Umeko Marubayashi

Evidence that tryptophan reduces mechanical efficiency and running performance in rats

It has been reported that exercise increases brain tryptophan (TRP), which is related to exhaustive fatigue. To study this further, the effect of increased TRP availability on the central nervous system (CNS) with regard to mechanical efficiency, oxygen consumption (VO(2)) and run-time to exhaustion was studied in normal untrained rats. Each rat was anesthetized with thiopental (30 mg/kg ip b. wt.) and fitted with a chronic guiding cannula attached to the right lateral cerebral ventricle 1 week prior to the experiments. Immediately before exercise, the rats were randomly injected through these cannulae with 2.0 microl of 0.15 M NaCl (n=6) or 20.3 microM L-TRP solution (n=6). Exercise consisted of running on a treadmill at 18 m min(-1) and 5% inclination until exhaustion. TRP-treated rats presented a decrease in their mechanical efficiency (21.25+/-0.84%, TRP group vs. 24.31+/-0.98%, saline-treated group; P< or =.05), and increased VO(2) at exhaustion (40.3+/-1.6 ml kg(-1) min(-1), TRP group vs. 36.0+/-0.8 ml kg(-1) min(-1), saline group; P< or =.05), indicating that the metabolic cost of exercise was higher in the former group. In addition, a highly significant reduction was also observed in run-time to exhaustion of TRP animals compared to those of the saline-treated group (15.2+/-1.52 min, TRP group vs. 50.6+/-5.4 min, saline group; P< or =.0001). It can be deduced from the data that intracerebroventricular TRP injection in rats increases O(2) consumption and reduces mechanical efficiency during exercise, diminishing running performance.

Tryptophan-induced central fatigue in exercising rats is related to serotonin content in preoptic area.

To assess the effects of increased hypothalamic tryptophan (TRP) availability on 5-HT content in preoptic area on thermoregulation and work production during exercise on treadmill, 20.3 microM of L-TRP (n=7) or 0.15M NaCl (n=6) was injected into the lateral cerebral ventricle of male Wistar rats immediately before the animals started running (18 m min(-1) 5% inclination). Exercise time to fatigue (min), and workload (kgm) were analysed. Core temperature was measured by telemetry. At fatigue, brains were quickly removed and preoptic area (POA), hypothalamus (HP), frontal cortex (FC), hippocampi (HC) were rapidly dissected and frozen immediately in dry ice. Serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were measured by HPLC. TRP-exercised rats showed the highest content of 5-HT in the POA and the lowest in the hippocampi compared to the rested and SAL-exercised rats. An inverse relationship between TF and a direct correlation with body temperature changes and POA-5HT levels were observed. A correlation between HC 5-HT content and TF was also found. However, there was no correlation between HC 5-HT content and changes in Tb at fatigue. Finally, our results bring further evidences that increased 5-HT content in POA is involved with an increase in heat production during exercise. In addition, the direct correlation of 5-HT level in hippocampi and TF of TRP-exercised rats suggests that this brain area is also related to motor activity control during exercise. In conclusion, our data indicated that tryptophan-induced central fatigue in exercising rats is related to serotonin content in preoptic area.

Evidence that brain nitric oxide inhibition increases metabolic cost of exercise, reducing running performance in rats.

To assess the role of nitric oxide (NO) in the metabolic rate and running performance of rats submitted to exercise on a treadmill, 1.43 micromol (2 microL) of Nomega-nitro-L-arginine methyl ester (L-NAME, n=6), a NO synthase inhibitor, or 2 microL of 0.15M NaCl (SAL, n=6) was injected into the lateral cerebral ventricle of male Wistar rats immediately before the animals started running (18m min(-1), 5% inclination). Oxygen consumption (VO2) was measured at rest, during the exercise until fatigue and thereafter during the 30 min of recovery using the indirect calorimetry system. Mechanical efficiency (ME) was also calculated during the running period. During the first 11 min of exercise, there was a similar increase in VO2 while ME remained the same in both groups. Thereafter, VO2 remained stable in the SAL group but continued to increase and remained higher in the L-NAME group until fatigue. The L-NAME-treated rats also showed a sharper decrease in ME than controls. In addition, there was a significant reduction in workload performance by L-NAME-treated animals compared to SAL-treated animals. This suggests that central blockage of nitric oxide increases metabolic cost during exercise, reduces mechanical efficiency and decreases running performance in rats.

