神经肽Y系统在高剂量咖啡因、可可碱对大鼠焦虑行为影响的作用机制
摘要
目的：观察高剂量咖啡因、可可碱对大鼠焦虑行为的影响,探讨神经肽Y系统在其中具体的作用机制。
方法：成年雄性Wistar大鼠30只,体重280±20g。随机数字表法将大鼠分为5组,每组6只。5组分别为生理盐水对照组、咖啡因30mg/kg组、咖啡因60mg/kg组、可可碱30mg/kg组、可可碱60mg/kg组,给药方式均为灌胃给药。给药后30min进行高架十字迷宫测试大鼠焦虑行为,分别观察各组药物对大鼠焦虑行为的影响；用荧光定量PCR技术检验大鼠海马神经肽Y、Y1受体、Y2受体mRNA的表达水平,并通过与对照组比较,判断神经肽Y系统在高剂量咖啡因、可可碱对大鼠焦虑行为的影响。
结果：1.大鼠运动活性结果：摄入60mg/kg咖啡因可使动物的运动活性明显下降(F=21.774,P=0.001),而30mg/kg可可碱则可明显升高(F=6.869,P=0.026)。摄入60mg/kg咖啡因较30mg/kg咖啡因的动物运动活性明显下降(F=15.168,P=0.003),而60mg/kg可可碱较30mg/kg可可碱也明显下降(F=12.331,P=0.006)。摄入30mg/kg咖啡因(F=1.244,P=0.291)与60mg/kg可可碱(F=1.553,P=0.241)对动物运动活性无显著影响,且两组药物对动物运动活性影响相当(F=0.023,P=0.882)
2.大鼠焦虑行为结果：摄入60mg/kg咖啡因可使动物开臂次数%(F=5.336,P=0.044)及开臂时间%(F=7.750,P=0.019)明显均下降。摄入60mg/kg可可碱可使动物开臂次数%(F=5.022,P=0.049)明显下降。60mg/kg咖啡因组较30mg/kg咖啡因组开臂次数%明显减少(F=7.569, P=0.020)。60mg/kg可可碱组较30mg/kg可可碱组无论是开臂次数%(F=24.587,P=0.001)及开臂时间%(F=17.065,P=0.002)均显著减少。
3.与空白对照组相比(NPY/GAPDH值),摄入60mg/kg咖啡因动物NPY明显升高(F=13.961, P=0.004),30mg/kg可可碱NPY明显降低(F=7.597,P=0.020)。摄入60mg/kg咖啡因动物NPY明显较30mg/kg咖啡因高CA_30(F=7.169,P=0.023)。摄入30mg/kg咖啡因(F=32.963,P=0.000)及30mg/kg可可碱动物(F=10.541,P=0.009)Y1 mRNA明显升高。摄入60mg/kg可可碱动物Y1 mRNA较空白组有增高趋势(F=4.227,P=0.067)。摄入30mg/kg可可碱动物Y2 mRNA较空白组增高(F=4.532,P=0.050)。
结论：
本论文阐述了高剂量咖啡因及可可碱对大鼠焦虑行为的影响,并从分子生物学的层面解释了其具体的分子机制。
过往研究中发现高剂量咖啡因具有致焦虑作用,但其具体作用机制尚未完全揭示；而高剂量可可碱对焦虑的具体作用仍未见报道。本研究首先采用了动物焦虑研究广泛应用的高架十字迷宫(EPM)模型,进行了动物行为学的研究,并详细阐述了两种物质分别对大鼠焦虑行为的具体作用。其后,采用目前较先进的荧光定量PCR技术精确检测海马区NPY、Y1、Y2mRNA表达水平,并发现咖啡因、可可碱在不同焦虑行为影响作用下分子生物学层面的差异,首次阐述了咖啡因、可可碱对NPY系统的具体作用。
本研究发现：
高剂量咖啡因致大鼠焦虑,且具有量效关系；并可能导致动物运动活性降低。分子生物学层面发现,其可导致NPY及Y1mRNA表达明显升高,且具有量效关系；对Y2mRNA没有影响。
高剂量可可碱可产生抗焦虑作用,且可能导致动物运动活性增高；但过高剂量则产生致焦虑作用。分子生物学层面发现,高剂量可可碱可导致NPY mRNA表达明显降低,且通过上调Y2 mRNA水平达到该作用；过高剂量可可碱则导致NPY及Y1mRNA表达明显升高,对Y2mRNA没有影响。
综上所述,咖啡因、可可碱是人们生活中常食用的物质,其对焦虑的作用及其具体机制的研究具有重大的意义,然而目前该领域的研究仍有待深入。本研究对高剂量咖啡因、可可碱影响机体焦虑的作用有了更深刻的认识,并探讨了其具体的作用机制。结果有望对咖啡因、可可碱的食用习惯、NPY靶点抗焦虑药物的开发等起到一定的指导作用。
Object:We designed this experiment to observe the effects of high dose Caffeine and Theobromine on the anxiety behavior in rats, and to explore the molecular mechanism of neuropeptide Y (NPY) in the potential effects.
