压力增强恐惧学习--创伤后应激障碍的健壮啮齿动物模型
Summary
在这里, 我们描述了进行压力增强恐惧学习 (大我) 实验所需的详细方法, 这是在大鼠和小鼠创伤后应激障碍的临床前模型。该模型利用巴甫洛夫恐惧调理和冻结的各个方面作为对啮齿动物增强恐惧的指标。
Abstract
恐惧行为对于生存是重要的, 但过高的恐惧会增加患精神疾病的脆弱性, 如创伤后应激障碍 (PTSD)。要了解 PTSD 中恐惧失调的生物学机制, 首先必须从有效的疾病动物模型入手。本议定书描述了在大鼠和小鼠中进行压力增强恐惧学习 (大我) 实验的方法, 即 PTSD 的临床前模型。大我被开发来重述 PTSD 的关键方面, 包括对急性压力源引起的恐惧学习的长期敏感性。大我使用巴甫洛夫恐惧调理的一些方面, 但产生明显的和强健的敏感的恐惧反应远远大于正常条件恐惧反应。创伤程序包括将啮齿动物放置在调理室中, 并在90分钟内随机分布15非终止状态冲击 (用于大鼠实验; 对于小鼠实验, 使用10非终止状态随机分布60分钟以上的冲击).在2天, 啮齿类动物被放置在一个新的调理环境中, 他们收到一个单一的冲击;然后, 在3天, 它们被放回与2天相同的上下文中, 并测试了冻结级别的变化。与第一天没有受到冲击的人相比, 以前受到创伤的啮齿类动物在测试日的冻结水平有所提高。因此, 这种模式下, 单一的高度紧张的经验 (创伤) 产生极端恐惧与创伤事件相关的刺激。
Introduction
恐惧是生存的关键行为, 使个人能够识别和应对威胁。然而, 夸张的恐惧反应可能有助于精神障碍的发展, 如创伤后应激障碍 (PTSD)。PTSD 的一个特点是对轻度压力源的夸张反应, 特别是那些让人想起最初的创伤, 以及发展新恐惧的倾向1,2。在实验室里, 恐惧通常是通过冰冻行为来衡量的, 这是人类和啮齿类动物3、4的可靠和 ethologically 有效的恐惧指数。虽然众所周知, ptsd 涉及恐惧和增强恐惧表达的失调, 但缺乏强有力的创伤后应激障碍动物模型, 可靠地捕捉到相对无害刺激的这种增强恐惧反应。
本议定书提供了在大鼠和小鼠中进行压力增强恐惧学习 (大我) 实验的详细方法, 这是一种可靠和稳健的创伤后应激障碍临床前模型。大我利用巴甫洛夫恐惧调理的一些方面, 但它产生了不同的反应从正常恐惧调理和概括创伤后应激障碍患者的增强恐惧5,6。在这个模型中, 单一的高度紧张的经验 (称为创伤) 导致持久的行为改变, 包括与创伤事件相关的刺激的极端恐惧, 增加的焦虑, 增加惊吓反应性, 并改变糖皮质激素信号7,8。大我的主要特点是, 在不同的环境下接触创伤性压力源 (一系列非终止状态冲击) 后, 动物对轻度压力源 (例如, 单次休克) 在另一背景下表现出夸大的恐惧反应。重要的是, 大我效应不是由于从创伤语境到新的语境或增加的休克敏感性5的泛化。在我们的模型中, 我们有针对性地利用程序, 减少任何泛化到一个新的背景, 如独特的运输, 气味和网格地板模式。因此, 与正常的恐惧调理不同, 大我是一个非关联的过程, 导致一种新的恐惧学习, 这与与创伤经验不直接相关的环境线索不成比例地相关。广泛的工作表明, 一个90分钟的会话包含15个不可预知的冲击在大鼠 (或一个单一的60分钟的会话包含10个不可预知的冲击, 小鼠), 导致长期持续的警觉性恐惧调理随着焦虑和失调基底皮质酮的昼夜节律。相比之下, 对单个电击的预曝光不会产生大我9。此外, 大我可在大鼠和小鼠中可靠地使用。
因此, ptsd 的大我模型是一个强大的工具, 用于探讨创伤后应激障碍病理生理学的生物学机制。使用大我, 研究人员可以研究如何暴露于创伤会影响未来的恐惧学习。此外, 此模型可用于调查可能涉及的特定细胞和分子机制, 以调节在 PTSD 中观察到的增强的恐惧表达。
Protocol
Representative Results
Discussion
大我是一种强大的创伤后应激障碍行为模型, 可在大鼠和小鼠中概括, 可用于研究创伤后应激障碍的致敏性恐惧反应。在创伤后的压力下, 啮齿动物在截然不同的语境中表现出增加的恐惧反应, 只有在这种情况下, 与温和的压力源配对, 才能提醒人们以前的创伤经历。在创伤性压力下, 啮齿类动物在2天返回创伤性压力语境时毫不意外地表现出高水平的恐惧感, 这表明创伤性压力的记忆是完整的 (<strong c…
Declarações
The authors have nothing to disclose.
