社会威胁安全测试揭示了与社会心理压力相关的表型
Summary
社会威胁安全测试允许同时评估社会回避发展，作为厌恶条件学习和社会威胁安全辨别能力的测量，两者都用于在一组长期社会失败的雄性小鼠中识别压力易感和压力弹性个体。
Abstract
社会压力是精神障碍发展的主要原因。为了提高临床前研究的转化价值，社会压力体验及其对小鼠的行为影响应与人类相当。慢性社交失败 （CSD） 利用一种涉及身体攻击和感官威胁的社会压力来诱发类似于人类情感障碍的精神功能障碍。为了加强 CSD 的社会心理成分，应用了为期 10 天的 CSD 方案，其中每日物理攻击被标准化为三次 10 秒发作，然后是 24 小时的感觉阶段。在第 10个 感觉阶段之后，CSD 协议之后是称为社会威胁安全测试 （STST） 的精细行为测定。应激后行为测定需要确定社会压力源如何以及在多大程度上影响了行为。STST允许长期被社会击败的雄性小鼠与2个新的雄性个体（社会目标）相互作用：一个来自CSD期间遇到的攻击菌株的社会目标，另一个来自新菌株。两者同时呈现在三室测试场的不同隔间中。该测试能够同时评估社会回避的发展，以衡量成功的厌恶条件学习和社会威胁安全辨别能力。对这两种菌株的社会回避的发展反映了一种普遍的厌恶反应，因此也反映了压力易感性的测量。同时，社会回避的发展仅反映了威胁-安全歧视，从而反映了压力弹性的衡量标准。最后，缺乏对攻击压力的社会回避反映了厌恶条件学习受损。该协议旨在通过包括转化标准，特别是威胁安全歧视和厌恶反应泛化，来完善目前使用的压力易感性/复原力小鼠模型，将一组长期被社会击败的动物分类为复原力和易感亚群，最终推进未来的转化方法。
Introduction
压力被定义为由身体或心理刺激引起的体内平衡破坏1.众所周知，压力是导致创伤后应激障碍、抑郁和焦虑等精神障碍发展的主要危险因素 2,3。特别是，社会压力被认为是与压力相关的精神障碍发展的主要危险因素4.一种在研究中变得特别重要的社会压力是社会从属压力5.小鼠和人类一样，具有丰富的社会行为6，使它们适合于涉及社会压力的调查。在实验室环境中，当成年小鼠被群体饲养时，它们建立了一个社会结构，涉及形成7级。因此，设计了菌落模型来研究自然建立的社会等级制度对混合性别小鼠群体的影响8。多年来，已经开发了群体模型的变体来利用社会从属压力，包括同性群体模型、社会不稳定模型和入侵者-群体模型。然而，近年来，一种被称为男性居民入侵者模型的特殊变体在文献中流行起来，将社会复杂性简化为两只小鼠：居民和入侵者。感兴趣的动物，称为入侵者，被放入一个更大、更老、更退休的饲养员的笼子里，称为居民或侵略者。然后，居民对入侵者进行人身攻击，作为一种对抗方法，建立一种社会等级制度，其中居民占主导地位，入侵者处于从属地位。当对抗是一次性事件时，它们被归类为“急性”（“急性社会失败模型”），而持续数天（通常为 10 天）的重复对抗被称为“慢性”（“慢性社会失败模型”）。在慢性社交失败 （CSD） 模型中，发作是间歇性的，通常局限于 5-10 分钟9 的时间段，称为物理阶段。在物理阶段之后，入侵者和居民被关在同一个笼子里过夜，用网状墙隔成两半，允许除物理接触以外的所有形式的互动。这种配置被称为感觉阶段，通过威胁的持续出现而不是直接的身体对抗来诱发压力。2018 年，van der Kooji 及其同事引入了一种改良的慢性社会失败治疗，通过标准化和严格限制物理阶段10 来关注模型的社会心理成分。修改后的模型将物理攻击限制为三个 10 秒的发作，与不同的居民发生，发生在感觉阶段的 15 分钟发作间隔内。在第三次身体发作之后，感觉阶段持续一夜。这个循环连续重复 10 天，每集都有新居民。改进后的治疗增强了慢性社会失败模型的转化效度，因为入侵者的身体伤害被最小化，并且减少了不同物理攻击持续时间的结果变异性。
由于 CSD 模型用于研究与压力相关的疾病（例如，抑郁症、焦虑症、创伤后应激障碍），因此选择行为后检测，包括但不限于攻击性、记忆力和快感缺乏的行为检测。近年来，小鼠的 CSD 后行为测定经常评估社交能力如何以及在多大程度上受到影响9.社交能力被定义为小鼠对社会互动的天生偏好，而不是在社会上避免同种。由于社交能力会受到压力影响，因此建立了仅评估社交回避发展的测定法。压力引起的社交回避具有转化相关性，因为它代表了人类社交焦虑和抑郁的主要行为症状之一11.与人类类似，并非所有小鼠在CSD治疗后都会出现社交回避，这表明压力反应中存在个体性。Cohen 及其同事提出了截止行为标准，认为这是研究个体性神经生物学的一种有前途的方法12.基于行为的动物选择导致群体划分，强调了基因环境研究的基础。随后，不同的亚组通常表现出特定遗传变异/修饰的不同富集，这反过来又可以在不同的环境条件下进行研究13.因此，利用社会回避发展中的个体性将长期社会失败的雄性小鼠分为两个亚组：压力易感（社交回避）和压力弹性（社交非回避9,14）。然而，应在治疗（此处为CSD）和治疗后行为测定的整体背景下考虑将小鼠的社会回避表型解释为适应不良或适应性行为。此外，理想情况下，选择的治疗后行为测定将评估社交能力的其他方面，而不仅仅是社交回避的发展。我们最近的工作揭示了条件学习在 CSD 引起的社交回避中的参与15。具体来说，CSD诱发的社会回避是对居民应变的特征的厌恶性条件反应，作为对无条件刺激的条件刺激，即居民的攻击。此外，在社会回避型亚组中，一些个体可以区分厌恶型居民菌株的特征和其他安全新型菌株的特征，而另一些个体则对这两种菌株表现出普遍的社会回避性。我们在这里提出了一种改进的 CSD 后行为检测：社会威胁安全测试 （STST）15。与其他社会互动测试9不同，STST可以同时评估社会回避发展，作为正确厌恶条件反应（即成功的条件学习）和社会威胁安全辨别能力的测量，这两者都用于识别压力易感和压力弹性个体在一组长期社会失败的雄性小鼠中。社会威胁-安全歧视与厌恶反应泛化的评估扩展了用于将长期被社会击败的单一动物群体分类为弹性和易感亚群的转化标准。
Protocol
Representative Results
Discussion
