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. 2011 Nov 7;590(Pt 3):607–616. doi: 10.1113/jphysiol.2011.217919

Brain glycogen supercompensation following exhaustive exercise

Takashi Matsui 1, Taro Ishikawa 1, Hitoshi Ito 1, Masahiro Okamoto 1, Koshiro Inoue 1, Min-chul Lee 1, Takahiko Fujikawa 2, Yukio Ichitani 3, Kentaro Kawanaka 4, Hideaki Soya 1
PMCID: PMC3379704  PMID: 22063629

Abstract

Non-technical summary

Exercise training elicits an increase in the basal level of muscular glycogen. This happens when glycogen recovers to above its basal level (supercompensation) after it decreases with acute exercise. Although untested, it is hypothesized that, similar to that of skeletal muscle, brain glycogen supercompensation occurs after acute exhaustive exercise. We provide evidence that exhaustive exercise induces glycogen supercompensation not only in skeletal muscles, but also in the brain. Furthermore, we observed exercise training-induced increases in basal glycogen levels in the cortex and hippocampus, which are involved in motor control and cognitive function. This suggests that, like skeletal muscles, the brain adapts metabolically, probably to meet the increased energy demands of exercise training.

Abstract

Brain glycogen localized in astrocytes, a critical energy source for neurons, decreases during prolonged exhaustive exercise with hypoglycaemia. However, it is uncertain whether exhaustive exercise induces glycogen supercompensation in the brain as in skeletal muscle. To explore this question, we exercised adult male rats to exhaustion at moderate intensity (20 m min−1) by treadmill, and quantified glycogen levels in several brain loci and skeletal muscles using a high-power (10 kW) microwave irradiation method as a gold standard. Skeletal muscle glycogen was depleted by 82–90% with exhaustive exercise, and supercompensated by 43–46% at 24 h after exercise. Brain glycogen levels decreased by 50–64% with exhaustive exercise, and supercompensated by 29–63% (whole brain 46%, cortex 60%, hippocampus 33%, hypothalamus 29%, cerebellum 63% and brainstem 49%) at 6 h after exercise. The brain glycogen supercompensation rates after exercise positively correlated with their decrease rates during exercise. We also observed that cortical and hippocampal glycogen supercompensation were sustained until 24 h after exercise (long-lasting supercompensation), and their basal glycogen levels increased with 4 weeks of exercise training (60 min day−1 at 20 m min−1). These results support the hypothesis that, like the effect in skeletal muscles, glycogen supercompensation also occurs in the brain following exhaustive exercise, and the extent of supercompensation is dependent on that of glycogen decrease during exercise across brain regions. However, supercompensation in the brain preceded that of skeletal muscles. Further, the long-lasting supercompensation of the cortex and hippocampus is probably a prerequisite for their training adaptation (increased basal levels), probably to meet the increased energy demands of the brain in exercising animals.

Introduction

Brain glycogen, which localizes in astrocytes, is an important energy source for neurons (Brown, 2004)). Astrocytic glycogen is degraded into lactate to provide fuel for neurons during sleep deprivation, hypoglycaemia and memory formation (Kong et al. 2002; Herzog et al. 2008; Suzuki et al. 2011)), suggesting a role for glycogen in energy provision during sustained neuronal activity. Exercise also increases neuronal activity and creates an energy demand in the brain (Vissing et al. 1996; Saito & Soya, 2004; Ohiwa et al. 2007; Soya et al. 2007a,b)). Prolonged exhaustive exercise with hypoglycaemia elicits a decrease in brain glycogen due to an increase in brain monoamines (i.e. noradrenaline (NA) and 5-hydroxytryptamine (5-HT)), which are not only the inducing factors for central fatigue (Newsholme et al. 1992)), but also glycogenolysis-enhancing factors in astrocytes (Brown, 2004)). Hence, brain glycogen may be part of a new mechanism for central fatigue during acute prolonged exhaustive exercise (Matsui et al. 2011)).

