then completed a cycling performance test. Plasma volume restoration was highest in IV with oral glycerol > IV > oral glycerol > oral. Urine volume was reduced in both IV trials compared with oral. IV and IV with oral glycerol resulted in lower aldosterone levels during rehydration and performance, and lower cortisol levels during rehydration. IV with oral glycerol resulted in the greatest fluid retention. In summary, the IV conditions resulted in greater fluid retention compared with oral and lower levels of fluid regulatory and stress hormones compared with both oral conditions.

Intravenous (IV) fluid rehydration is an attractive technique for use by some athletes during and/or following competition. Despite being banned by the World AntiDoping Agency (WADA) in 2005 (under Section M2 of their list of prohibited substances and methods), and having risks that are not associated with oral fluid intake (e.g., infection, thrombophlebitis, air embolus, needle stick injury, bleeding, hematoma, soft tissue extravasation), the ongoing use of IV infusions is highlighted by a recent survey finding that 75% of National Football League (United States) teams reported regularly infusing 1.5 L of normal saline in up to 20 players, 2 h before games (Fitzsimmons et al., 2011). A major ergogenic appeal of IV fluids is the subsequent rapid enhancement of plasma volume which is achieved since the fluids bypass the gastrointestinal absorption delays associated with oral rehydration (Casa et al., 2000; Kenefick et al., 2006). The higher circulating plasma volume theoretically allows enhanced thermoregulation and attenuated cardiovascular strain. In spite of this, previous reports indicate that the rapid influx of fluid into the vasculature following IV rehydration is transient and equilibrates between the fluid compartments within 35 min during passive rehydration (Kenefick et al., 2000), or between 5 and 25 min when exercise begins soon after rehydration is completed (Casa et al., 2000; Kenefick et al., 2000, 2006; Maresh et al., 2001), regardless of whether hypo-

tonic or isotonic IV fluids are used (Kenefick et al., 2000). As a result, these studies have shown few sustained benefits with IV compared with oral rehydration. However, dehydration should be considered from a whole-body perspective since the majority of the fluid deficits occur from intracellular and interstitial spaces exclusive of plasma volume. When dehydrated by 2–4% bodyweight, the extracellular and intracellular compartments contribute approximately 60% and 40% of the fluid lost, respectively (Nose et al., 1988b). Thus, it is imperative that rehydration occurs to the extravascular spaces. Replenishment of these compartments also tends to be more rapid following IV compared with oral rehydration. Here, the downside of IV rehydration is that the rapid increases to intravascular fluid volumes also promote a greater production of urine and therefore an overall greater loss of infused fluids relative to fluids taken orally (Kenefick et al., 2006). Theoretically, the provision of an osmotic agent (e.g., an oral sports drink containing glycerol) with the IV fluid will enhance the retention of the fluid and improve rehydration. Glycerol is an osmotically active solute that promotes fluid retention and reduces urine output when given mixed with an oral fluid bolus (van Rosendal et al., 2010a). Most glycerol hydration research has provided glycerol with fluid as a method to hyperhydrate preexercise since it disperses throughout the entire total

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Rehydration with IV fluid and oral glycerol body water and therefore promotes fluid retention in all spaces. An additional three studies have investigated the addition of glycerol to rehydration solutions and all found that beverages containing glycerol were associated with significantly more rapid and complete restoration of plasma volume than water alone (Scheett et al., 2001; Magal et al., 2003; Kavouras et al., 2006). Thus, the aim of this study was to test the effects of IV and oral rehydration regimes, with and without glycerol, on rehydration following acute exercise-induced dehydration. We investigated subsequent biochemical and hormonal variables during exercise in a hot/humid environment. It was hypothesized that: (a) combining IV rehydration with oral glycerol would result in the greatest fluid retention of the four protocols; (b) glycerol would reduce urine output relative to the same trial without glycerol; and (c) the enhanced hydration in IV trials would be associated with lower aldosterone, antidiuretic hormone (ADH), and cortisol concentrations. The effects of these rehydration regimes on subsequent exercise performance have been reported elsewhere (van Rosendal et al., 2012). Performance was significantly (P < 0.05) improved when rehydrating with oral glycerol (improved time to complete 40 km time trial by 3.7%), IV (3.5%), and IV with oral glycerol (4.1%), compared with the oral rehydration trial (van Rosendal et al., 2012). Methods Ethical approval Subjects were informed of all the risks and stresses of the study, and gave their written informed consent. The University of Queensland Medical Research Ethics Committee approved the study.

Subjects Nine endurance-trained male cyclists took part in this study. Their mean (SD) physical characteristics were: age 22.8 (3.9) years; bodyweight 74.1 (4.9) kg; height 179.2 (4.7) cm; VO2max 64.8 (5.7) mL/kg/min. All subjects trained regularly and were non-smokers. Inclusion criteria for the study were: (a) male aged 18–40 years; (b) experienced cyclist (raced competitively in cycling and/or triathlon events, of equivalent to B grade or better, for an average of 6 races per year for the last 3 years); (c) high endurance training status (VO2max > 55 mL/kg/min); and (d) consistently high training volumes (minimum of 140 km cycling and/or 70 km running per week) for at least the preceding 2 months. Based on data collected in a pretrial screening questionnaire, subjects were excluded from participating in the study if they reported: (a) a history of current or previous renal, hepatic, cardiovascular, thermoregulatory, or endocrine disorders; (b) contraindications to exercising in the heat; (c) any current or chronic health problems or injuries; (d) the use of any diuretic during the preceding 4 weeks; and (e) having made a blood donation in the preceding 3 months. Subjects had their trial session rescheduled if they: (a) presented with an abnormally high temperature (> 38 °C); (b) reported dehydrated based on their baseline urine osmolality reading; or (c) failed to abide by the pretrial conditions detailed in the pretrial preparation section. None of the subjects were taking any nutritional or performance supplements during the course of the study.

Graded exercise test Each subject initially completed a graded cycle ergometer test to volitional exhaustion to determine their maximal oxygen uptake (VO2peak) (van Rosendal et al., 2012). The cycle ergometer used for the VO2max test and all subsequent trials was their own road bike mounted on a stationary wind trainer (Cyclosimulator CS-1000; Cateye Co. Ltd., Osaka, Japan), as described previously (van Rosendal et al., 2012). Therefore, the bike setup was identical for all trials.