这里的行为方案描述了社会威胁-安全测试，用于将 CSD 治疗后的一组分为三个不同的亚组，作为一种研究压力易感性和弹性的潜在生物学以及测试潜在疗法的方法。需要仔细考虑生物学背景和技术细节，以指导彻底的实验设计。
不同的住房条件会改变攻击性社交水平，从而可能影响从治疗后测试中获得的结果28,29,<sup class="x…
Divulgations
The authors have nothing to disclose.
Acknowledgements
这项研究得到了合作研究中心 1193 子项目 Z02 的支持，该子项目由德国国家研究基金会（SFB1193，弹性神经生物学）和勃林格殷格翰基金会（授予莱布尼茨弹性研究所和个体表型和高分辨率自动行为分析）。我们要感谢 Konstantin Radyushkin 博士和 Sandra Reichel 女士的技术援助，以及 Hanna Kim 女士的英语语言支持。资金来源没有参与模型设计;数据的收集、分析和解释;在协议的编写中;以及提交议定书供公布的决定。
Materials
| Arenas | Noldus, Sociability cage, Wageningen, the Netherlands | https://www.noldus.com/applications/sociability-cage | Three-chambered, rectangle in shape with a total size of 60 cm x 40 cm, made of acrylic transparent walls and smooth floors |
| Camera for video recording | Basler AG, Germany An der Strusbek 60-62 22926 Ahrensburg |
ace Classic acA1300-60gc |
If using automatic detection program, make sure cameras are compatible |
| Camera objective | KOWA Kowa Optimed Deutschland GmbH Fichtenstr. 123 40233 Duesseldorf: LMVZ4411 | 1/1.8" 4.4~11mm Varifokal Objektiv |
Part-No. 10504 | |
| Detection program/Timer | Noldus, EthoVision-XT, Wageningen, the Netherlands | https://www.noldus.com/ethovision-xt | Detection can be achieved either manually (using a timer or a software for manual scoring) or automatically |
| Housing cages | ZOONLAB GmbH, Hermannstraße 6, 44579 Castrop-Rauxel |
3010010 | Type 2 cages: 265 mm x 205 mm x 140 mm (l x w x h) i.e. 360 cm² bottom area. Made of Polycarbonate (Makrolone©) and Polysulfone. Lids are made of stainless steel. European standard cages for up to 5 mice (20–25 g). Autoclavable up to 134 °C |
| Mesh enclosures | Part of the Arena Package: Noldus, Sociability cage, Wageningen, the Netherlands | https://www.noldus.com/applications/sociability-cage | Small acrylic or metal cage-like with a diameter of 100 mm and a height of 200 mm with openings of a 10 mm in size. Two mesh enclosures per arena would work but four is preferable (see point 2.5 in protocol) |
| Mesh wall | selfmade | N/A | Acrylic or metal, one for each cage. Size depends on cages used. The walls must not allow the two animals to have a physical contact |
| Social targets: Mice of the strains CD-1 and 129/Sv; retired male breeders | Mice provided by Charles River: Strain name: CD-1®IGS Mouse 129S2/SvPasCrl |
Crl:CD1(ICR); 129S2/SvPasCrl | CD-1 and 129/Sv retired male breeders, single-housed, novel (unknown) conspecifics to the animals of interest. If retired male breeders are not available then males older than 1 year from both strains would suffice |
References
- Hyman, S. E. How mice cope with stressful social situations. Cell. 131 (2), 232-234 (2007).