Nitric oxide pathway is an important modulator of heat loss in rats during exercise

To assess the role of nitric oxide (NO) in central thermoregulatory mechanisms during exercise, 1.43 micromol (2 microL) of N(omega)-nitro-L-arginine methyl ester (L-NAME, n=6), a NO synthase inhibitor, or 2 microL of 0.15M NaCl (SAL, n=6) was injected into the lateral cerebral ventricle of male Wistar rats immediately before the animals started running (18 m min(-1), 5% inclination). Core (Tb) and skin tail (Ttail) temperatures were measured. Body heating rate (BHR), threshold Tb for tail vasodilation (TTbV), and workload (W) were calculated. During the first 11 min of exercise, there was a greater increase in Tb in the L-NAME group than in the SAL group (BRH=0.17+/-0.02 degrees C min(-1), L-NAME, versus 0.09+/-0.01 degrees C min(-1), SAL, p<0.05). Following the first 11 min until approximately 40 min of exercise, Tb levels remained stable in both groups, but levels remained higher in the L-NAME group than in the SAL group (39.16+/-0.04 degrees C, L-NAME, versus 38.33+/-0.02 degrees C, SAL, p<0.01). However, exercise went on to induce an additional rise in Tb in the SAL group prior to fatigue. These results suggest that the reduced W observed in L-NAME-treated rats (10.8+/-2.0 kg m, L-NAME, versus 25.0+/-2.1 kg m, SAL, p<0.01) was related to the increased BHR in L-NAME-treated animals observed during the first 11 min of exercise (r=0.74, p<0.01) due to the change in TTbV (39.12+/-0.24 degrees C, L-NAME, versus 38.27+/-0.10 degrees C, SAL, p<0.05). Finally, our data suggest that the central nitric oxide pathway modulates mechanisms of heat dissipation during exercise through an inhibitory mechanism.

Intracerebroventricular tryptophan increases heating and heat storage rate in exercising rats.

The role of increased hypothalamic tryptophan (TRP) availability on thermoregulation and rates of core temperature increase and heat storage (HS) during exercise was studied in normal untrained rats running until fatigue. The rats were each anesthetized with 2.5% tribromoethanol (1.0 ml kg(-1) ip) and fitted with a chronic guiding cannula attached to the right lateral cerebral ventricle 1 week prior to the experiments. Immediately before exercise, they were randomly injected through these cannulae with 2.0 microl of 0.15 M NaCl (SAL; n=6) or 20.3 microM L-TRP solution (n=7). Exercise consisted of running on a treadmill at 18 m min(-1) and 5% inclination until fatigue. Body temperature was recorded before and during exercise with a thermistor probe implanted into the peritoneal area. Rates of core temperature increase (HR, degrees C min(-1)) and heat storage (HSR, cal min(-1)) were calculated. TRP-treated rats showed a rapid increase in body temperature which was faster than that observed in the saline-treated group during the exercise period. The TRP group also showed a higher rate of core temperature increase and HS. TRP-treated rats that presented higher HR and HSR also fatigued much earlier than saline-treated animals (16.8+/-1.1 min TRP vs. 40+/-3 min SAL). This suggests that the reduced running performance observed in TRP-treated rats is related to increased HR and HSR induced by intracerebroventricular injection of TRP in these animals.

Effect of intracerebroventricular injection of atropine on metabolic responses during exercise in untrained rats

To investigate the role of the central cholinergic system in the regulation of metabolism during exercise, we injected atropine (5 x 10(-7) mol) into the lateral cerebral ventricle of normal and adrenodemedullated (ADM) untrained rats submitted to exercise on a treadmill (15 m min(-1), 5% grade) until exhaustion. Concentrations of blood glucose, plasma free fatty acids (FFA), and lactate were measured before and every 10 min after the start of exercise for a period of 60 min. Adrenomedullectomy had no effect on the maximal capacity of exercise (MCE), but atropine administered intracerebroventricularly (i.c.v.) reduced the maximal capacity of exercise of both normal and ADM rats. In normal rats, blood concentrations of glucose and plasma free fatty acids remained essentially unchanged compared to the levels at rest, whereas in ADM rats a rapid increase in plasma glucose and plasma free fatty acids levels occurred during exercise. These data indicate that adrenomedullectomy disrupted the accuracy of the feedback mechanism that regulates the mobilization of extramuscular fuels during exercise in normal rats. In addition, ADM rats showed an increased lipid mobilization as a source of energy during exercise, which might explain the increased plasma glucose by an inhibition of muscle glucose uptake. These results suggest that central cholinergic neurons might be involved in the control of energy substrate adjustment during exercise, thereby reducing the maximal capacity of exercise. In addition, the results of this study suggest that the adrenal glands are important for an accurate feedback mechanism during exercise.

Muscarinic cholinoceptors in the ventromedial hypothalamic nucleus facilitate tail heat loss during physical exercise

The aim of this study was to evaluate the participation of ventromedial hypothalamic nucleus (VMH) muscarinic cholinoceptors in heat balance and central fatigue during treadmill exercise (24 m min(-1), 5% inclination). The animals were anesthetized with pentobarbital sodium (50 mg/kg body weight i.p.) and fitted with bilateral cannulae into the VMH 1 week prior to the experiments. Tail skin (T(tail)) and core body temperatures (T(b)) were measured after the injection of 0.2 microL of 5 x 10(-9) mol methylatropine (Matr) or 0.15 M NaCl solution (Sal) into the hypothalamus. Methylatropine injection into the VMH greatly increased heat storage rate (HSR) measured until fatigue (19.7+/-4.6 cal min(-1) Matr versus 9.7+/-3.3 cal min(-1) Sal; P<0.05) and attenuated the exercise-induced tail vasodilation as seen by T(tail) (23.98+/-0.43 degrees C Matr versus 25.52+/-0.85 degrees C Sal; at 6.5 min; P<0.05), indicating inhibition of the heat loss process. The 2 min delay and the increased DeltaT(b), which triggered the heat loss mechanisms observed in Matr-treated rats, are associated with increased HSR and may be responsible for the decreased running performance of these animals (21.0+/-2.9 min Matr versus 33.5+/-3.4 min Sal; P<0.001). In fact, a close negative correlation was observed between HSR and time to fatigue (r=-0.61; P<0.01). In conclusion, VMH muscarinic cholinoceptors facilitate tail heat loss mechanisms, and a delay in this adjustment would lead to a decrease in physical exercise performance due to excess heat storage.