Familiarization trial and performance test reliability At least 1 week after the VO2max test, subjects completed a familiarization trial of the performance test to be used during the experimental trials. For the first 30 min, subjects cycled at a self-selected intensity and cadence with a heart rate corresponding to 85% of their ventilatory threshold. During this stage, heart rate (used to maintain intensity) was the only variable on display to the subjects. They were informed when they passed 15 min and were given 0.5 mL/kg of water at the 15 and 25 min marks. After 30 min they stopped cycling for 5 min and were allowed to dismount. This break was included to keep the protocol consistent with that used in the experimental trials where subjects were given the chance to void before starting the time trial. They then remounted and completed a 40-km time trial. Subjects were given 0.5 mL/kg of water after 5, 15, 25, and 35 km and were informed when they passed 10, 20, 30, 35, 36, 37, 38, 39, and 40 km. All performance variables including speed, distance, heart rate, and time were removed from their view. No specific encouragement was given to the subjects at any stage during the familiarization trial or performance tests. The room temperature and humidity for familiarization trials were 34 °C and 60% relative humidity.

Pretrial preparation Subjects kept a food diary for 3 days preceding each experimental trial and were asked to reproduce this diet as consistently as possible before each subsequent trial. Food diaries were analyzed for total energy intake and for the intake of protein, fat (total, saturated, polyunsaturated, and monounsaturated), cholesterol, carbohydrate, water, sodium, and potassium (Foodworks Professional 2007, Xyris Software (Australia) Pty Ltd.). Subjects were asked to refrain from strenuous training for 48 h, and alcohol and caffeine for 24 h prior to each experimental trial. During this preceding 24-h period, subjects were asked to consume at least 2 L of water. On the morning of each experimental trial, subjects were instructed to ensure euhydration by consuming a further 500 mL of plain tap water upon waking. They also consumed a standardized breakfast comprising two slices of multigrain toast, two sachets of butter, two sachets of honey, a yoghurt top muesli bar, and an individualized volume of Sustagen®, Nestle Australia, Tongala, Victoria, Australia to make a total caloric intake of 70 kJ/ kg. These measures were employed to minimize variability in preexperimental hydration, hormonal and metabolic status between subjects and treatments.

Experimental protocol At least 1 week after the familiarization trial, subjects reported to the laboratory between 07:00 and 08:00 h for their first experimental trial. Subsequent trials commenced at the same time for each individual subject to control for circadian variations and were conducted at least 2 weeks apart to limit the change in heat acclimation of individual subjects. Each subject completed four experimental trials which differed only by the rehydration treatments provided. The four experimental treatments were: (a) 100% oral

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van Rosendal et al.

Fig. 1. Experimental protocol for dehydration, rehydration, and exercise performance. Vertical lines indicate time points at which blood samples (A, 16 mL; B, 4 mL; C, 12 mL; D, 6 mL) were taken and outcome measures were assessed. RH = relative humidity, BW = bodyweight, Thirst = thirst sensation. fluid (of which 64% was carbohydrate-electrolyte sports drink and 36% was distilled water; oral); (b) 100% oral fluid with oral glycerol (of which 64% was carbohydrate-electrolyte sports drink with a total dose of 1.5 g/kg glycerol and 36% was distilled water; oral glycerol); (c) 50% oral fluid (of which 88% was carbohydrateelectrolyte sports drink and 12% was distilled water), 50% IV fluid (0.9% NaCl; IV); and (d) 50% oral fluid (of which 88% was carbohydrate-electrolyte sports drink with a total dose of 1.5 g/kg glycerol and 12% was distilled water), 50% IV fluid (0.9% NaCl; IV with oral glycerol). As detailed in the rehydration section, the differences in the relative amounts of oral fluid given as sports drink between the trials were due to maintaining a constant experimental solution volume for all conditions (even though the total volume of oral fluid was halved in the IV trials). These rehydration volumes were based on the American College of Sports Medicine’s (Sawka et al., 2007) recommendation of 150% of fluid lost during dehydration. The combinations of sports drink/water and IV fluids were based on protocols anecdotally used by professional sporting teams at the time of study design and because subjects reported feeling nauseous during pilot trials when trying to consume the whole rehydration volume as Gatorade®, Schweppes Australia, Tullamarine, Victoria, Australia. Trials were run in a randomized order predetermined using a computer-based random number generator. An individual not involved with the study conducted the randomization and allocation. Treatments were given in a doubleblind manner in regard to glycerol ingestion. None of the subjects had consumed glycerol before participating in the study. Figure 1 outlines the experimental protocol.

Baseline measurements Subjects were asked not to urinate in the morning before arriving at the laboratory. Upon arrival, subjects provided a urine sample for the determination of urine osmolality (Wescor 5500 vapor pressure osmometer, Wescor Inc, Logan, Utah, USA) to ensure euhydration. Adequate hydration was represented by an osmolality < 700 mOsmol (Popowski et al., 2001). Tympanic temperature was measured using a tympanic thermometer (FirstTemp Genius® Infrared thermometer, model 3000A, Sherwood Medical, St Louis, Missouri, USA). Subjects then had their bodyweight recorded to the nearest 0.05 kg (Wedderburn Precision Digital Floor Scale, Wedderburn, Brisbane, Australia) while wearing only Lycra cycling pants. Subjects entered the environment chamber and rested for 5 min to equilibrate to the hot environment. A 21-gauge indwelling venous cannula was inserted into a forearm vein for blood extractions throughout the trial. A 10-cm extension and three-way stop cock were attached to the cannula for blood sampling. Blood was taken with the subject in a seated posture to be consistent with

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samples taken during exercise. Blood collection procedures are described in detail elsewhere (van Rosendal et al., 2012).

Dehydration Subjects then mounted the cycle ergometer to begin the dehydration protocol which was divided into 30 min blocks. They cycled at a moderate intensity for the first 23 min. Then they rested for 7 min during which time they had blood taken, dismounted, toweled dry, urinated if needed, and were weighed to monitor fluid loss. This procedure was repeated until subjects had lost 4% of baseline bodyweight. Tympanic temperature was monitored continuously throughout the dehydration protocol and the session was terminated if it reached 40 °C. Urine collected throughout the dehydration session was included as part of the weight loss. A 3 mL aliquot of all urine samples was collected and stored at −80 °C. Once dehydrated to −4% bodyweight, subjects were asked to report their thirst sensation using a 9-point thirst scale ranging from 1 (not thirsty) to 9 (very thirsty) (Maresh et al., 2001). Whole-body sweat rates throughout the trials were calculated as bodyweight changes, corrected for fluid intake and urine output, divided by total exercise duration (Deschamps et al., 1989).