In skeletal muscle, exhaustive exercise results in glycogen depletion, but glycogen returns to above basal levels (supercompensation) approximately 24 h after cessation of exercise (Bergstrom & Hultman, 1966)). An adaptation to meet the increased energy demand in skeletal muscle with exercise training is based on glycogen supercompensation following exercise (James & Kraegen, 1984)). Additionally, 3 days of a hyper-carbohydrate diet increases basal levels of muscle glycogen (also known as ‘muscle glycogen loading’), and prolongs the time required to exercise exhaustion (Bergstrom et al. 1967; Pitsiladis & Maughan, 1999)). These data indicate that the glycogen level of an exercising muscle is a critical factor in performing endurance exercise to exhaustion.

Interestingly, in the brain, neuronal activation time in the cortex during hypoglycaemia was prolonged by elevating the basal level of brain glycogen with an injection of a glycogen phosphorylase inhibitor, CP-316,819; this inhibitor causes glycogen accumulation under normoglycaemic conditions but permits glycogen utilization under hypoglycaemic conditions (Suh et al. 2007)). Therefore, if we can increase brain glycogen levels with exercise, as we can in skeletal muscles, endurance performance can be increased through a sustained central command from the motor cortex to skeletal muscle, thereby mitigating central fatigue during exercise.

It is, however, still unknown whether brain glycogen supercompensation occurs following exhaustive exercise. Secher and colleagues have measured the arterial–jugular venous glucose, lactate, oxygen concentration differences and their ratios in exercising humans (Nybo & Secher, 2004)), suggesting that quick replenishment of brain glycogen could occur after exhaustive exercise. Moreover, using a biopsy technique, the hippocampal glycogen levels of patients with epilepsy, a disease characterized by abnormally elevated neuronal activity in the temporal lobe and hippocampus, were found to be higher compared with other normal brain regions (Dalsgaard et al. 2007)), as observed in well-trained skeletal muscle. These results led to the hypothesis that glycogen supercompensation occurs in the brain, as well as in skeletal muscle, following exhaustive exercise. This hypothesis remains to be tested.

In the present study, to test the above-mentioned hypothesis, we determined brain glycogen levels after exhaustive exercise by employing an accurate high-power (10 kW) focused microwave irradiation (MI) as a gold standard method, which momentarily inactivates glycogenolytic and glycosynthetic enzymes (Kong et al. 2002; Matsui et al. 2011)).

Methods

Experimental design

The experiments are composed of two major steps. First, we investigated whether brain glycogen supercompensation occurs following exhaustive exercise. Next, we examined the effect of exercise training on glycogen levels in several brain loci. Our experiments comply with the policies and regulations of The Journal of Physiology (Drummond, 2009)).

Materials

All chemicals, including amyloglucosidase, hexokinase, NADP+-dependent glucose-6-phosphate dehydrogenase, NADP+, ATP, EDTA, MgSO4, glucose, glucose-6-phosphate, KOH, imidazole, perchloric acid, Tris-HCl, 5,5-dithiobis (2-nitrobenzoate) (DTNB), acetyl-CoA and oxaloacetate (OAA) were purchased from Sigma (St Louis, MO, USA) and Nacalai Tesque (Kyoto, Japan).

Animals

Adult male Wistar rats (250–300 g) (SLC Inc., Shizuoka, Japan) were housed and cared for in an animal facility, fed a standard pellet diet (MF, Oriental Yeast Co., Ltd, Tokyo, Japan), and given water ad libitum. The room temperature was maintained between 22 and 24°C under a 12 h light–12 h dark cycle (lights on: 07:00–19:00). All experimental protocols were conducted in accordance with University of Tsukuba Animal Experiment Committee Guidelines.

Habituation to treadmill running

After a 1 week acclimatization period, rats were habituated to run on a treadmill (SN-460, Shinano, Tokyo, Japan) for a total of five sessions over 6 days. The running duration was 30 min day−1, and the running speed was gradually increased from 5 to 25 m min−1 (Nishijima & Soya, 2006; Soya et al. 2007a; Nishijima et al. 2011)).

Surgery

Surgery was performed according to methods described by Soya et al. (2007a)). After habituation to treadmill running, the rats were anaesthetized with sodium pentobarbital (50 mg kg−1, i.p.), and a silicone catheter was inserted into the jugular vein and fixed with a silk thread (32 mm). The external distal end of the catheter was fixed at the nape of the animal. The following experiments and analyses were carried out 2 days after surgery.