Methods:A total of 30 male Wistar rats, with body weight range from 260 to 300 g,were randomizedly divided into 5 groups on average. The 5 groups were as follows:NS (control), 30mg/kg Caffeine,60mg/kg Caffeine,30mg/kg Theobromine,60mg/kg Theobromine,. The agentia were intragastric administration. We used the elevated plus-maze(EPM) to test the anxiety behavior of rats 30mins after agentia administrated. To observe the effect of NS, Caffeine and Theobromine on the anxiety behavior. And we measured the level of NPY, Y1 and Y2 mRNA through fluorescent quantitation PCR. The results are compared with control group and we develop the role of NPY system in the effects of high dose Caffeine and Theobromine on the anxiety behavior in rats.
Results:
1. Locomotor activity results:60mg/kg Caffeine administration decreased the locomotor activity in rats (F=21.774, P=0.001) while 30mg/kg Theobromine administration increase the locomotor activity in rats (F=6.869, P=0.026).60mg/kg Caffeine administration shows decreased locomotor activity compared with 30mg/kg Caffeine (F=15.168, P=0.003). 60mg/kg Theobromine administration shows decreased locomotor activity compared with 30mg/kg Theobromine (F=12.331, P=0.006). Each group of 30mg/kg Caffeine (F=1.244, P=0.291)and 60mg/kg Theobromine (F=1.553, P=0.241)administration shows no significant impact on locomotor activity and no significant difference were found between the two groups (F=0.023, P=0.882).
2. Anxiety behavior results in rats:The open arm entries %(F=5.336, P=0.044) and open arm time % (F=7.750, P=0.019) were decreased in 60mg/kg Caffeine administration group. The open arm entries% (F=5.022, P=0.049) are decreased in 60mg/kg Theobromine.60mg/kg Caffeine group shows significant less open arm entries% (F=7.569, P=0.020) than 30mg/kg Caffeine group. The open arm entries% (F=24.587, P=0.001) and open arm time % (F=17.065, P=0.002) of 60mg/kg Theobromine were lower than 30mg/kg Theobromine.
3. fluorescent quantitation PCR results shows that when compared with the control group (NPY/GAPDH), NPY mRNA level are increased in 60mg/kg Caffeine group (F=13.961, P=0.004) and are decreased in 30mg/kg Theobromine group (F=7.597, P=0.020). NPY mRNA level are higher in 60mg/kg Caffeine group than in 30mg/kg Caffeine group (F=7.169, P=0.023). Y1 mRNA level are increased in 30mg/kg Caffeine group (F=32.963, P=0.000) and in 30mg/kg Theobromine group (F=10.541, P=0.009). It shows increased tendency of Y1 mRNA level in 60mg/kg Theobromine group (F=4.227, P=0.067)and Y2 mRNA level in 30mg/kg Theobromine group (F=4.532, P=0.059).
Conclusion:
This paper described that the effects of high dose Caffeine and Theobromine on anxious behavior in rats. We also explained the specific mechanism of the effects mentioned above through molecular biological methods.
In the past, high dose caffeine was reported to have anxigenic effects. But the mechanism of it hasn't been discovered completely. Effects of high dose Theobromine on anxious behavior has not even been reported. Firstly, we studied the behavioral effects through EPM which was most widely used animal model for anxiety and we explained the specific effects of the two substance on anxious behavioral in rats. Secondly, we used the fluorescent quantitation PCR, an advanced molecular biological technology, to test the expression of NPY mRNA and Yl mRNA and Y2 mRNA and we found the molecular biological difference in Caffeine and Theobromine on different anxious effects. We explained the role of NPY system in high dose Caffeine and Theobromine on anxious behavior in rats first time.