Acknowledgements
这项工作由国家卫生研究院 R01AA026530 (msf)、珠联璧合脑和行为健康中心 (msf)、NRSA-F32 MH10721201A1 和 NARSAD 26612 (AKR) 和 NSF DGE-1650604 (SG) 资助。
Materials
| Fear Conditioning Chamber for Low Profile Floors | Med Associates Inc. | VFC-008-LP | Fear conditioning chamber |
| Sound Attenuating Cubicle | Med Associates Inc. | NIR-022SD | Sound-attentuaing cubicle to prevent intrusion of outside noise |
| NIR/White Light Control Box | Med Associates Inc. | NIR-100VR | Light control box capable of delivering white and near-infrared light |
| NIR VFC Light Box | Med Associates Inc. | NIR-100L2 | White overhead houselight |
| Windex Original Glass Cleaner | Windex | Solution for cleaning and scenting fear conditioning chambers between animals | |
| Acetic acid | Fisher Scientific | A38-212 | Solution for cleaning and scenting fear conditioning chambers between animals |
| A-Frame Chamber Insert | Med Associates Inc. | ENV-008-IRT | Black Plexiglas triangular insert to differentiate internal layout of Contexts A and B |
| Curved Wall Insert | Med Associates Inc. | VFC-008-CWI | White plastic sheet to differentiate internal layout of Contexts A and B |
| Low Profile Contextual Grid Floor with 1/8" Grid Rods for Mouse | Med Associates Inc. | VFC-005A | Flat grid floor for mice |
| Low Profile Contextual Grid Floor with Alternating 1/8" & 3/16" Grid Rods Mouse | Med Associates Inc. | VFC-005-S | Staggered grid floor for mice |
| Low Profile Contextual Grid Floor with 1/8" Staggered Grid Rods for Mouse | Med Associates Inc. | VFC-005A-L | Alternating grid floor for mice |
| Low Profile Contextual Grid Floor with 3/16" Grid Rods for Rat | Med Associates Inc. | VFC-005 | Flat grid floor for rats |
| Low Profile Contextual Grid Floor with Alternating 3/16" & 3/8" Grid Rods | Med Associates Inc. | VFC-005-L | Alternating grid floor for rats |
| Low Profile Contextual Grid Floor with 3/16" Staggered Grid Rods for Rat | Med Associates Inc. | VFC-005-S | Staggered grid floor for rats |
| Metal pans | Med Associates Inc. | Metal pans to catch droppings underneath grid floors | |
| Standalone Aversive Stimulator/Scrambler | Med Associates Inc. | ENV-414S | Shock generator and scrambler for footshock delivery |
| Multimeter | Fluke | 87-5 | Tool for measuring footshock amplitude |
| VideoFreeze Software | Med Associates Inc. | SOF-843 | VideoFreeze software for controlling shock delivery |
| High Speed Firewire Monochrome Video Camera | Med Associates Inc. | VID-CAM-MONO-4 | Video camera capable of recording in near-infrared light |
Referências
- Bremner, J. D., Krystal, J. H., Southwick, S. M., Charney, D. S. Functional neuroanatomical correlates of the effects of stress on memory. Journal of Traumatic Stress. 8 (4), 527-553 (1995).
- Dykman, R. A., Ackerman, P. T., Newton, J. E. Posttraumatic stress disorder: a sensitization reaction. Integrative Physiological and Behavioral Science. 32 (1), 9-18 (1997).
- Fanselow, M. S., Bolles, R. C. Naloxone and shock-elicited freezing in the rat. Journal of Comparative and Physiological Psychology. 93 (4), 736-744 (1979).
- Fanselow, M. S. What is Conditioned Fear?. Trends in Neurosciences. 7, 460-462 (1984).
- Rau, V., DeCola, J. P., Fanselow, M. S. Stress-induced enhancement of fear learning: an animal model of posttraumatic stress disorder. Neuroscience and Biobehavioral Reviews. 29 (8), 1207-1223 (2005).
- Perusini, J. N., Fanselow, M. S. Neurobehavioral perspectives on the distinction between fear and anxiety. Learning and Memory. 22 (9), 417-425 (2015).
- Poulos, A. M., et al. Sensitization of fear learning to mild unconditional stimuli in male and female rats. Behavioral Neuroscience. 129 (1), 62-67 (2015).
- Perusini, J. N., et al. Induction and Expression of Fear Sensitization Caused by Acute Traumatic Stress. Neuropsychopharmacology. 41 (1), 45-57 (2016).
- Rau, V., Fanselow, M. S. Exposure to a stressor produces a long lasting enhancement of fear learning in rats. Stress. 12 (2), 125-133 (2009).
- Poulos, A. M., et al. Amnesia for early life stress does not preclude the adult development of posttraumatic stress disorder symptoms in rats. Biological Psychiatry. 76 (4), 306-314 (2014).
- Fanselow, M. S., Sigmundi, R. A. Species-specific danger signals, endogenous opioid analgesia, and defensive behavior. Journal of Experimental Psychology: Animal Behavior Processes. 12 (3), 301-309 (1986).
- Jacobs, N. S., Cushman, J. D., Fanselow, M. S. The accurate measurement of fear memory in Pavlovian conditioning: Resolving the baseline issue. Journal of Neuroscience Methods. 190 (2), 235-239 (2010).
- Anagnostaras, S. G., et al. Automated assessment of pavlovian conditioned freezing and shock reactivity in mice using the video freeze system. Frontiers in Behavioral Neuroscience. , (2010).
- Long, V. A., Fanselow, M. S. Stress-enhanced fear learning in rats is resistant to the effects of immediate massed extinction. Stress. 15 (6), 627-636 (2012).
- Craske, M. G., et al. Optimizing inhibitory learning during exposure therapy. Behaviour Research and Therapy. 46 (1), 5-27 (2008).
- Paylor, R., Tracy, R., Wehner, J., Rudy, J. W. DBA/2 and C57BL/6 mice differ in contextual fear but not auditory fear conditioning. Behavioral Neuroscience. 108 (4), 810-817 (1994).
- Graham, L. K., et al. Strain and sex differences in fear conditioning: 22 kHz ultrasonic vocalizations and freezing in rats. Pyschology and Neuroscience. 2 (2), 219-225 (2009).