Central nitric oxide inhibition modifies metabolic adjustments induced by exercise in rats.

The influence of the central nervous system on metabolic function is of interest in situations deviating from basal states, such as during exercise. Our previous study in rats demonstrated that central nitric oxide (NO) blockade increases metabolic rate, reducing mechanical efficiency during exercise. To assess the role of brain nitric oxide in the plasma glucose, lactate and free fatty acids (FFAs) concentrations of rats submitted to an incremental exercise protocol on a treadmill until fatigue, 1.43 micromol (2 microl) of N(omega)-nitro-l-arginine methyl ester (L-NAME, n=6), a NO synthase inhibitor, or 2 microl of 0.15M NaCl (SAL, n=6) was injected into the lateral cerebral ventricle (icv) of male Wistar rats immediately before exercise (starting at 10 m/min, with increments of 1m/min every 3 min until fatigue, 10% inclination). Blood samples were collected through a chronic jugular catheter at rest and during exercise until fatigue. During exercise, the L-NAME-treated animals had the following metabolic response compared to controls: (1) an increased hyperglycemic response during the first 60% of time to fatigue; (2) higher plasma lactate levels; and (3) a significant transitory increase in plasma free fatty acids during the dynamic phase of exercise that returned to basal levels earlier than controls during the steady state phase of exercise. In addition L-NAME-treated rats fatigued earlier than controls. The data indicate that the inhibition of the brain nitrergic system induced by icv L-NAME treatment disrupted the accuracy of the neural mechanism that regulates plasma glucose and free fatty acids mobilization during exercise in rats.

The medial preoptic area modulates the increase in plasma glucose and free fatty acid mobilization induced by acute cold exposure

The effect of cold exposure (0 degrees C, 1 h) on free fatty acid (FFA) mobilization and plasma glucose was studied in freely moving male rats injected with adrenergic blockers (phentolamine or propranolol) into the medial preoptic area (MPOA). The rats were implanted with chronic jugular catheters for blood sampling and with unilateral intracerebral cannulas placed just above the MPOA. Blood samples were taken 2 min before and 10, 20, 40 and 60 min after cold exposure. After cold exposure plasma glucose and plasma FFA levels rose rapidly, reaching a peak at 20 min post-stimulus. Previous administration of phentolamine (50 nmol), but not propranolol (100 nmol), into the MPOA blocked the glycemic response to cold exposure. On the other hand, previous administration of propranolol, but not phentolamine, into the MPOA blocked the increased FFA mobilization in response to cold exposure. On the basis of these results, we propose that MPOA alpha-adrenergic synapses relay impulses activating the sympathetic outflow expressed by neurally mediated hyperglycemia, and beta-adrenergic synapses relay impulses activating the sympathetic outflow to the adipose tissue, increasing FFA acid mobilization.

Prolactin release during exercise in normal and adrenodemedullated untrained rats submitted to central cholinergic blockade with atropine.

To study the role of the central cholinergic system in pituitary prolactin (PRL) release during exercise we injected atropine (5 x 10(-7) mol) into the lateral cerebral ventricle of intact or adrenodemedullated (ADM) untrained rats, at rest or submitted to exercise on a treadmill (18 m x min(-1), 5% grade) until exhaustion. The rats were implanted with chronic jugular catheters for blood sampling and with unilateral intracerebroventricular (icv) cannulas placed in the right lateral ventricle. Blood prolactin concentrations were measured before and every 10 min after the start of exercise for a period of 60 min. After the animals started running, plasma prolactin levels rose rapidly in both normal and ADM rats, reaching near maximum at 10 min. Close to exhaustion (19.8 +/- 2.9 min for intact rats and 23.5 +/- 4.1 min for ADM) they were still high, remained increased until 30 min, and returned to preexercise levels at 40 min. Icv injections of atropine decreased the time to exhaustion by 67% in intact rats and by 96.2% in ADM and also reduced the exercise-induced PRL release in both intact (50%) and ADM rats (90%). The results showed that prolactin release induced by exercise was dependent on the exercise workload and could be observed as early as after 10 min of running, remaining increased until 30 min. These data indicate that adrenodemedullation does not affect prolactin secretion induced by exercise, although adrenodemedullated rats proved to be more sensitive to the reducing effect of central cholinergic blockade on their maximal capacity for exercise.