We have found:
High dose Caffeine has anxigenic effects with dose-effects relationship. In addition, it decreases the locomotion activity of rats. We also found it can increase the expression of NPY mRNA and Yl mRNA with dose-effects relationship. But it has no effects on Y2 mRNA expression.
High dose Theobromine has anxiolytic effects, and it could increase the locomotion activity of rats. But too high dose Theobromine makes rats anxious. We also found high dose Theobromine can decrease the expression of NPY mNRA which mediated by increasing the expression of Y2 mRNA. But too high dose Theobromine increased the expression of NPY mRNA and Y1 mRNA, and has no effects on Y2 mRNA.
In summary, Caffeine and Theobromine are widely consumed in the world. The effects of them on anxious behavior and the specific mechanism of that are important. But the more researches are still needed. Our research has contributions to understand the effects of high dose Caffeine and Theobromine on anxious behavior and the specific mechanism of them. The results of this study are expected to guide the consuming of Caffeine and Theobromine, and the development of anxiolytic agents targeting on NPY system.
方法：成年雄性Wistar大鼠30只,体重280±20g。随机数字表法将大鼠分为5组,每组6只。5组分别为生理盐水对照组、咖啡因30mg/kg组、咖啡因60mg/kg组、可可碱30mg/kg组、可可碱60mg/kg组,给药方式均为灌胃给药。给药后30min进行高架十字迷宫测试大鼠焦虑行为,分别观察各组药物对大鼠焦虑行为的影响；用荧光定量PCR技术检验大鼠海马神经肽Y、Y1受体、Y2受体mRNA的表达水平,并通过与对照组比较,判断神经肽Y系统在高剂量咖啡因、可可碱对大鼠焦虑行为的影响。
结果：1.大鼠运动活性结果：摄入60mg/kg咖啡因可使动物的运动活性明显下降(F=21.774,P=0.001),而30mg/kg可可碱则可明显升高(F=6.869,P=0.026)。摄入60mg/kg咖啡因较30mg/kg咖啡因的动物运动活性明显下降(F=15.168,P=0.003),而60mg/kg可可碱较30mg/kg可可碱也明显下降(F=12.331,P=0.006)。摄入30mg/kg咖啡因(F=1.244,P=0.291)与60mg/kg可可碱(F=1.553,P=0.241)对动物运动活性无显著影响,且两组药物对动物运动活性影响相当(F=0.023,P=0.882)
2.大鼠焦虑行为结果：摄入60mg/kg咖啡因可使动物开臂次数%(F=5.336,P=0.044)及开臂时间%(F=7.750,P=0.019)明显均下降。摄入60mg/kg可可碱可使动物开臂次数%(F=5.022,P=0.049)明显下降。60mg/kg咖啡因组较30mg/kg咖啡因组开臂次数%明显减少(F=7.569, P=0.020)。60mg/kg可可碱组较30mg/kg可可碱组无论是开臂次数%(F=24.587,P=0.001)及开臂时间%(F=17.065,P=0.002)均显著减少。
3.与空白对照组相比(NPY/GAPDH值),摄入60mg/kg咖啡因动物NPY明显升高(F=13.961, P=0.004),30mg/kg可可碱NPY明显降低(F=7.597,P=0.020)。摄入60mg/kg咖啡因动物NPY明显较30mg/kg咖啡因高CA_30(F=7.169,P=0.023)。摄入30mg/kg咖啡因(F=32.963,P=0.000)及30mg/kg可可碱动物(F=10.541,P=0.009)Y1 mRNA明显升高。摄入60mg/kg可可碱动物Y1 mRNA较空白组有增高趋势(F=4.227,P=0.067)。摄入30mg/kg可可碱动物Y2 mRNA较空白组增高(F=4.532,P=0.050)。
结论：
本论文阐述了高剂量咖啡因及可可碱对大鼠焦虑行为的影响,并从分子生物学的层面解释了其具体的分子机制。
过往研究中发现高剂量咖啡因具有致焦虑作用,但其具体作用机制尚未完全揭示；而高剂量可可碱对焦虑的具体作用仍未见报道。本研究首先采用了动物焦虑研究广泛应用的高架十字迷宫(EPM)模型,进行了动物行为学的研究,并详细阐述了两种物质分别对大鼠焦虑行为的具体作用。其后,采用目前较先进的荧光定量PCR技术精确检测海马区NPY、Y1、Y2mRNA表达水平,并发现咖啡因、可可碱在不同焦虑行为影响作用下分子生物学层面的差异,首次阐述了咖啡因、可可碱对NPY系统的具体作用。
本研究发现：
高剂量咖啡因致大鼠焦虑,且具有量效关系；并可能导致动物运动活性降低。分子生物学层面发现,其可导致NPY及Y1mRNA表达明显升高,且具有量效关系；对Y2mRNA没有影响。
高剂量可可碱可产生抗焦虑作用,且可能导致动物运动活性增高；但过高剂量则产生致焦虑作用。分子生物学层面发现,高剂量可可碱可导致NPY mRNA表达明显降低,且通过上调Y2 mRNA水平达到该作用；过高剂量可可碱则导致NPY及Y1mRNA表达明显升高,对Y2mRNA没有影响。