Exhaustive exercise

Figure 1A shows the experimental procedure. Rats were fasted before exercise for 2 h to obtain stable glycogen levels in the brain. They were exercised on a treadmill at 20 m min−1 to exhaustion (n = 8–13 rats per group). A sedentary group of animals (n = 8 rats) was placed on a stationary treadmill for 90 min. Exhaustion was considered to have occurred when the rat was unable to keep pace with the treadmill, lay flat on, and stayed on the grid positioned at the back of the treadmill for a period of 30 s despite being gently pushed with sticks or breathed on (Hasegawa et al. 2008)). At exhaustion, rats were injected with 50% glucose (0.2 ml (300 g body wt)-1) through the catheter to recover to a normoglycaemic condition from hypoglycaemia (Suh et al. 2007)). Immediately after exercise, and at 3, 6 and 24 h post-exercise, rats were killed to prepare tissues.

Figure 1. Blood parameters and glycogen levels in skeletal muscles and liver after exhaustive exercise.

Figure 1

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A, experimental procedure; B, blood glucose and lactate levels; C, skeletal muscle glycogen levels; D, liver glycogen levels; E, blood insulin levels. Data represent the mean ± SEM (n = 8 – 13 rats). *P < 0.05; **P < 0.01 compared to pre-exercised rats (Dunnett's post hoc test).

Exercise training

Two days after the habituation period, rats were subjected to running training at 20 m min−1 for 60 min on a treadmill during the dark phase, 5 days week−1 for 3 weeks. At the same time, a sedentary group of animals was placed on a stationary treadmill as control. Seventy-two hours after the last session of exercise training, rats were killed to prepare tissues.

Tissue preparation

Following exhaustive exercise and exercise training, rats were anaesthetized with isoflurane (a mixture of 30% vol/vol isoflurane in propylene glycol, Dainippon Sumitomo Pharma Co., Ltd, Osaka, Japan) in a bell jar and killed using focused microwave irradiation (MI) (10 kW, 1.2 s; NJE-2603, New Japan Radio Co., Ltd, Tokyo, Japan). Following the MI, five brain loci (cortex, hippocampus, hypothalamus, cerebellum and brainstem) were collected using a method modified from Hirano et al. (2006). Two skeletal muscles (soleus and plantaris), liver and blood were also collected. All samples were stored at –80°C for subsequent biochemical analyses.

Blood glucose, lactate and insulin assays

Blood glucose and lactate levels were measured using an automated glucose–lactate analyser (2300 Stat Plus, Yellow Springs Instruments, USA). Plasma level of insulin was determined using a commercially available rat insulin ELISA kit (Shibayagi Co. Ltd, Gunma, Japan).

Glycogen assays

Glycogen measurements were taken according to the method described by Matsui et al. (2011). Tissues were homogenized (Polytron, Kinematica, Kriens-Luzern, Switzerland; setting 6; 60 s) in ice-cold 6% perchloric acid (PCA) containing 1 mm EDTA. For tissue glycogen content measurements, glycogen was hydrolysed to glucose in 100 μl aliquots of homogenate that were removed and incubated for 1 h at 37°C with 1 ml of 0.2 m sodium acetate, 20 μl of 1.0 m KHCO3 and 20 U ml−1 of amyloglucosidase. The addition of 0.5 ml of PCA solution stopped the reaction. After centrifugation (14,000 g for 10 min at 4°C), the supernatant was neutralized with a KOH solution consisting of 3 m KOH, 0.3 m imidazole and 0.4 m KCl. The supernatant was then centrifuged (16,000 g for 10 min at 4°C) and assayed for glucose content. To measure endogenous (background) glucose levels, non-hydrolysed samples were obtained by centrifuging the homogenate (14,000 g for 10 min at 4°C) and adjusting the pH of the supernatants to a final level of 6–8 with KOH solution. Neutralized samples were mixed thoroughly, centrifuged (16,000 g for 10 min at 4°C), and assayed for endogenous glucose levels. The glucose content assay was performed in 96-well plates using a coupled enzyme assay method modified from a previous study (Passonneau & Lauderdale, 1974)). A total of 200 μl of a reaction solution containing 50 mm Tris-HCl (pH 8.1), 0.5 mm ATP, 0.5 mm NADP+, 5 mm MgSO4 and 0.1 U ml−1 glucose-6-phosphate dehydrogenase was added to each well. The plate was then placed in a fluorescence plate reader (Arvo, Perkin Elmer, Groningen, the Netherlands) and shaken, and measurements of the resultant NADPH were taken at 350 nm excitation and 450 nm emission. After the addition of hexokinase (0.3 U) to each well, plates were shaken and measurements were taken after a 30 min incubation period. Tissue glycogen levels, indicated as glucose units, were calculated by subtracting the final micromolar concentration of glucose per gram of wet weight of the non-hydrolysed tissue sample from the micromolar concentration of glucose per gram of wet weight of the hydrolysed tissue sample.