- Kessler, R. C. The effects of stressful life events on depression. Annual Review of Psychology. 48 (1), 191-214 (1997).
- Vos, T., et al. Global, regional, and national incidence, prevalence, and years lived with disability for 301 acute and chronic diseases and injuries in 188 countries, 1990-2013: A systematic analysis for the Global Burden of Disease Study, 2013. The Lancet. 386 (9995), 743-800 (2015).
- Björkqvist, K. Social defeat as a stressor in humans. Physiology & Behavior. 73 (3), 435-442 (2001).
- Blanchard, R. J., McKittrick, C. R., Blanchard, D. C. Animal models of social stress: effects on behavior and brain neurochemical systems. Physiology & Behavior. 73 (3), 261-271 (2001).
- Singleton, G. R., Krebs, C. J. Chapter 3- The Secret World of Wild Mice. The Mouse in Biomedical Research. American College of Laboratory Animal Medicine. The Mouse in Biomedical Research (Second Edition). 1, 25-51 (2007).
- Kondrakiewicz, K., Kostecki, M., Szadzińska, W., Knapska, E. Ecological validity of social interaction tests in rats and mice. Genes, Brain, and Behavior. 18 (1), e12525 (2019).
- Martinez, M., Calvo-Torrent, A., Pico-Alfonso, M. A. Social defeat and subordination as models of social stress in laboratory rodents: a review. Aggressive Behavior: Official Journal of the International Society for Research on Aggression. 24 (4), 241-256 (1998).
- Golden, S. A., Covington III, H. E., Berton, O., Russo, S. J. A standardized protocol for repeated social defeat stress in mice. Nature Protocols. 6 (8), 1183 (2011).
- van der Kooij, M. A., et al. Chronic social stress-induced hyperglycemia in mice couples individual stress susceptibility to impaired spatial memory. Proceedings of the National Academy of Sciences of the United States of America. 115 (43), E10187-E10196 (2018).
- Chartier, M. J., Walker, J. R., Stein, M. B. Considering comorbidity in social phobia. Social Psychiatry and Psychiatric Epidemiology. 38 (12), 728-734 (2003).
- Cohen, H., Zohar, J., Matar, M. A., Kaplan, Z., Geva, A. B. Unsupervised fuzzy clustering analysis supports behavioral cutoff criteria in an animal model of post-traumatic stress disorder. Biological Psychiatry. 58 (8), 640-650 (2005).
- Scharf, S. H., Schmidt, M. V. Animal models of stress vulnerability and resilience in translational research. Current Psychiatry Reports. 14 (2), 159-165 (2012).
- Krishnan, V., et al. Molecular adaptations underlying susceptibility and resistance to social defeat in brain reward regions. Cell. 131 (2), 391-404 (2007).
- Ayash, S., Schmitt, U., Müller, M. B. Chronic social defeat-induced social avoidance as a proxy of stress resilience in mice involves conditioned learning. Journal of Psychiatric Research. 120, 64-71 (2020).
- Ayash, S., et al. Fear circuit-based neurobehavioural signatures mirror resilience to chronic social stress in mouse. Proceedings of the National Academy of Sciences of the United States of America. 120 (17), e2205576120 (2023).
- Duits, P., et al. Updated meta-analysis of classical fear conditioning in the anxiety disorders. Depression and Anxiety. 32 (4), 239-253 (2015).
- Coifman, K. G., Bonanno, G. A., Rafaeli, E. Affect dynamics, bereavement, and resilience to loss. Journal of Happiness Studies. 8 (3), 371-392 (2007).
