Studies On The Antifatigue Effects Of Capsaicin On Mice And Its Mechanisms

Partâ… BackgroundFatigue is a normal physiological phenomenon and it occurs when mental or physical strength reseaches a certain level.Fatigue is a complicated physiological change.With the speeding up of the social rhythm and increasing of social competition,people always stay in tension when working and studying and their physical strength and energy are always in the long-term overdrawn.According to the investigation result of WHO,over 35%of people are in fatigue state,the rate of fatigue among middle aged men reached to 60.70%,nearly 70% of the middle-aged are in fatigue state in China.Fatigue can lead to the decline of people's performance.If the fatigue can't recover in time, "Overtraining syndrome,OTS" and "Chronic fatigue syndrome,CFS" would occur and lead to endocrine disturbance,immunity decrease and threaten people's health.At present,various measures to eliminate fatigue include rest,sleep, massage and other physical methods;People also depend on nutritive fatigue supplements and traditional medical therapy aiming at relieving. However,these effects also have something unsatisfactory.Consequently, active substances extracted from the natural products have become the focus of the research for a safer,more effective methords of relieving fatigue since they have less side effects and not contain harmful components of doping for people.Capsaicin,the major pungent ingredient in red pepper,exerts extensive biological and pharmacological activities via activating vanilloid receptor-1(VR1),which may be fully released in the cerebral and periphery nervous system.Recently,it is reported that capsaicin could improve the capacity of exercise,which means that capsaicin may have the effect of antifatigue.However,so far,the study of the effect of capsaicin on antifatigue and its dose-effect relation is still not so profound and systematic now and there are no reports about the related signal molecular and the signal translation of capsaicin in the antifatigue effect study.Partâ…¡.The Antifatigue effects of Capsaicin on miceAIM:To explore the antifatigue effects of capsaicin on mice.METHODS:168 normal male KunMing mice were randomly divided into 7 groups based on body weight:low,medium,medium-high, high dose capsaicin intervention groups,control group and positive control group.The control group was given distilled water,the four intervention groups were given different doses of capsaicin(3,6,10,15mg/kg·bw) respectively and the positive control group were treated with Rhodiola Rosea(1g/kg·bw/d).After 4 weeks,the weight of each mouse was measured.In addition,the pole-climbing time,the loading swimming time,serum urea nitrogrn,the hepatic glycogen,the muscle glycogen,the content of blood lactic acid and the activity of lactate dehydrogenase are all taken as the index of the antifatigue effects;and the toxicosis experiment of the second sodium,the survival time of mice under anoxia are taken as the index of anti-anoxic.RESULTS:(1) There is no significant difference between the capsaicin treated group and the control group.(2) Capsaicin significantly increased the time of climbing pole and loading swimming time of the mice,the maximum effect of capsaicin was observed at 15mg/kg·bw.(3) There was no significant difference of the survival time of mice under normal pressure anoxia and acute ischemic hypoxia between the low dose capsaicin treated group(3mg/kg) and the control group (P＞0.05);different concentration of capsaicin(6,10,15mg/kg)can significantly extend the survival time of mice under the normal pressure anoxia and acute ischemic hypoxia compared with the control group(P＜0.05);the maximal effect was observed at 15mg/kg·bw,the survival time under normal pressure anoxia and acute ischemic hypoxia were extended 64%and 107%respectively,there was no significant difference between the capsaicin 15mg/kg·bw treated group and the Rhodiola Rosea(1g/kg·bw/d) treated group.(4) Compared with the control group,the capsaicin has significantly improved the content of hepatic glycogen and muscle glycogen and the activity of the lactate dehydrogenase after exercise,meanwhile,the level of serum urea nitrogrn and the content of blood lactic acid were significantly decreased,the maximal effect were both observed at 15mg/kg.(5) Compared with the control group,capsaicin of different concentration can significantly lengthen the survival time under the toxicosis experiment of the sodium nitrite(P＜0.05).CONCLUSION:Capsaicin has obvious antifatigue effects.It has significantly lengthened the time of pole climbing and loading swimming of the mice,improved the capacity of the anti-hypoxia,and inhibited the generation of blood lactic acid and serum urea nitrogen after exercise, maintained the content of the hepatic and muscle glycogen.Partâ…¢.The antifatigue mechanism of CapsaicinAIM:To investigate the antifatigue mechanism of Capsaicin on mice from several aspects including antioxidant activity,energy metabolism and central system using the appropriate concentration based depending on the results from Partâ… .METHODS:(1) antioxygen system:â‘ measuring the activity of hepatic superoxide dismutase,glutathion peroxidase and the content of hepatic malonaldehyde before and after exercise;â‘¡measuring the plasma content of creatine kinase and MB isoenzyme of creatine kinase of the mice before and after exercise as the indexes of the damage of muscle cells.(2)energy metabolism system:â‘ detecting the blood glucose,muscle and hepatic glycogen of mice before and after exercise; â‘¡detecting the content of plasma free fatty acid before and after exercise;â‘¢measuring content of total protein,albumin and globulin plasma.