Rehydration Oral and oral glycerol trials. Throughout rehydration, subjects remained in a 22 °C environment. Figure 2 shows a schematic representation of the 2 h rehydration period. In the first hour of the oral and oral glycerol trials, subjects consumed 100% of the fluid they lost during dehydration in six separate oral fluid volumes, consumed every 10 min. The first bolus was the first experimental solution comprising a volume of carbohydrate-electrolyte beverage (Gatorade®) equal to 40% of the total first hour rehydration volume, with (oral glycerol) or without (oral) a glycerol dose of 1 g/kg bodyweight. The remaining 60% of the first hour fluid volume was broken into five boluses of 12% each. The second, fourth, and sixth fluid boluses were distilled water while the third and fifth fluid boluses were a carbohydrate-electrolyte beverage (Gatorade®). In the second hour of rehydration, a volume of fluid equal to 50% of the bodyweight lost during dehydration was consumed so that the total fluid replacement equaled 150% of bodyweight lost. The second hour fluid regime mirrored that used in the first hour, except half the volume of fluid was given during each 10-min period. For the oral glycerol trials, the glycerol dose for the second experimental solution was also halved to 0.5 g/kg so that the total glycerol dose was 1.5 g/kg bodyweight. IV and IV with oral glycerol trials. For the IV and IV with oral glycerol trials, half of the total rehydration fluid volume was IV

Rehydration with IV fluid and oral glycerol

Fig. 2. Timeline delineating the differences between the rehydration protocols used for the four trials. aFluid bolus consumed every 10 min as a % of the total oral fluid volume consumed in that hour. bType of fluid consumed in each bolus. CHO-elect = carbohydrateelectrolyte beverage. cGlycerol dose for oral with glycerol and IV with oral glycerol trials. fluid (0.9% NaCl) and half was oral fluid. Again, the total volume of fluid provided in the first hour was 100% of the fluid lost during dehydration, with the remaining 50% provided in the second hour. Therefore, the first hour IV and oral volumes were each 50% of the total bodyweight loss during dehydration, while the second hour IV and oral volumes were each 25% of the bodyweight loss. For the IV fluid, the saline bags were attached to the cannula already in place and flow was adjusted to run at a rate of approximately 0.80 mL/kg/min so that the correct volume of fluid was administered by the 50 min mark of the first hour. The IV line was then interrupted and the subject rested for the remaining 10 min to allow the saline to disperse from within the vein before the first hour rehydration blood sample was withdrawn. The second hour IV fluid was then attached to the cannula and the line reopened. The flow was adjusted to approximately 0.40 mL/kg/min so that the fluid was again administered over the first 50 min of the hour. While the IV fluid was being administered the subject remained seated and consumed the oral component of the rehydration volume. The oral protocol for the IV and IV with oral glycerol trials was the same as that described for the oral and oral glycerol trials, except that the experimental solutions were 80% of the total oral fluid to be consumed that hour and the remaining five drinks were each 4% of the total volume of oral fluid to be consumed that hour. These percentage changes ensured that the experimental solutions were the same volume as in the oral trials. For the IV with oral glycerol trials, the experimental solutions once again comprised glycerol doses of 1 g/kg (first hour) and 0.5 g/kg bodyweight (second hour) in a carbohydrate-electrolyte beverage (Gatorade®).

All experimental solutions were mixed with sports drinks to ensure they had a similar temperature, color, texture, and flavor to mask the taste of glycerol. After consuming each experimental solution, the subjects were asked if they could distinguish whether they were receiving glycerol or not. Only one subject was able to correctly identify when they were on the glycerol trial. Subjects were also questioned regarding adverse reactions and no subjects reported experiencing side effects during any trial. All oral and IV fluids were given at 22 °C to ensure a similar effect on core temperature between conditions. Other measurements taken during rehydration are displayed in Fig. 1. Post-rehydration equilibration period. Subjects remained in a temperate environment (22 °C) for the 60-min post-rehydration equilibration period. During this time they consumed a standardized lunch comprising two pieces of multigrain toast, two sachets of butter, two sachets of strawberry jam and a yoghurt top muesli bar, and a calculated amount of solid carbohydrate (jellybeans) to correct for the greater carbohydrate consumption in the oral and oral glycerol trials from the larger volume of carbohydrateelectrolyte sports drinks consumed. The total caloric intake of 70 kJ/kg aimed to offset the energy used during the dehydration component of the trial. Subjects then entered the environment chamber (34 °C, 60% relative humidity) and were prepared to begin the exercise performance test. Exercise performance test. The performance aspects of the study have been presented in another manuscript (van Rosendal et al., 2012) and are therefore not described in detail here. The

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van Rosendal et al. performance test itself was conducted as described in the familiarization trial. Measurements were collected as in Fig. 1. Blood and urine analyses. All variables were analyzed in duplicate and third measures were taken if the variation between samples was greater than 3%. The average of the values was used for analysis. Urine and plasma were analyzed for osmolality using a vapor pressure osmometer (Wescor 5500 vapor pressure osmometer, Wescor Inc) with Coefficient of variations (CVs) 0.6% and 0.7%, respectively. Hemoglobin and hematocrit were measured from whole blood and the percentage change in plasma volume was calculated using the equations of Dill and Costill (1974). Plasma aldosterone and cortisol were measured simultaneously using a High-performance liquid chromatography (HPLC)tandem mass spectrometric method that we developed (Taylor et al., 2010). Serum was measured for ADH using commercially available Fluorescent EIA kits (Phoenix Pharmaceuticals AVP [Arg8] kit, Burlingame, California, USA). In our hands, this assay yielded a CV of 11.8%. Total serum protein and serum concentrations of glycerol, Na+, K+, and Cl− were all measured via automated analysis (Cobas Mira, Roche Diagnostic Systems, Rotkreuz, Switzerland). CVs were: total protein 1.2%, glucose 1.9%, glycerol 2.6%, Na+ 0.4%, K+ 0.4%, and Cl− 0.6%.