综上所述,咖啡因、可可碱是人们生活中常食用的物质,其对焦虑的作用及其具体机制的研究具有重大的意义,然而目前该领域的研究仍有待深入。本研究对高剂量咖啡因、可可碱影响机体焦虑的作用有了更深刻的认识,并探讨了其具体的作用机制。结果有望对咖啡因、可可碱的食用习惯、NPY靶点抗焦虑药物的开发等起到一定的指导作用。
Object:We designed this experiment to observe the effects of high dose Caffeine and Theobromine on the anxiety behavior in rats, and to explore the molecular mechanism of neuropeptide Y (NPY) in the potential effects.
Methods:A total of 30 male Wistar rats, with body weight range from 260 to 300 g,were randomizedly divided into 5 groups on average. The 5 groups were as follows:NS (control), 30mg/kg Caffeine,60mg/kg Caffeine,30mg/kg Theobromine,60mg/kg Theobromine,. The agentia were intragastric administration. We used the elevated plus-maze(EPM) to test the anxiety behavior of rats 30mins after agentia administrated. To observe the effect of NS, Caffeine and Theobromine on the anxiety behavior. And we measured the level of NPY, Y1 and Y2 mRNA through fluorescent quantitation PCR. The results are compared with control group and we develop the role of NPY system in the effects of high dose Caffeine and Theobromine on the anxiety behavior in rats.
Results:
1. Locomotor activity results:60mg/kg Caffeine administration decreased the locomotor activity in rats (F=21.774, P=0.001) while 30mg/kg Theobromine administration increase the locomotor activity in rats (F=6.869, P=0.026).60mg/kg Caffeine administration shows decreased locomotor activity compared with 30mg/kg Caffeine (F=15.168, P=0.003). 60mg/kg Theobromine administration shows decreased locomotor activity compared with 30mg/kg Theobromine (F=12.331, P=0.006). Each group of 30mg/kg Caffeine (F=1.244, P=0.291)and 60mg/kg Theobromine (F=1.553, P=0.241)administration shows no significant impact on locomotor activity and no significant difference were found between the two groups (F=0.023, P=0.882).
2. Anxiety behavior results in rats:The open arm entries %(F=5.336, P=0.044) and open arm time % (F=7.750, P=0.019) were decreased in 60mg/kg Caffeine administration group. The open arm entries% (F=5.022, P=0.049) are decreased in 60mg/kg Theobromine.60mg/kg Caffeine group shows significant less open arm entries% (F=7.569, P=0.020) than 30mg/kg Caffeine group. The open arm entries% (F=24.587, P=0.001) and open arm time % (F=17.065, P=0.002) of 60mg/kg Theobromine were lower than 30mg/kg Theobromine.
3. fluorescent quantitation PCR results shows that when compared with the control group (NPY/GAPDH), NPY mRNA level are increased in 60mg/kg Caffeine group (F=13.961, P=0.004) and are decreased in 30mg/kg Theobromine group (F=7.597, P=0.020). NPY mRNA level are higher in 60mg/kg Caffeine group than in 30mg/kg Caffeine group (F=7.169, P=0.023). Y1 mRNA level are increased in 30mg/kg Caffeine group (F=32.963, P=0.000) and in 30mg/kg Theobromine group (F=10.541, P=0.009). It shows increased tendency of Y1 mRNA level in 60mg/kg Theobromine group (F=4.227, P=0.067)and Y2 mRNA level in 30mg/kg Theobromine group (F=4.532, P=0.059).