Citrate synthase activity assays

Citrate synthase (CS) activity was measured using the method described by Siu et al. (2003). The soleus muscle was homogenized on ice in 0.1 m Tris buffer containing 0.1% Triton X-100, pH 8.35. The homogenates were frozen under liquid nitrogen and thawed four times to disrupt the mitochondria and expose the CS. The assay system contained a total volume of 200 μl of 100 mm Tris buffer (pH 8.35), 5 mm DTNB, 22.5 mm acetyl-CoA, 25 mm OAA and 4 μl of homogenate of muscle. The principle of the assay was to initiate the reaction of acetyl-CoA with OAA and link the release of free CoA-SH to a colourimetric reagent, DTNB. The rate change in colour was monitored at a wavelength of 400 nm at 15 s intervals for 3 min using a plate reader (Arvo). All measurements were performed in duplicate, with the same settings at 20–22°C. The CS activity was then normalized to the total protein content and was reported as micromoles per milligram protein per minute.

Statistics

Data are expressed as mean ± standard error (SEM) and were analysed using Prism 5 (MDF Co., Ltd, Tokyo, Japan). Group comparisons were performed using a one-way ANOVA with Dunnett's post hoc tests. Comparisons of two groups were performed using Student's t test for unpaired data. Correlations were calculated using Pearson's product–moment correlations. Statistical significance was assumed at P values < 0.05.

Results

Blood parameters and glycogen in the skeletal muscles and liver after exhaustive exercise

At the end of exhaustive exercise (time to exhaustion, 81.2 ± 2.7 min), blood glucose levels significantly decreased compared with pre-exercise levels (F = 70.0, P < 0.01), and quickly recovered after cessation of exercise (Fig. 1B)). Blood lactate levels also significantly increased compared with pre-exercise levels (F = 106.6, P < 0.01), and returned to pre-exercise levels within 3 h after exercise (Fig. 1B)). Glycogen levels in both slow-twitch soleus and fast-twitch plantaris muscles were depleted by 82% and 90% (P < 0.01), respectively, with exhaustive exercise. After the cessation of exercise, depleted muscle glycogen levels in both types of muscles returned to pre-exercise levels within 3 h and increased by 43–46% above pre-exercise levels (glycogen supercompensation) at 24 h after exercise (P < 0.01) (Fig. 1C)). Liver glycogen was also depleted by 96% with exhaustive exercise (P < 0.01), but supercompensation did not occur (Fig. 1D)). Blood insulin levels were significantly lower as compared with pre-exercise levels, and returned to pre-exercise levels at 24 h post-exercise (P < 0.05) (Fig. 1E)). Blood insulin levels at 6 and 24 h after exercise were positively correlated with liver glycogen levels (r = 0.64, P < 0.01), but showed no correlation with glycogen levels in skeletal muscles (data not shown).

Brain glycogen after exhaustive exercise

Glycogen levels in the five brain loci were significantly decreased by 50–63.6% (whole brain 57%, cortex 61%, hippocampus 55%, hypothalamus 50%, cerebellum 64% and brainstem 54%) with exhaustive exercise (P < 0.01), and supercompensation occurred of 29–63% within 6 h after the cessation of exercise (P < 0.05) (Fig. 2)). Glycogen levels in the cortex and brainstem were supercompensated within 3 h after exercise (P < 0.05), and supercompensated glycogen levels in the cortex and hippocampus were sustained for 24 h after exercise (P < 0.05) (Fig. 2)). In sedentary rats, brain glycogen levels did not change at 6 and 24 h after being placed on a stationary treadmill (data not shown). Brain glycogen levels did not correlate with blood insulin levels (data not shown).