Data analysis Data were initially tested for normality (Shapiro–Wilk normality test). General linear model with two-way repeated measures analysis of variance (ANOVA) with Tukey’s post-hoc test was used to assess time by trial interactions (for plasma volume and osmolality, total serum protein, Na+, K+ and Cl−, glycerol, aldosterone, cortisol, ADH, and thirst sensation). Mauchly’s tests were used to examine sphericity for the two-way repeated measures ANOVA and datum that was not significant based on the Mauchly’s test was corrected based on the epsilon value. If epsilon was less than 0.75 then Greenhouse–Geisser corrections were used, while Huynh–Feldt corrections were used if epsilon was greater than 0.75. Significance was then determined from these corrected significance values. One-way repeated measures ANOVA with Tukey’s post-hoc tests was used to compare group means for environment conditions; pretrial intake of total energy, protein, fat (total, saturated, polyunsaturated, and monounsaturated), cholesterol, carbohydrate, water, sodium, and potassium; bodyweight change and net fluid balance at each time point; dehydration time; whole-body sweat rates; experimental solution volumes; carbohydrate intake (solid and total); total Na+ intake; cumulative urine volumes and urine osmolality; and tympanic temperature. Pearson’s correlations measured the relationships between variables (between aldosterone and plasma volume, tympanic temperature, osmolality, Na+, and K+; between cortisol and plasma volume and osmolality; and between ADH and plasma volume and osmolality). One-way repeated measures ANOVA was analyzed using GraphPad Prism 5 (GraphPad Software Inc., La Jolla, California, USA) and two-way repeated measures ANOVA was analyzed using SPSS for Windows, version 17.0 (SPSS Inc., Chicago, Illinois, USA). The significance level was set at P = 0.05. All values presented are mean ± SD unless otherwise specified.

Results Fifteen individuals were recruited into the study; however, six withdrew. Of these, three withdrew due to an inability to tolerate exercise in the hot/humid environment, two withdrew because they moved interstate before completing the experimental trials, and one was

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unable to be cannulated for blood collection. Data are therefore presented for nine subjects. Pretrial diet There were no between-trial differences in pretrial nutritional dietary intakes (van Rosendal et al., 2012). Dehydration Climate chamber conditions during dehydration were similar (P > 0.05) between conditions [temperature (°C) and relative humidity (%); oral 34.9 ± 0.6 and 68.6 ± 8.3; oral glycerol 34.9 ± 0.6 and 68.9 ± 7.4; IV 35.2 ± 0.6 and 66.1 ± 7.4; IV with oral glycerol 35.3 ± 0.7 and 72.2 ± 6.7]. There was no difference in baseline hemoglobin between trials (Table 1). Baseline bodyweight was also similar between conditions, and decreased significantly following dehydration (P < 0.001), but to the same extent in all four trials (Table 1). Based on the technique used to dehydrate, the reduction in bodyweight was attributable to water loss, with no difference seen between conditions (P > 0.05). There were no between-trial differences for the average time taken for dehydration or for average sweat rate (P > 0.05) (Table 1). Rehydration Bodyweight increased significantly (P < 0.001) and to a similar extent in all trials following rehydration (Table 1). There were no differences for total rehydration fluid volumes between trials or for the volumes of experimental solutions 1 and 2 (Table 1). The total carbohydrate consumed from carbohydrate-electrolyte beverage plus solid carbohydrate was similar between trials (Table 2). Plasma volume, osmolality, serum protein, and glycerol Figure 3(a) shows the percentage change for plasma volume during the course of the study. Plasma volume decreased during dehydration in all groups (P < 0.001), but to a similar extent (P > 0.05) between groups. During rehydration, plasma volume increased in all groups (P < 0.001). Overall, both IV trials showed significantly greater increases for plasma volume than the two oral trials. The addition of glycerol to the rehydration solutions also increased plasma volume significantly higher than the corresponding fluid regime without glycerol. Therefore, plasma volume was highest in IV with oral glycerol, followed by IV, then oral glycerol and oral (P < 0.01 for all of these comparisons). Plasma osmolality was similar between trials at baseline and increased (P < 0.01) to a similar extent during dehydration in all conditions (Fig. 3(b)). Large variations were seen between conditions during rehydration

Rehydration with IV fluid and oral glycerol Table 1. Changes to hydration-related variables throughout dehydration, rehydration, and the performance test

Baseline Bodyweight (kg) Hemoglobin (g/dL) Dehydration Decrease in bodyweight (%) Time taken for dehydration (min) Sweat rate (L/h) Rehydration Increase in bodyweight (%) Total rehydration fluid volume (L) Volume of experimental solution 1 (L) Volume of experimental solution 2 (L) Total volume of sports drink (L) Solid CHO consumed (g) Total CHO (from sports drink + solid) (g) Total Na+ intake from rehydration fluids (g) Performance Decrease in bodyweight (%) Net fluid balance (change from baseline) Post-dehydration (kg) Post first hour rehydration (kg) Post second hour rehydration (kg) Post-equilibration (kg) Post-performance (kg) Urine osmolality Baseline (mOsmol) Cumulative urine volume (mL) Post first hour rehydration Post second hour rehydration Post-equilibration Post-performance

Oral

Oral glycerol

IV

IV with oral glycerol

73.67 ± 4.65 15.1 ± 1.0

74.02 ± 5.09 15.4 ± 1.3

74.28 ± 5.11 15.0 ± 1.1

74.41 ± 4.91 15.1 ± 0.9

4.01 ± 0.86 106.1 ± 14.4 1.5 ± 0.4

4.05 ± 0.65 112.8 ± 11.5 1.5 ± 0.4

4.02 ± 0.79 107.8 ± 16.6 1.5 ± 0.6

3.88 ± 0.82 106.1 ± 12.2 1.5 ± 0.4

5.24 ± 1.49 4.4 ± 0.9 1.2 ± 0.3 0.6 ± 0.1 2.8 ± 0.6* 10.4 ± 12.6 189.2 ± 30.1 1.33 ± 0.28

5.68 ± 1.27 4.5 ± 0.7 1.2 ± 0.2 0.6 ± 0.1 2.9 ± 0.5* 7.9 ± 13.3 189.2 ± 31.7 1.35 ± 0.22

5.24 ± 1.11 4.5 ± 0.9 1.2 ± 0.2 0.6 ± 0.1 2.0 ± 0.4 66.2 ± 19.0† 190.4 ± 30.8 8.86 ± 1.80†

5.79 ± 1.93 4.3 ± 0.9 1.2 ± 0.2 0.6 ± 0.1 1.9 ± 0.4 68.7 ± 16.7† 188.8 ± 34.0 8.57 ± 1.80†