Conclusion:
This paper described that the effects of high dose Caffeine and Theobromine on anxious behavior in rats. We also explained the specific mechanism of the effects mentioned above through molecular biological methods.
In the past, high dose caffeine was reported to have anxigenic effects. But the mechanism of it hasn't been discovered completely. Effects of high dose Theobromine on anxious behavior has not even been reported. Firstly, we studied the behavioral effects through EPM which was most widely used animal model for anxiety and we explained the specific effects of the two substance on anxious behavioral in rats. Secondly, we used the fluorescent quantitation PCR, an advanced molecular biological technology, to test the expression of NPY mRNA and Yl mRNA and Y2 mRNA and we found the molecular biological difference in Caffeine and Theobromine on different anxious effects. We explained the role of NPY system in high dose Caffeine and Theobromine on anxious behavior in rats first time.
We have found:
High dose Caffeine has anxigenic effects with dose-effects relationship. In addition, it decreases the locomotion activity of rats. We also found it can increase the expression of NPY mRNA and Yl mRNA with dose-effects relationship. But it has no effects on Y2 mRNA expression.
High dose Theobromine has anxiolytic effects, and it could increase the locomotion activity of rats. But too high dose Theobromine makes rats anxious. We also found high dose Theobromine can decrease the expression of NPY mNRA which mediated by increasing the expression of Y2 mRNA. But too high dose Theobromine increased the expression of NPY mRNA and Y1 mRNA, and has no effects on Y2 mRNA.
In summary, Caffeine and Theobromine are widely consumed in the world. The effects of them on anxious behavior and the specific mechanism of that are important. But the more researches are still needed. Our research has contributions to understand the effects of high dose Caffeine and Theobromine on anxious behavior and the specific mechanism of them. The results of this study are expected to guide the consuming of Caffeine and Theobromine, and the development of anxiolytic agents targeting on NPY system.
引文
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[27]. Gehlert DR, Yang P, George C, Wang Y, Schober D, Gackenheimer S, Johnson D, Bea-vers LS, Gadski RA, Baez M. Cloning and characterization of rhesus monkey neuropep-tide Y receptor subtypes. Peptides.2001;22:343-350.
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normal human volunteers. Psychopharmacology 1989;98:81-88.
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[34]. Takashi Yamada. Anxiolytic effects of short-and long-term administration of cacao mass on rat elevated T-maze test. Journal of Nutritional Biochemistry,2009;20(12):948-55.
[35]. Arborelius L, Owens MJ, Plotsky PM, Nemeroff CB. The role of corticotrophin-releasing factor in depression and anxiety disorders. J Endocrinol 1999;160:1-12
[36]. Mckinney WT. Overview of the past contributions in animal models and their changing place in psychiatry. Sem Clin Psychiatry 2001;6:68-78.
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[38]. Kalueff AV. Problems of the Study of Stress-Related Behavior(in Rassian). KSF, Kiev, 1999;99p.
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125-133.
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[48]. Flint J. Animal models of anxiety and their molecular dissection. Sem Cell Devel Biol 2003;14:37-42.
[49]. Kalueff AV. Today and tomorrow of anxiety research. Stress Behav 2003;8:145-147.
[50]. King JA, Messenger T, Ferris CF. Seed finding in golden hamsters:a potential animal model for screening anxiolytic drugs. Neuropsychobiol 2002;45:150-155.
[51]. Newport DJ, Stowe ZN, Nemeroff CB. Parental depression:animal models of an adverse life event. Am J Psychiatry 2002;159:1265-1283.
[52]. Makarchuk NE, Kalueff AV. Olfaction and Behavior (in Russian). KSF, Kiev,2000;149.
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[56]. Kawachi I, Willett WC, Colditz GA, Stampfer MJ and Speizer FE. A prospective study of coffee drinking and suicide in women. Arch Intern Med 1996;156:521-525.
[57]. Pechlivanova D, Tchekalarova J, Nikolov R, Yakimova K. Dose-dependent effects of caffeine on behavior and thermoregulation in a chronic unpredictable stress model of depression in rats.Behav Brain Res.2010 Jun 19;209(2):205-11.
[58]. Daly JW. Mechanism of action of caffeine, in Caffeine, Coffee, and Health (Garattini S ed).1993;97-150, Raven Press, New York.
[59]. Ferre'S, Fuxe K, von Euler G, Johansson B and Fredholm BB. Adenosinedopamine interactions in the brain. Neuroscience 1992;51:501-512.