Figure 2. Glycogen levels in the brain after exhaustive exercise.

Figure 2

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Data represent the mean ± SEM (n = 8 – 13 rats). *P < 0.05; **P < 0.01 compared to pre-exercised rats (Dunnett's post hoc test).

Glycogen supercompensation rates in the skeletal muscle, liver and brain

The rates of supercompensation peak were 29–63% (soleus 43%, plantaris 46%, liver –2.9%, whole brain 46%, cortex 60%, hippocampus 33%, hypothalamus 29%, cerebellum 63% and brainstem 49%; Fig. 3A)). In brain regions, glycogen supercompensation rates positively correlated with their corresponding decrease rates (r = 0.89, P < 0.05; Fig. 3B)). The peak of skeletal muscle glycogen supercompensation was confirmed at 24 h after exercise, while the peak of glycogen supercompensation in the brain occurred at 6 h after exercise. Liver glycogen supercompensation was not observed in this study (Fig. 4)).

Figure 3. Rates of glycogen decrease and supercompensation peak following exhaustive exercise.

Figure 3

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A, rates of glycogen decrease and supercompensation peak in skeletal muscles, liver and brain. Data of the supercompensation peak in skeletal muscle and liver are at 24 h after exercise, and data of the supercompensation peak in the brain are at 6 h after exercise. Data represent the mean ± SEM (n = 8–13 rats). **P < 0.01 compared to pre-exercised rats (unpaired t test). B, correlation between rates of glycogen decrease and rates of supercompensation peak in the whole brain (Pearson's product–moment correlations).

Figure 4. The glycogen dynamics during and following exhaustive exercise in the brain, skeletal muscle and liver.

Figure 4

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Data from pre-exercise to 24 h after exercise are taken from Figs 1 and 2, and data from 48 h to 72 h after exercise are extrapolated based on our unpublished data. During exhaustive exercise, brain glycogen decreases by approximately 50–60%, while glycogen in skeletal muscle and liver decreases by approximately 80–90%. Following exercise, in the resting phase, skeletal muscle glycogen supercompensation occurs at 24 h after exercise, and returns to pre-exercise level at 72 h after exercise. Brain glycogen supercompensation also occurs and reaches the peak of supercompensation at 6 h after exercise, and returns to pre-exercise level at 48 h after exercise. Liver glycogen is not completely replenished until 48 h after exercise.

Brain glycogen levels after exercise training

Both body and fat weight decreased significantly in exercise-trained rats compared with sedentary rats, and both soleus and plantaris muscle weights increased significantly (P < 0.01) (Fig. 5B and C)). The CS activity and glycogen level in the soleus muscle also increased (P < 0.01) (Fig. 5D and E)). Glycogen levels in the cortex and hippocampus of trained rats were 7% and 9% higher than those of sedentary rats (P < 0.05), while glycogen levels in the hypothalamus, cerebellum and brainstem were not changed (Fig. 5F)).

Figure 5. Glycogen levels in the brain after exercise training.

Figure 5

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A, experimental procedure; B, body and epididymal fat weight; C, skeletal muscle weights; D, soleus CS activity; E, glycogen levels in the soleus and liver; F, glycogen levels in the brain. Data represent the mean ± SEM (n = 9–10 rats). *P < 0.05; **P < 0.01 compared to sedentary rats (unpaired t test).

Discussion

To the best of our knowledge, this is the first study testing the hypothesis that brain glycogen supercompensation occurs following exhaustive exercise as it does in skeletal muscles, and the extent of supercompensation is dependent on that of glycogen decrease during exercise. We also observed that cortical and hippocampal glycogen supercompensation were sustained until 24 h after exercise (long-lasting supercompensation), and their basal glycogen levels increased with 4 weeks of exercise training (60 min day−1, 5 days week−1 at 20 m min−1) compared to sedentary rats.