−3.94 ± 0.62

−3.51 ± 0.95

−3.76 ± 0.80

−3.58 ± 0.63

−2.96 ± 0.62 −0.27 ± 0.17 0.74 ± 0.43 0.07 ± 0.46 −2.84 ± 0.55

−2.99 ± 0.47 −0.19 ± 0.14 1.03 ± 0.41 0.44 ± 0.35 −2.16 ± 0.62**

−2.99 ± 0.61 −0.32 ± 0.42 0.73 ± 0.50 0.66 ± 0.38§ −2.16 ± 0.65**

−2.89 ± 0.61 0.01 ± 0.32 1.23 ± 0.79‡ 1.09 ± 0.53§,¶ −1.61 ± 0.49**

400.8 ± 290.8

332.6 ± 131.0

317.3 ± 182.5

367.7 ± 143.7

37 ± 74 442 ± 239 1183 ± 364 1487 ± 482

23 ± 36 426 ± 236 992 ± 290 1233 ± 405

0±0 373 ± 369 711 ± 444†† 886 ± 466††

15 ± 45 256 ± 294 580 ± 243‡‡,§§ 755 ± 300‡‡,§§

*Oral and oral glycerol > IV and IV with oral glycerol (P < 0.001). † IV and IV with oral glycerol > oral and oral glycerol (P < 0.001). ‡ IV with oral glycerol > oral and IV (P < 0.05). § IV and IV with oral glycerol > oral (P < 0.01). ¶ IV with oral glycerol > oral glycerol and IV (P < 0.01). **Oral glycerol, IV and IV with oral glycerol > oral (P < 0.01). †† IV < oral (P < 0.01). ‡‡ IV with oral glycerol < oral (P < 0.001). §§ IV with oral glycerol < oral glycerol (P < 0.05)

and performance. From the first hour of rehydration until the end of the 40 km time trial, IV with oral glycerol was higher than oral (P < 0.01). With the exception of the post-equilibration time point, oral glycerol was also higher than oral from the first hour of rehydration until the end of 15-min steady-state exercise (P < 0.05). Furthermore, IV with oral glycerol was higher than oral glycerol from the second hour of rehydration until the end of the 40-km time trial (P < 0.05), except at the mid-equilibration time point. Total serum protein concentration was similar between conditions at baseline and increased (P < 0.01) in all conditions during dehydration (Fig. 3(c)). Total protein then decreased (P < 0.01) during rehydration, before increasing (P < 0.01) again during the performance test in all trials. The decrease during rehydration was greater in the IV and IV with oral glycerol trials compared with oral and oral glycerol (P < 0.01). IV and IV with oral glycerol remained lower (P < 0.05) than

oral and oral glycerol until the end of the 30-min steadystate component of the performance test. Na+ increased (P < 0.01) by a similar magnitude in all trials during dehydration and decreased significantly (P < 0.05) during the first two hours of rehydration (Fig. 4(a)). Na+ was then significantly lower in the oral glycerol trial compared with IV (P < 0.05) throughout equilibration and performance. In addition, Na+ was lower in oral glycerol compared with IV with oral glycerol from post-equilibration until 20 km into the time trial (P < 0.05) and in oral glycerol compared with oral during the first 20 km of the time trial (P < 0.05). There was no significant change to K+ during dehydration in any condition (Fig. 4(b)). K+ decreased (P < 0.05) in all trials during rehydration before increasing in all trials during the performance test (P < 0.05), but with no significant between-trial differences at any time. During dehydration, Cl− increased (P < 0.01) by a similar amount in all conditions (Fig. 4(c)). During rehydration

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van Rosendal et al. Table 2. Pearson’s correlations between fluid regulatory and stress hormones and factors known to influence their release

Aldosterone Oral Oral glycerol IV IV with oral glycerol Cortisol Oral Oral glycerol IV IV with oral glycerol Antidiuretic hormone Oral Oral glycerol IV IV with oral glycerol

Plasma volume

Tympanic temperature

Osmolality

Na+

K+

−0.94‡ −0.94‡ −0.89‡ −0.83‡

0.74† 0.76† 0.77† 0.77†

0.91‡ 0.50 0.72† 0.10

0.90‡ 0.75† 0.82‡ 0.96‡

0.81‡ 0.87‡ 0.75† 0.81‡

−0.89‡ −0.83‡ −0.88‡ −0.89‡

0.61* 0.60* 0.76† 0.83‡

– – – –

– – – –

– – – –

−0.83§ −0.72 −0.70 −0.76

– – – –

0.58 0.90* 0.92* 0.27

– – – –

– – – –

Data presented as correlation coefficients (r). n = 9. *(P < 0.05). † (P < 0.01). ‡ (P < 0.001). § (P = 0.09). IV, intravenous.

and equilibration, Cl− decreased (P < 0.001) in oral and oral glycerol but not in either IV trial so both IV trials had significantly higher Cl− than both oral trials at most time points. During the performance test, Cl− remained higher in both IV trials compared with oral glycerol (P < 0.05) but only IV was higher than oral (P < 0.01). Blood glycerol concentrations are presented in Fig. 5. Glycerol increased significantly (P < 0.001) after the first hour of rehydration in both the oral glycerol and IV with oral glycerol trials. For the oral glycerol condition, peak glycerol concentration (16.4 ± 1.7 mmol/L) occurred at the end of the second hour of rehydration. For IV with oral glycerol, the peak concentration was similar (16.3 ± 1.9 mmol/L) but occurred 30 min later at the mid-equilibration time point. There were no differences in blood glycerol concentrations between oral glycerol and IV with oral glycerol at any stage.

Hormonal responses There were no significant differences (P > 0.05) in ADH between conditions at any time point (Fig. 6(a)). In all conditions, both cortisol (Fig. 6(b)) and aldosterone (Fig. 6(c)) increased significantly during dehydration, decreased significantly during rehydration, and increased significantly during the exercise performance test (P < 0.001). Both hormones were lower in the IV and IV with oral glycerol trials compared with oral and oral glycerol at several points during rehydration and performance (P < 0.05). Correlations for these three hormones and factors known to affect their release are shown in Table 2.

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Aldosterone showed strong negative correlations with percentage change in plasma volume and positive correlations with tympanic temperature, Na+ and K+, in all four groups. Aldosterone was also well correlated with osmolality in the oral and IV trials and moderately correlated with osmolality in the oral glycerol condition. In all trials, cortisol was highly negatively correlated with plasma volume and moderate-to-highly positively correlated with tympanic temperature. ADH was highly negatively correlated with plasma volume in all trials while ADH and osmolality were well correlated in the oral glycerol and IV conditions, moderately correlated in the oral trial. Urine osmolality and volume Urine osmolality was similar between trials at baseline (Table 1). The total cumulative urine volumes throughout rehydration and the performance test are presented in Table 1. There were no significant differences (P > 0.05) between oral and oral glycerol, or between IV and IV with oral glycerol, at any time point. However, by the end of the equilibration period, significantly (P < 0.01) lower urine volumes were seen in the IV trials compared with the oral trials and these were maintained following the performance test (P < 0.01). Thirst sensation Thirst sensation at the end of dehydration (oral 6 ± 2; oral glycerol 7 ± 1; IV 7 ± 1; IV with oral glycerol 6 ± 1) and the first hour of rehydration (oral 2 ± 1; oral glycerol 2 ± 1; IV 3 ± 1; IV with oral glycerol 3 ± 2) was