[60]. Jin S, Johansson B and Fredholm BB. Effects of adenosine A1 and A2 receptor
activation on electrically evoked dopamine and acetylcholine release from rat striatal slices. J Pharmacol Exp Ther 1993;267:801-808.
[61]. Fernstrom JD and Fernstro"m MH. Effects of caffeine on monoamine neurotransmitters in the central and peripheral nervous system, in Caffeine:Perspectives from Recent Research (Dews PB ed) 1984; 107-118, Springer-Verlag, Heidelberg.
[62]. Bickford PC, Fredholm BB, Dunwiddie TV and Freedman R. Inhibition of Purkinje cell firing by systemic administration of phenylisopropyl adenosine:effect of central noradrenaline depletion by DSP4. Life Sci 1985;37:289-297.
[63]. Fredholm BB and Jonzon B. Adenosine receptors:Agonists and antagonists, in Role of Adenosine in Cerebral Metabolism and Circulation (Stefanovich V and Okayayoz-Baklouti I eds) 1988; 17-46, VSP BV, Utrecht.
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[67]. Madsen TM, Greisen MH, Nielsen SM, Bolwig TG, Mikkelsen JD. Electroconvulsive stimuli enhance both neuropeptide Y receptor Y1 and Y2 messenger RNA expression and levels of binding in the rat hippocampus. Neuroscience 2000;98:33-39.
[68]. Husum H, Bolwig TG, Sanchez C, Mathe AA, Hansen SL. Levetiracetam prevents changes in levels of brain-derived neurotrophic factor and neuropeptide Y mRNA and of Y1-and Y5-like receptors in the hippocampus of rats undergoing amygdale kindling: implications for antiepileptogenic and mood-stabilizing properties. Epilepsy Behav 2004;5:204-215.
[69]. Dumont Y, Jacques D, Bouchard P, Quirion R. Species differences in the expression and distribution of the neuropeptide Y Y1, Y2, Y4, and Y5 receptors in rodents, guinea pig, and primates brains. J Comp Neurol 1998;402:372-384.
[70]. King PJ, Williams G, Doods H, Widdowson PS. Effect of a selective neuropeptide Y Y(2) receptor antagonist, BIIE0246 on neuropeptide Y release. Eur J Pharmacol 2000;396:R1-R3.
[71]. Caberlotto L, Hurd YL. Neuropeptide Y Y(1) and Y(2) receptor mRNA expression in the prefrontal cortex of psychiatric subjects. Relationship of Y(2) subtype to suicidal behavior. Neuropsychopharmacology 2001;25:91-97.
[72]. Wetherill L, Schuckit MA, Hesselbrock V, Xuei X, Liang T, Dick DM, Kramer J, Nurnberger JI Jr, Tischfield JA, Porjesz B, Edenberg HJ, Foroud T. Neuropeptide Y receptor genes are associated with alcohol dependence, alcohol withdrawal phenotypes, and cocaine dependence. Alcohol Clin Exp Res.2008 Dec-32(12):2031-40.
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[25]. Ants Kask, Jaanus Harro, Stephan Von Horsten, et al. The neurocircuitry and receptor sub-types mediating anxiolytic-like effects of neuropeptide Y. Nrueoscience nd Biobehavioral Re-views,26:259-283
[26]. King PJ, Williams G, Doods H, Widdowson PS. Effects of a selective neuropeptide Y Y2 receptor antagonist, BIIE0246 on neuropeptide Y release. Eur Pharmacol, 2000;396:R1-R3
[27]. Gehlert DR, Yang P, George C, Wang Y, Schober D, Gackenheimer S, Johnson D, Bea-vers LS, Gadski RA, Baez M. Cloning and characterization of rhesus monkey neuropep-tide Y receptor subtypes. Peptides.2001;22:343-350.
[28]. Greden JF. Anxiety or caffeinism:A diagnostic dilemma. Am J Psychiatry 1974; 131:1089-1092.
[29]. Stern KN, Chait LD and Johanson CE. Reinforcing and subjective effects of caffeine in
normal human volunteers. Psychopharmacology 1989;98:81-88.
[30]. Daly JW. Mechanism of action of caffeine, in Caffeine, Coffee, and Health (Garattini S ed) 1993;97-150, Raven Press, New York
[31]. Eaton WW and McLeod J. Consumption of coffee or tea and symptoms of anxiety. Am J Public Health 1984;74:66-68.