The extent of post-exercise brain glycogen supercompensation is dependent on that of glycogen decrease during exercise, across brain regions. Greater glycogen depletion induces greater glycogen supercompensation in skeletal muscle (Goforth et al. 2003)), suggesting muscle glycogen supercompensation is inducible in more active muscle for metabolic adaptation. In the present study, the highest rate of glycogen decrease and supercompensation was seen in the cerebellum (64 vs. 63%), the second highest was in the cortex (61 vs. 60%) and the lowest was seen in the hypothalamus (50 vs. 29%) (Fig. 3A)). Vissing et al. (1996) examined glucose utilization as an index of functional neuronal activity during 30 min of high-intensity running (28 m min−1, ≤85% of maximum oxygen consumption), and found that cerebellar and cortical glucose utilization are higher than hypothalamic use. Thus, in the brain, greater neuronal activity could induce greater glycogen supercompensation as seen in skeletal muscle.

During the recovery phase after exhaustive exercise, glycogen supercompensation in the brain occurs earlier than that of skeletal muscles and the liver (for the brain, 6 h; for skeletal muscle and liver, 24 h and above; Fig. 4)). These results are consistent with the previous findings that skeletal muscles exhibit glycogen supercompensation at 24 h after exhaustive exercise (Gaesser & Brooks, 1980)), and that the brain exhibits rapid glycogen replenishment and supercompensation at 4-7 h after insulin-induced hypoglycaemia in rats (Choi et al. 2003; Canada et al. 2011)). These findings support the ‘Selfish Brain Theory’ regarding competition for energy resources throughout the whole body (Peters et al. 2004)). Brain glycogen depletion with severe, insulin-induced hypoglycaemia elicits neuronal death (Suh et al. 2007)). Severe hypoglycaemia in animals and humans induces a hypoglycaemic coma to save energy and, ultimately, their life (Sokoloff, 1971)). Thus, rapid and preferential brain glycogen supercompensation following exhaustive exercise with hypoglycaemia may also be induced in an attempt to prevent neuronal death and to save the animals’ lives (preservation of species), and could cause metabolic adaptation in the brain for delaying hypoglycaemic coma.

Perhaps increased basal glycogen levels in the cortex and hippocampus with exercise training are a result of the accumulation of long-lasting supercompensation with acute exercise. In the exhaustive exercise test, the long-lasting glycogen supercompensation following exhaustive exercise was confirmed at 24 h after exercise in the cortex and hippocampus, but not in the cerebellum, hypothalamus or brainstem (Fig. 2)). In a previous study, without MI, glycogen supercompensation in the whole brain was confirmed following insulin-induced hypoglycaemia, but long-lasting supercompensation was not observed (Canada et al. 2011)). These findings suggest that the long-lasting glycogen supercompensation could be an exercise-specific effect with exercise-induced neuronal activation in the brain.

Increased brain glycogen levels must be adaptive metabolic changes in the brain in response to increased energy demand with endurance training. Increases in cortical and hippocampal glycogen levels accompany the typical indications of endurance training, such as reduced body and fat weights, and increased wet weights, glycogen levels and CS activity in the soleus muscle (slow-twitch type) (Fig. 5B–D)) (James & Kraegen, 1984; Siu et al. 2003)). Brain mitochondrial biogenesis increases with exercise training, as in skeletal muscle (Davis et al. 2009; Steiner et al. 2011)). In addition to increased mitochondrial biogenesis, an increase in brain glycogen with exercise training could be an important phenomenon of the adaptation to meet the increased energy demands of the brain in exercising animals. If so, the mRNA and protein content of glucose transporters and monocarboxylic acid transporters in astrocytes and neurons might also increase for activation of glycogen metabolism. However, these were not examined here because the heat of MI would have damaged mRNA and protein. We are examining whether the detection of mRNA and protein in the microwaved brain is possible.