Rehydration with IV fluid and oral glycerol

Fig. 3. Percent change in plasma volume (a), plasma osmolality (b), and total serum protein (c) during exercise-induced dehydration (−4% bodyweight), rehydration with 150% of fluid lost, passive equilibration, and an exercise performance test. See Methods section for details of the experimental procedures. Data are presented as mean ± SEM (n = 9). (P < 0.05)*oral vs IV, †oral vs IV with oral glycerol, ‡oral glycerol vs IV with oral glycerol, §oral vs oral glycerol, #IV vs IV with oral glycerol, $oral glycerol vs IV. BW = bodyweight, Rehyd = rehydration.

similar in all conditions. Thirst was higher (P < 0.05) in IV with oral glycerol compared with the oral glycerol trial following the second hour of rehydration (oral 2 ± 1; oral glycerol 1 ± 1; IV 2 ± 1; IV with oral glycerol 3 ± 1) and after equilibration (oral 2 ± 1; oral glycerol

Fig. 4. Change in serum electrolytes Na+ (a), K+ (b), and Cl– (c) during exercise-induced dehydration (−4% bodyweight), rehydration with 150% of fluid lost, passive equilibration, and an exercise performance test. See Methods section for details of the experimental procedures. Data are presented as mean ± SEM (n = 9). (P < 0.05)*oral glycerol vs IV, †oral glycerol vs IV with oral glycerol, ‡oral vs oral glycerol, §oral vs IV, #oral vs IV with oral glycerol. BW = bodyweight, Rehyd = rehydration.

2 ± 1; IV 2 ± 1; IV with oral glycerol 3 ± 1). Thirst was then similar between all conditions during the performance test (van Rosendal et al., 2012). Discussion The present study explored the simultaneous effects of rehydrating with IV fluid and oral glycerol. The follow-

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Fig. 5. Serum glycerol concentrations during rehydration, equilibration, and performance in the oral with glycerol and IV with oral glycerol trials. Glycerol levels were undetectable during the dehydration period and for the full duration of the oral and IV trials. Data are presented as mean ± SEM (n = 9).

ing findings emerged: (a) plasma volume was restored more quickly and completely with IV therapy compared with oral therapy; (b) the IV with oral glycerol trial was associated with the greatest fluid retention and lowest urine volume of the four protocols; and (c) compared with oral rehydration, IV fluids resulted in a lower plasma cortisol concentration during rehydration and a lower aldosterone concentration during rehydration and exercise performance. Exercise in the heat induces a state of hypovolemichyperosmolality that is intricately related to many of the physiological responses that allow the body to cope with heat stress. In the current study, several of the variables underpinning the adaptive mechanisms of the body (such as plasma volume, osmolality, and electrolyte concentrations) were affected to differing degrees between the four rehydration conditions. The biggest difference between trials was seen for plasma volume. Previous studies have shown that plasma volume is consistently restored more rapidly and completely when rehydrating with IV compared with oral fluid, whether following sustained (i.e., overnight) (Casa et al., 2000) or acute (Kenefick et al., 2000, 2006) exercise-induced dehydration. Similarly, glycerol ingestion has been associated with enhanced plasma volume when used as a hyperhydrating agent (Coutts et al., 2002; Magal et al., 2003), or as a substrate to enhance rehydration during (Murray et al., 1991; Siegler et al., 2008) or after (Scheett et al., 2001; Magal et al., 2003; Kavouras et al., 2006) exercise. Our hypothesis that combining IV fluid with oral glycerol would augment plasma volume to the greatest extent during rehydration was confirmed.

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Fig. 6. Changes to ADH (a), cortisol (b), and aldosterone (c) concentrations during exercise-induced dehydration (−4% bodyweight), rehydration with 150% of fluid lost, passive equilibration, and an exercise performance test. See Methods section for details of the experimental procedures. Data are presented as mean ± SEM (n = 9). (P < 0.05)*oral glycerol > IV with oral glycerol, †oral > IV with oral glycerol, ‡oral > IV, §oral glycerol > IV. BW = bodyweight, Rehyd = rehydration.

Importantly for athletes exercising in hot/humid environments, the current findings are the first to show a prolonged benefit from IV fluids, independent of the addition of glycerol. The enhanced plasma volumes seen in both IV trials were maintained throughout the course of the exercise performance test, some 210 min after

Rehydration with IV fluid and oral glycerol rehydration began. Previous studies have indicated that the rapid increase of fluid in the vasculature is transient and equilibrates within 35 min during passive rehydration (Kenefick et al., 2000) and even more rapidly with the onset of exercise immediately after rehydration (Casa et al., 2000; Kenefick et al., 2000, 2006; Maresh et al., 2001). While this redistribution of fluid to the extravascular spaces is good from a whole-body rehydration perspective, the rapid influx of the infused fluids also stimulates a greater urine production (Kenefick et al., 2006) and therefore a reduced overall benefit to net fluid balance. In a recent review (van Rosendal et al., 2010b), we concluded that IV rehydration may therefore offer a short-term physiological advantage when little time is available for rehydration; however, if periods of longer than approximately 1 h are available for fluid absorption and equilibration, then few differences between IV and oral rehydration are observed. Discrepancies between the current study and the previous findings showing no sustained benefit are most likely explained by differences in study design. We used a total fluid volume as recommended by the American College of Sports Medicine for rapid and complete recovery following exercise (150% of the bodyweight lost) (Sawka et al., 2007). In previous IV rehydration studies, subjects were also dehydrated by ∼ 4% bodyweight (∼ 2.8 L) but only rehydrated with 50% of the fluid lost (0.45% NaCl in Casa et al., 2000 and 0.45% and 0.9% NaCl in Kenefick et al., 2000). In Kenefick et al.’s (2006) study, the subjects were dehydrated to −2.4% bodyweight (∼ 1.7 L) and rehydrated with 100% of fluid lost (0.45% NaCl). Normal saline is a replacement crystalloid solution that has a [Na+] (154 mmol/L) similar to that of the extracellular fluid. Because Na+ is the principle determinant of extracellular osmolality, the distribution of normal saline is effectively limited to the extracellular space. The fluid then distributes between the extracellular constituents in proportion to their volumes (∼3/4 in interstitial fluid, ∼1/4 in plasma). IV rehydration with half normal saline still replenishes the extracellular fluid first; however, the lower [Na+] means the fluid is not restricted to the extracellular compartment and a gradient is initiated favoring the movement into the intracellular fluid. These fluid shifts would be comparatively more rapid following IV compared with oral rehydration because an immediate and large hydrostatic pressure gradient is induced by administering the fluid directly into the vasculature, without the time constraints of waiting for gastrointestinal absorption. Thus, the use of 0.45% NaCl coupled with the smaller total fluid volumes in the previous trials (∼1.3–1.9 L) resulted in a much smaller volume of fluid acting to expand the extracellular compartments (including plasma volume) and the equilibration between compartments could be expected to be fairly rapid. Although only half of the rehydration volume was IV fluid in the