[32]. Lee MA, Cameron OG and Greden JF. Anxiety and caffeine consumption in people with anxiety disorders. Psychiatry Res 1985; 15:211-217.
[33]. Rihs M, Mu ller C and Baumann P. Caffeine consumption in hospitalized psychiatric patients. Eur Arch Psychiatry Clin Neurosci 1996;246:83-92.
[34]. Takashi Yamada. Anxiolytic effects of short-and long-term administration of cacao mass on rat elevated T-maze test. Journal of Nutritional Biochemistry,2009;20(12):948-55.
[35]. Arborelius L, Owens MJ, Plotsky PM, Nemeroff CB. The role of corticotrophin-releasing factor in depression and anxiety disorders. J Endocrinol 1999;160:1-12
[36]. Mckinney WT. Overview of the past contributions in animal models and their changing place in psychiatry. Sem Clin Psychiatry 2001;6:68-78.
[37]. Allan VK, Pentti T. Experiental modeling of anxiety and depression. Acta Neurobiol Exp 2004;64:439-448.
[38]. Kalueff AV. Problems of the Study of Stress-Related Behavior(in Rassian). KSF, Kiev, 1999;99p.
[39]. Kalueff AV, Makarchuk NE, Samohvalov VP, Deryagina MA. Urination and Behavior (in Russian). KSF, Kiev,2001;134 p.
[40]. Aguilar R, Gil L, Flint J, Gray JA, Dawson GR, Driscoll P, Gimenez-Llort L, Escorihuela RM, Fernandez-Teruel A, Tobena A. Learned fear, emotional reactivity and fear of heights:a factor analyticmap from a large F2 intercross of Roman rat strains. Brain Res Bull 2002;57:17-26.
[41]. Moyaho A, Eguibar JR, Diaz JL. Induced grooming transitions and open field behavior differ in high-and low-yawning sublines of Sprague-Dawley rats. Animal Behav 1995;50: 61-72.
[42]. Lapin IP. Models of anxiety in mice:experimental verification and critical methodology. Exp Clin Pharm 2000;63:58-62.
[43]. Andrade MM, Tome MF, Santiago ES, Lucia-Santos A, de Andrade TG. Longitudinal study of daily variation of rats'behavior in the elevated plus maze. Physiol Behav 2003;78:
125-133.
[44]. Kalueff AV. Grooming and Stress (in Russian). Avix, Moscow,2002; 148 p.
[45]. BelzungC. Measuring rodent exploratory behavior. In:Handbook of Molecular Genetics for Brain and Behavior Research (Eds. W.E. Cruzio and T.T. Gerlai). Elsevier, New York, 1999;p.77-99.
[46]. Chapillon P, Manneche C, Belzung C, Caston J. Rearing environment enrichment in two inbred strains of mice:1. Effect of emotional reactivity. Behav Genet 1999;29:41-46.
[47]. Chen SW, Xin Q, Kong WX, Min L, Li JF. Anxiolytic-like effect of succinic acid in mice. Life Sci 2003;73:3257-3264.
[48]. Flint J. Animal models of anxiety and their molecular dissection. Sem Cell Devel Biol 2003;14:37-42.
[49]. Kalueff AV. Today and tomorrow of anxiety research. Stress Behav 2003;8:145-147.
[50]. King JA, Messenger T, Ferris CF. Seed finding in golden hamsters:a potential animal model for screening anxiolytic drugs. Neuropsychobiol 2002;45:150-155.
[51]. Newport DJ, Stowe ZN, Nemeroff CB. Parental depression:animal models of an adverse life event. Am J Psychiatry 2002;159:1265-1283.
[52]. Makarchuk NE, Kalueff AV. Olfaction and Behavior (in Russian). KSF, Kiev,2000;149.
[53]. Pellow, S., Chopin, P.File, S. E. Validation of open:closedarm entries in an elevated plus2maze as a measure of anxiety in the rat J Neurosci Methods.1985;14 (3):149-167.
[54]. Arcelo, C.Bulletin, B. R. Evaluation of the elevated T-maze as an animal model of anxiety in the mouse. Jardim 1999;48 (4):407-411.