(3) central system:â‘ After sleep deprivation for 48h and 72h,all the rats were put to an openfield test and then the calculation of the score of openfield test follows;â‘¡Killed the rats and took the organs of hypothalamus,brain stem immediately,then measured the 5-hydroxytryptamine, NE,Dopamine,Ad,the content of 5-HIAA of the hypothalamus and brain stem,and detecting the content of plasma NH₃.RESULTS:(1) antioxygen system:â‘ There is no significant differentce in the activity of hepatic SOD,GSH-Px and the level of MDA between the control and the capsaicin treated group before exercise(P＞0.05;after exercise,the activity of hepatic SOD,GSH-Px of each group all decreased,however,capsaicin significantly increased the activityies of hepatic SOD,GSH-Px compared with the control group after exercise and reduced the content of MDA(P＜0.05 or P＜0.01 ),the maximal effect was observed at 15mg/kg·bw.â‘¡Before exercise,there is no significant difference of the levels of CK and CK-MB in plasma between the control and the capsaicin treated group.After exercise,capsaicin significantly reduced the levels of CK and CK-MB in plasma compared with the control group(P＜0.05 or P＜0.01 ),the maximal effect was observed at 15mg/kg·bw.(2) energy metabolism system:â‘ There is no significant differentce in the blood glucose between the control and the capsaicin treated group before exercise(P＞0.05);After exercise,the blood glucose all decreased,however,the capsaicin significantly increased the blood glucose(P＜0.05 or P＜0.01 ),the maximal effect was observed at 15mg/kg·bw.Capsaicin significantly increased the content of hepatic and muscle glycogen after exercise.â‘¡Capsaicin increased the serum free fatty acid content after exercise,the maximal effect was observed at 15mg/kg·bw the high-dose Capsaicin treated group and the control (P＜0.01)â‘¢There is no significant difference among each group before and after exercise. (3) central system:â‘ The open field test scores of the mice after sleep deprivation for 72h was significantly reduced compared with normal sleep group;caffeine and capsaicin significantly improved the open field test scores after sleep deprivation for 72h compared with the rats with normal sleep(P＜0.05 or P＜0.01 );the the maximal effect was observed at 15mg/kg·bw(74±9 vs 49±7,P＜0.01 ).â‘¡The content of hypothalamus and brain stem 5-HTand 5-HIAA after sleep deprivation for 72h was significantly increased compared with normal sleep group (14.38±1.37vs 12.76±1.25,P＜0.01);caffeine and capsaicin significantly reduced the content of hypothalamus and brain stem 5-HTand 5-HIAA after sleep deprivation for 72h compared with the normal sleep group(P＜0.05 or P＜0.01 );and the the maximal effect was observed at 15mg/kg·bw(11.19±1.26vs 14.38±1.37,P＜0.01 ) and there is no significant difference compared with caffeine treated group(P＞0.05).â‘¢The content of Ad,NE and DA in the brain stem was significantly reduced after sleep deprivation for 72h compared with normal sleep group(P＜0.01);Caffeine and Capsaicin significantly decreased the the content of Ad,NE and DA in the brain stem(P＜0.05 or P＜0.01 );and the the maximal effect was observed at 15mg/kg·bw and there is no significant difference compared with caffeine treated group(P＞0.05).â‘£There is no significant difference in the content of blood NH₃ among each group before sleep deprivation.The content of blood NH₃ after sleep deprivation for 72h was significantly increased compared with normal sleep group;Caffeine and Capsaicin significantly decreased the content of blood NH₃ after sleep deprivation for 72h compared with the rats with normal sleep(P＜0.05 or P＜0.01 );and the the maximal effect was observed at 15mg/kg·bw(158.5±27.9vs 318.6±34.8,P＜0.01 ) and there is no significant difference compared with caffeine treated group(P＞0.05).CONCLUSIONS:Capsaicin has significant antifatigue effect and the mechanism may be related toâ‘ increase the antioxidase activity, eliminate free radicals,and reduce the damage of cardic muscle;â‘¡promote the metabolism of fatty acids,reduce glycogen consumption and maintain the level of blood glucose after exercise;â‘¢decrease the generation of 5-HT and blood NH₃,and simultaneously increase the content of NA,Ad and DA,thus maintain the ratio of DA/5-HT at a high level to adjust the central activities.Partâ…£.Effect of capsaicin on I/R-induced apoptotic death of primary rat hippocampal neurons and the mechanismsAIM:To observe the effect of capsaicin on I/R-induced apoptotic death of primary rat hippocampal neurons and to explore its underlying mechanisms.METHODS:Primary cultures of rat hippocampal neurons were prepared from day 18 Sprague-Dawley rat embryos,and all experiments were performed on cells cultured for 8-10 days.Neuronal damage was induced by three-hour hypoxia(1%O₂) and consequent 24-hour reoxygenation.Apoptosis was evaluated by in situ terminal uridine nick end labeling(TUNEL) analysis and caspase-3 activity.Akt activation was characterized by western bloting with antibodies to phosphorylated Akt.Intracellular oxidant productions were measured using H₂DCF,a fluorescent indicator.RESULTS:(1)At 8th day of in vitro culture,embryonic rat hippocampus grown were identified as 94.5±2.1%microtubule-associated protein 2-positive cells,suggesting these cells contained mainly neurons.(2)H/R significantly increased the apoptotic death of hippocampal neurons as shown by increases in both TUNEL-positive cell number and caspase-3 activity.Pretreatment with capsaicin(3-30μM) or caspase-3 specific inhibitor DEVD-CHO(100μM) could markedly attenuate H/R-induced apoptosis in hippocampal neurons.(3)Capsaicin markedly induced the phosphorylation of Akt and PI3K inhibitor LY294002(10μM) prevented the capsaicin-induced survival effect in hippocampal neurons.(4)Intracellular reactive oxygen species(ROS) levels were significantly increased after H/R,which was concentration-dependent inhibited by capsaicin.The effects of capsaicin on intracellular ROS levels and apoptosis were blocked by LY294002(10μM).(5)VR1 antagonist capsazepine(1μM) or intracellular Ca²⁺ chelator BAPTA/AM(5μM) could eliminate these effects of capsaicin mentioned above.CONCLUSION:The present data suggest that capsaicin protects against H/R-induced apoptosis of hippocampal neurons via VR1-[Ca²⁺]_i-dependent and PI3K/Akt-mediated signaling pathway, which is related to inhibiting oxidative stress and caspase-3 activation.