present study, the absolute volumes of IV saline infused were somewhat comparable to those used in previous studies, even though the total volume of fluid was much higher. Additionally, the use of 0.9% NaCl provided twice the [Na+] as studies using 0.45% NaCl, and therefore ensured a much greater proportion of the total rehydration fluid volume was acting to expand the extracellular compartments for a longer period. Because of the relatively greater [Na+] in saline compared with sports drinks, the total amount of sodium provided was much higher in the IV trials compared with the oral trials. One would expect this to have contributed to the greater plasma volume expansion in the IV trials; however, with the exception of oral glycerol vs IV, there was no sustained pattern of differences for serum [Na+] between the conditions. The criticism of IV rehydration providing only a transient hydration benefit is supported by increases in urine volume following IV infusion compared with the ingestion of oral fluids when 100% of the fluid was replaced (Kenefick et al., 2006), although similar urine volumes were seen when only half of the fluid was replaced (Kenefick et al., 2007). The present results also differ from these previous observations. Urine volume was consistently significantly reduced by approximately 40% in the IV trials compared with the oral trials at the post-equilibration and postperformance time points. These data are supported by the persistent expansion of plasma volume and lower serum protein levels in the IV trials compared with the oral trials from first hour of rehydration until postperformance, and the lower aldosterone in the IV trials during rehydration and performance. The reason for this disparity is again most likely related to the relative amounts of Na+ provided in the trials. The previous trials used 0.45% NaCl as both the IV and oral rehydration solutions meaning that Na+ replacement was equal between trials. Oral rehydration in the current trials provided a much smaller total [Na+] and therefore the fluids were distributed throughout the entire total body water. These results are consistent with others who have shown that urine production following rehydration was inversely related to the Na+ concentration of the ingested fluids (Shirreffs & Maughan, 1998). The addition of glycerol was expected to provide a further osmotic drive favoring the retention of the rehydration fluids. Because glycerol is slowly absorbed and metabolized, plasma levels were still very high at the end of the performance test. Thus, the glycerol trials were expected to extend the duration required for fluid shifts from extracellular to intracellular compartments by increasing the extracellular osmolality, therefore favoring fluid retention in that space. However, plasma volumes between the oral and oral glycerol trials were not significantly different throughout the study duration (with the exception of mid-second hour rehydration time point). There was also no significant benefit to

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van Rosendal et al. plasma volume in the IV with oral glycerol relative to the IV groups. These results are in contrast to all three previous studies using glycerol in rehydration that showed an enhanced plasma volume in the glycerol trials (Scheett et al., 2001; Magal et al., 2003; Kavouras et al., 2006). Over time, glycerol is dispersed throughout the total body water compartments leading to anticipated benefits to net fluid balance. Therefore, another unexpected finding was that net fluid balance and urine volumes were not consistently reduced with the addition of glycerol to the two modes of rehydration. Overall, the greatest differences for net fluid balance and urine volume occurred between the oral and IV groups. Therefore, the mode of rehydration was more influential in reducing urine volume than the inclusion of the osmotic substrate into the rehydration solution. This is in stark contrast to the overwhelming body of hyperhydration literature showing that glycerol decreases urine volume (van Rosendal et al., 2010a). However, it is similar to the three previous studies investigating glycerol use in rehydration (Scheett et al., 2001; Magal et al., 2003; Kavouras et al., 2006), which all showed no significant differences for urine volume when monitored for 80–180 min following oral rehydration with or without glycerol. Based on Scheett et al.’s (2001) study, we recently proposed that the influence of glycerol on total fluid retention might be related to rehydration duration (van Rosendal et al., 2010a). Urine volume was similar between the trials with and without glycerol for the first 2 h of rehydration, but increased dramatically during the third hour of the water only trial (Scheett et al., 2001). Should such a pattern have continued, fluid excretion in the water trials would have rapidly increased compared with the glycerol trial. However, we monitored urine volumes for a longer duration in the current experiment and the results did not support that assertion. The decreases in urine volume in the two glycerol trials compared with the corresponding trials without glycerol were established after the postequilibration period (3 h after the onset of rehydration) and almost identical percentage differences were seen after the performance test, some 90 min later. Another interesting finding was that while plasma Cl− was maintained following IV rehydration, likely due to the Cl− in the saline, plasma Na+ dropped following rehydration in all conditions despite the saline providing equivalent Na+. The reason for this disparity is not apparent. It may be that atrial natriuretic peptide (Mannix et al., 1990), or more likely urodilatin (Drummer et al., 1996), was released in response to the increased plasma volume in the IV trials. These hormones promote Na+ loss at the kidney. However, as neither was measured in the current study, this is only speculative. Despite the vast improvements in fluid retention in the IV with oral glycerol condition, thirst sensation was significantly higher in IV with oral glycerol compared