Increased basal glycogen levels in the cortex and hippocampus with exercise training could have an important implication for their function. Cortical neuronal activation time during hypoglycaemia was increased by elevating the basal level of cortical glycogen with the injection of a glycogen phosphorylase inhibitor (Suh et al. 2007)). Thus, we can hypothesize that increased basal levels of cortical glycogen in exercise-trained animals prevents the attenuation of neuronal activity in the cortex during prolonged exercise with hypoglycaemia and sustains central command from the motor cortex to skeletal muscles, and that this is a mitigating factor of central fatigue. An increase in the basal level of cortical glycogen may enhance endurance performance. On the other hand, an exercise training-induced increase in hippocampal glycogen may be involved in cognitive function. It is well known that exercise training enhances cognitive function, especially hippocampal function (van Praag, 2009)). Hippocampal glycogen levels in Zucker rats, a type 2 diabetes model animal, were lower than those seen in control rats, which may be related to the decline of cognitive function in these animals (Sickmann et al. 2010)). Furthermore, the inhibition of hippocampal glycogenolysis in rats by 1,4-dideoxy-1,4-imino-d-arabinitol (DAB, a glycogen phosphorylase inhibitor), interdicted long-term memory formation (Suzuki et al. 2011)). Therefore, it is tempting to speculate that increased hippocampal basal glycogen might be involved in promoting cognitive function with exercise training.

Underlying molecular mechanisms behind brain glycogen replenishment and supercompensation are controversial, with limited information provided by in vitro studies. Among them, insulin and insulin-like growth factor I (IGF-I) have been implicated in the regulation of astrocytic glycogen synthesis as well as that of peripheral organs (Dringen & Hamprecht, 1992)). However, we could not find a correlation between blood insulin and supercompensated brain glycogen levels during glycogen repletion after exercise, although we found a positive correlation between blood insulin and liver glycogen levels (r = 0.64, P < 0.01) (data not shown). On the other hand, we found that regional blood–brain barrier transport of IGF-I was enhanced by neuronal activation during exercise and/or exposure to an enriched environment (Nishijima et al. 2010)). Thus, we should explore whether IGF-I signalling is involved in the replenishment and supercompensation of brain glycogen.

Further, it is of interest that the neurotransmitters NA and vasoactive intestinal peptide (VIP), which have been identified as neuronal signals triggering glycogenolysis in astrocytes, cause the induction of protein targeting to glycogen (PTG) in a non-insulin-dependent fashion and the subsequent enhancement of glycogen synthesis in primary culture of astrocytes (Sorg & Magistretti, 1992; Allaman et al. 2000)). NA and its metabolism in the brain has been shown to increase during exercise (Pagliari & Peyrin, 1995; Matsui et al. 2011)). Thus, it is hypothesized that increased cerebral NA stimulates astrocytic glycogenolysis during exhaustive exercise and expression of PTG, which may in turn lead to glycogen supercompensation.

In conclusion, we have provided evidence that brain glycogen supercompensation occurs in the brain, as in skeletal muscle, following exhaustive exercise. Furthermore, the lower the brain glycogen level during exercise, the higher the extent of glycogen supercompensation after exercise. In addition, the brain glycogen supercompensation peak preceded that of skeletal muscles and liver. We also propose that long-lasting glycogen supercompensation is likely to be a prerequisite for training adaptation (increased basal levels), probably to meet the increased energy demand of the brain in exercising animals.

Acknowledgments

We thank Professor Randeep Rakwal (University of Tsukuba) for his assistance in preparing the manuscript. This work was supported in part by Grant-in-Aid for Challenging Exploratory Research of the Japan Society for the Promotion of Science (JSPS) (No. 21650166), by Grant-in-Aid for JSPS Fellows (No. 10J00513), by a grant from the Kozuki Foundation for Sports and Education, and by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) for the Body and Mind Integrated Sports Sciences (BAMIS) Project (2010). T.M. and M.O. are both Research Fellows of JSPS.

Glossary

Abbreviations

CS

citrate synthase

DTNB

5,5-dithiobis (2-nitrobenzoate)

5-HT

5-hydroxytryptamine

IGF-I

insulin-like growth factor I

MI

microwave irradiation

NA

noradrenaline

OAA

oxaloacetate

PCA

perchloric acid

PTG

protein targeting to glycogen

VIP

vasoactive intestinal peptide

Author contributions

Experiments were performed at the Laboratory of Exercise and Neuroendocrinology at the University of Tsukuba. Individual author contributions were as follows: conception and design of the experiments (T.M. and H.S.); collection of data (T.M., T.I., H.I., M.O., K.I. and M.-c.L.); analysis and interpretation of data (T.M., T.F., Y.I., K.K. and H.S.); drafting the article and revising it critically for important intellectual content (T.M. and H.S.). All authors approved the final version.

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