[55]. Carobrez, A. P.Bertoglio, L. J. Ethological and temporal analyses of anxiety-likebehavior:
[56]. Kawachi I, Willett WC, Colditz GA, Stampfer MJ and Speizer FE. A prospective study of coffee drinking and suicide in women. Arch Intern Med 1996;156:521-525.
[57]. Pechlivanova D, Tchekalarova J, Nikolov R, Yakimova K. Dose-dependent effects of caffeine on behavior and thermoregulation in a chronic unpredictable stress model of depression in rats.Behav Brain Res.2010 Jun 19;209(2):205-11.
[58]. Daly JW. Mechanism of action of caffeine, in Caffeine, Coffee, and Health (Garattini S ed).1993;97-150, Raven Press, New York.
[59]. Ferre'S, Fuxe K, von Euler G, Johansson B and Fredholm BB. Adenosinedopamine interactions in the brain. Neuroscience 1992;51:501-512.
[60]. Jin S, Johansson B and Fredholm BB. Effects of adenosine A1 and A2 receptor
activation on electrically evoked dopamine and acetylcholine release from rat striatal slices. J Pharmacol Exp Ther 1993;267:801-808.
[61]. Fernstrom JD and Fernstro"m MH. Effects of caffeine on monoamine neurotransmitters in the central and peripheral nervous system, in Caffeine:Perspectives from Recent Research (Dews PB ed) 1984; 107-118, Springer-Verlag, Heidelberg.
[62]. Bickford PC, Fredholm BB, Dunwiddie TV and Freedman R. Inhibition of Purkinje cell firing by systemic administration of phenylisopropyl adenosine:effect of central noradrenaline depletion by DSP4. Life Sci 1985;37:289-297.
[63]. Fredholm BB and Jonzon B. Adenosine receptors:Agonists and antagonists, in Role of Adenosine in Cerebral Metabolism and Circulation (Stefanovich V and Okayayoz-Baklouti I eds) 1988; 17-46, VSP BV, Utrecht.
[64]. Hadfield MG and Milio C. Caffeine and regional brain monoamine utilization in mice. Life Sci 1989;45:2637-2644.
[65]. Parker RM, Herzog H. Regional distribution of Y-receptor subtype mRNAs in rat brain. European J Neurosci 1999;11:1431-1448.
[66]. Larsen PJ, Sheikh SP, Jakobsen CR, Schwartz TW, Mikkelsen JD. Regional distribution of putative NPY Y1 receptors and neurons expressing Y1 mRNA in forebrain areas of the rat central nervous system. Eur J Neurosci 1993;5:1622-1637.
[67]. Madsen TM, Greisen MH, Nielsen SM, Bolwig TG, Mikkelsen JD. Electroconvulsive stimuli enhance both neuropeptide Y receptor Y1 and Y2 messenger RNA expression and levels of binding in the rat hippocampus. Neuroscience 2000;98:33-39.
[68]. Husum H, Bolwig TG, Sanchez C, Mathe AA, Hansen SL. Levetiracetam prevents changes in levels of brain-derived neurotrophic factor and neuropeptide Y mRNA and of Y1-and Y5-like receptors in the hippocampus of rats undergoing amygdale kindling: implications for antiepileptogenic and mood-stabilizing properties. Epilepsy Behav 2004;5:204-215.
[69]. Dumont Y, Jacques D, Bouchard P, Quirion R. Species differences in the expression and distribution of the neuropeptide Y Y1, Y2, Y4, and Y5 receptors in rodents, guinea pig, and primates brains. J Comp Neurol 1998;402:372-384.
[70]. King PJ, Williams G, Doods H, Widdowson PS. Effect of a selective neuropeptide Y Y(2) receptor antagonist, BIIE0246 on neuropeptide Y release. Eur J Pharmacol 2000;396:R1-R3.
[71]. Caberlotto L, Hurd YL. Neuropeptide Y Y(1) and Y(2) receptor mRNA expression in the prefrontal cortex of psychiatric subjects. Relationship of Y(2) subtype to suicidal behavior. Neuropsychopharmacology 2001;25:91-97.
[72]. Wetherill L, Schuckit MA, Hesselbrock V, Xuei X, Liang T, Dick DM, Kramer J, Nurnberger JI Jr, Tischfield JA, Porjesz B, Edenberg HJ, Foroud T. Neuropeptide Y receptor genes are associated with alcohol dependence, alcohol withdrawal phenotypes, and cocaine dependence. Alcohol Clin Exp Res.2008 Dec-32(12):2031-40.