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with oral after the second hour of rehydration and compared with oral glycerol post-equilibration. However, IV was not higher than either of the oral trials at any time point. Thirst has been shown to be influenced by plasma osmolality and Na+ concentration (Greenleaf, 1992). These mechanisms may have contributed to the present findings since plasma osmolality was much higher in the IV with oral glycerol trial compared with oral during rehydration and plasma Na+ was much higher in IV with oral glycerol compared with oral glycerol after the equilibration period. The effects of IV rehydration on thirst are an important consideration for athletes. It has recently been proposed that thirst may act in a feed-forward manner to involuntarily slow the athlete, as an attempt to prevent dehydration from becoming physiologically significant (Sawka & Noakes, 2007). Furthermore, thirst affects the subjective response to exercise and also influences factors such as the release of fluid regulating hormones via the act of drinking (e.g., via oropharyngeal stimulation). Wellrehydrated, though still thirsty, athletes must also be cautious not to overdrink plain water on top of the IV fluid they have received, otherwise they may increase the risk of hyponatremia (Noakes, 2002). The hypovolemic-hyperosmolar state caused by exercise also stimulates the renin-angiotensin-aldosterone system. This primarily occurs as a consequence of the reduction in arterial blood pressure, subsequent to the drop in extracellular fluid volume. Aldosterone increases during periods of dehydration and decreases with rehydration because its secretion is primarily influenced by plasma volume (Brandenberger et al., 1986; Nose et al., 1988a), plasma Na+ and K+ (Brandenberger et al., 1986, 1989), and plasma adrenocorticotropic hormone (ACTH) (Nose et al., 1988a). Cortisol secretion is controlled almost entirely by ACTH (Collins & Weiner, 1968). In turn, ACTH secretion is effected by plasma volume and thermosensitive control centers in the hypothalamus (Francis, 1979; Brandenberger et al., 1986). As a result, cortisol has been shown to be a sensitive measure of heat stress and increases are accompanied by subjective discomfort (Follenius et al., 1982). Cortisol fluctuations during exercise also therefore mirror the changes in aldosterone and reflect the increased physiological strain resulting from hypovolemia (Francis, 1979; Francesconi et al., 1985) and from increased heart rate and core temperature, and reduced sweat rate (Follenius et al., 1982). In the present study, aldosterone was well correlated with variables associated with hydration balance including change in plasma volume, Na+, K+, and osmolality, while cortisol was highly correlated with variables of exercise heat stress including change in plasma volume and tympanic temperature. As mentioned above, the lower aldosterone concentrations during rehydration and performance in the two IV conditions reflect the improved maintenance of plasma volume in those trials. The simultaneous

Rehydration with IV fluid and oral glycerol reduction to cortisol seen in the IV trials during rehydration indicates a decreased physiological stress. This is the first study to show such changes when rehydrating with oral and IV fluids following exercise-induced dehydration. It is also known that reduced plasma volume (Robertson & Athar, 1976) and increased osmolality (Moses & Miller, 1971), which typically occur simultaneously during dehydration, are potent stimulators for the secretion of ADH. The lack of significant differences for ADH in the present study likely reflects that plasma volume and osmolality were both highest in the IV with oral glycerol trial and lowest in the oral trial. It is therefore probable that the influence of the increased plasma volume, which should rapidly reduce ADH release, was effectively balanced by the increased osmolality, which stimulates ADH release, and vice versa. This postulate is supported by the correlations between ADH and plasma volume, which were high for all four conditions, and between ADH and osmolality, which were moderate to high for all trials, except in IV with oral glycerol. Previous studies have also concluded that ADH is reduced in an anticipatory manner in response to the ingestion of water and subsequent decrease in plasma osmolality (Geelen et al., 1984; Seckl et al., 1986). These studies indicate that oropharyngeal receptors may rapidly initiate inhibition of ADH before decreases to plasma osmolality or volume are even detected (Kenefick et al., 2000). The fact that all trials in the present study received at least half the volume of fluid orally may help explain why no differences for ADH were seen between conditions even though there were differences for osmolality and plasma volume. There were several limitations associated with the study that need to be acknowledged. During pilot testing, attempts were made to single blind the administration of IV fluid with respect to the subjects. Unfortunately, this was logistically difficult and ineffective because the subjects were able to feel the fluid being administered since it was cooler than body temperature. Thus, these plans were abandoned for the main study. Importantly, it is very unlikely that the athletes knowing they received IV fluid would have resulted in any changes to the physiological variables measured in this study. Future research should investigate whether glycerol is more effective at enhancing rehydration when combined with a sports drink, as in the present study, as opposed to water. Several authors have demonstrated that a combination of glycerol and glucose enhances the absorption of sodium and water in rat intestinal models (which is similar physiologically to the human intestine) (Wapnir et al., 1996; Allen et al., 1999). Sports drinks are used by athletes because they are generally considered to provide a better hydration potential than water alone, as well as providing sodium to assist in fluid retention and to attenuate hyponatremia. A randomized controlled trial

comparing glycerol ingestion with sports drinks and water is warranted. Further research examining the optimal rate and volume of IV fluid infusion would also be useful. In conclusion, we examined the combination of IV infusion and oral glycerol as a technique to enhance rehydration. Anecdotally, the use of IV fluids occurs frequently in sports that are not subject to the WADA’s jurisdiction (e.g., most professional leagues in the United States and the National Collegiate Athletic Association) despite there being little evidence to support its use. In order to accurately reflect real-life practice, athletes were rehydrated with a combination of IV and oral fluid, rather than giving IV fluid in isolation, as few athletes would typically rehydrate exclusively with IV fluid after moderate exercise-induced dehydration in the heat. By using a protocol that more accurately depicts the way athletes use IV fluid compared with previous research and by incorporating the American College of Sports Medicine’s guidelines for rehydration, this is the first study to show that rehydration with IV fluids between exercise bouts may be beneficial for athletes. Although the majority of cases of dehydration can be treated via oral fluid consumption, there exists a subgroup of athletes for whom the current findings may have a very important practical application. These include athletes who compete or train several times a day in hot humid environments, with short recovery times. The inclusion of glycerol with IV fluid further enhanced hydration. Thus, athletes who require large volumes of fluid for rehydration in short time frames will benefit from IV fluid and the inclusion of oral glycerol will further enhance rehydration.

Perspectives Rapid and complete rehydration is important for athletes who exercise in hot/humid conditions, especially when conducting multiple exercise sessions with little time for recovery. In these athletes, rehydration may be difficult with oral fluids alone. Anecdotally, many such athletes use IV fluids to assist in rehydration despite the technique carrying risks not applicable to oral rehydration (e.g., infection, hematoma, air embolus, extravasation), and despite previous research showing no sustained benefits from using IV vs oral fluids in athletes. By using a combination of IV and oral fluids and oral glycerol, this is the first study to show a benefit to athletes that may offset the associated risks. We achieved a more rapid and complete restoration of plasma volume and net fluid balance that was maintained throughout subsequent exercise and was associated with lower fluid regulatory and stress hormone levels. These changes were also associated with a 3.5– 4.1% performance improvement (van Rosendal et al.,

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van Rosendal et al. 2012). Thus, such a technique may be beneficial in situations where an athlete has limited recovery time. However, practitioners should be aware that both IV fluids and glycerol are banned by WADA and are therefore unable to be used in athletes competing under the WADA jurisdiction.

Acknowledgements

Key words: Aldosterone, athlete, cortisol, dehydration, exercise, heat.

Conflicts of interest: The authors of this study declare that they have no conflicts of interest.

The authors wish to thank the participants for volunteering to be a part of this study and the Queensland Academy of Sport for the use of their facilities. The expert technical assistance of Gary Wilson is also gratefully acknowledged. There was no funding received from National Institutes of Health, Wellcome Trust, Howard Hughes Medical Institute, or from any other sources for this work.

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