- Waugh, C. E., Thompson, R. J., Gotlib, I. H. Flexible emotional responsiveness in trait resilience. Emotion. 11 (5), 1059 (2011).
- Bonanno, G. A. Loss, trauma, and human resilience: Have we underestimated the human capacity to thrive after extremely aversive events. American Psychologist. 59 (1), 20 (2004).
- Bonanno, G. A. Resilience in the face of potential trauma. Current Directions in Psychological Science. 14 (3), 135-138 (2005).
- Yehuda, R., Flory, J. D., Southwick, S., Charney, D. S. Developing an agenda for translational studies of resilience and vulnerability following trauma exposure. Annals of the New York Academy of Sciences. 1071 (1), 379-396 (2006).
- Grillon, C., Morgan III, C. A. Fear-potentiated startle conditioning to explicit and contextual cues in Gulf War veterans with post-traumatic stress disorder. Journal of Abnormal Psychology. 108 (1), 134 (1999).
- Brewin, C. R. A cognitive neuroscience account of post-traumatic stress disorder and its treatment. Behaviour Research and Therapy. 39 (4), 373-393 (2001).
- Milad, M. R., Rauch, S. L., Pitman, R. K., Quirk, G. J. Fear extinction in rats: implications for human brain imaging and anxiety disorders. Biological Psychology. 73 (1), 61-71 (2006).
- Jovanovic, T., Norrholm, S. D. Neural mechanisms of impaired fear inhibition in post-traumatic stress disorder. Frontiers in Behavioral Neuroscience. 5, 44 (2011).
- Morton, D. B., Griffiths, P. H. Guidelines on the recognition of pain, distress and discomfort in experimental animals and an hypothesis for assessment. The Veterinary Record. 116 (6), 431-436 (1985).
- Goldsmith, J. F., Brain, P. F., Benton, D. Effects of the duration of individual or group housing on behavioural and adrenocortical reactivity in male mice. Physiology & Behavior. 21 (5), 757-760 (1978).
- Cairns, R. B., Hood, K. E., Midlam, J. On fighting in mice: Is there a sensitive period for isolation effects. Animal Behaviour. 33 (1), 166-180 (1985).
- Varlinskaya, E. I., Spear, L. P., Spear, N. E. Social behavior and social motivation in adolescent rats: role of housing conditions and partner’s activity. Physiology & Behavior. 67 (4), 475-482 (1999).
- Sial, O. K., Warren, B. L., Alcantara, L. F., Parise, E. M., Bolaños-Guzmán, C. A. Vicarious social defeat stress: Bridging the gap between physical and emotional stress. Journal of Neuroscience Methods. 258, 94-103 (2016).
- Brown, R. E., Brown, R. E., Macdonald, D. W. . The rodents II. Suborder Myomorpha. [In: Social Odours in Mammals]. 1, (1985).
- Haney, M., Miczek, K. A. Ultrasounds during agonistic interactions between female rats (Rattus norvegicus). Journal of Comparative Psychology. 107 (4), 373 (1993).
- Warren, B. L., et al. Neurobiological sequelae of witnessing stressful events in adult mice. Biological Psychiatry. 73 (1), 7-14 (2013).
- Malatynska, E., Knapp, R. J. Dominant-submissive behavior as models of mania and depression. Neuroscience & Biobehavioral Reviews. 29 (4-5), 715-737 (2005).
- Avgustinovich, D. F., Kovalenko, I. L., Kudryavtseva, N. N. A model of anxious depression: persistence of behavioral pathology. Neuroscience and Behavioral Physiology. 35 (9), 917-924 (2005).
- Vennin, C., et al. A resilience related glial-neurovascular network is transcriptionally activated after chronic social defeat in male mice. Cells. 11 (21), 3405 (2022).
- Ayash, S., Schmitt, U., Lyons, D. M., Müller, M. B. Stress inoculation in mice induces global resilience. Translational Psychiatry. 10 (1), 200 (2020).
- Yuan, R., et al. Long-term effects of intermittent early life stress on primate prefrontal-subcortical functional connectivity. Neuropsychopharmacology. 46 (7), 1348-1356 (2021).
- Lyons, D. M., Ayash, S., Schatzberg, A. F., Müller, M. B. Ecological validity of social defeat stressors in mouse models of vulnerability and resilience. Neuroscience & Biobehavioral Reviews. 145, 105032 (2023).
- Oizumi, H., et al. Influence of aging on the behavioral phenotypes of C57BL/6J mice after social defeat. PLoS One. 14 (9), e0222076 (2019).
