We are grateful for the instrumentation and laboratory provided byextend our gratitude to the Mongolian medical faculty of the Inner Mongolian Medical University, China.This study was supported by, for providing the necessary instrumentation and laboratory facilities. This study received financial support from the National Natural Sciences Foundation of China (Grant No. 81760762) and), the Inner Mongolia Medical University Project of China (Grant No. YKD2022MS074), and the Scientific Research Project of Higher Education in Inner Mongolia, China (Grant No. NJZY22661) and the Open Fund Project of Key Laboratory of Chinese and Mongolian Medicine in Inner Mongolia Autonomous Region,China (Grant No. MYX2023-K07).

The experimental protocols received approval from the Ethics of Animal Experiment Care Committee of Inner Mongolia Medical University (YKD202301172) and adhered to the National Institutes of Health guidelines for animal care and ethics. The license number for our animal center is NO.110324230102364187. Twenty-four male Sprague-Dawley (SD) rats, each 8 weeks old (200 g &#177; 20 g), were acquired and housed in a controlled environment with a temperature of 22&#176;C &#177; 2&#176;C and humidity of 55% &#177; 15%. Feed the...

Establishing and Validating a Chronic Unpredictable Mild Stress Rat Model

Depression is a mental disorder characterized by symptoms such as low mood, lack of pleasure, and reduced energy30. In the field of depression research, the establishment of a reliable animal model is crucial for advancing therapeutic interventions. Among various animal models, the CUMS model is particularly noteworthy for its high reliability, validity, and its congruence with the characteristics of human depression31. It is well-suited to mimic the cumulative effects of l...

Major depressive disorder (MDD) is a prevalent mental illness, ranking as the third leading cause of disability globally and affecting over 300 million people1,2,3. Notably, it is estimated that at least half of the affected individuals do not receive adequate treatment4. Given this gap, animal models serve as a crucial tool for investigating the etiology of depression. To date, more than 20 different animal models for depression exist5. Among these, the chronic unpredictable mild stress (CUMS) model, refined by Paul Winer in 1987, is the most frequently utilized6. The CUMS model operates on the premise that exposing rodents to a diverse range of socio-environmental stressors leads to symptoms akin to anxiety, tension, and depression. The methodology involves exposing animals to various mild stressors over several weeks, culminating in a range of behavioral alterations, including anhedonia and depressive-like behaviors7,8. These changes are accompanied by shifts in endocrine and neurotransmitter profiles, such as a reduction in 5-HT9,10. These outcomes closely mirror those observed in humans diagnosed with MDD, thereby validating the model's utility. The CUMS model is particularly valued for its effectiveness in evaluating antidepressants, manifesting high levels of surface, structural, and predictive validity11,12. Unlike other models, CUMS is sensitive to the effects of chronic administration of monoaminergic antidepressants. For instance, selective serotonin reuptake inhibitors (SSRIs) such as citalopram, paroxetine, and fluoxetine have been shown to prevent and reverse anhedonia under conditions of chronic stress12,13. Additionally, new rapid-acting antidepressants, such as ketamine, have also demonstrated efficacy in this model14,15. In contrast, other tests like the forced swimming test (FST) and tail suspension test (TST) are less reliable for modeling long-term behavioral changes, often reflecting adaptations to acute stress rather than enduring symptoms of depression16. These characteristics underscore the robust validity of the CUMS model in depression research. One of the most salient features of the CUMS model, recognized for its high reliability in classical studies, is anhedonia-the incapacity to experience pleasure or interest in everyday activities17,18. This phenomenon is commonly evaluated using sucrose preference tests, and many antidepressants have been shown to reverse reduced sucrose consumption. Several other metrics are also commonly employed in the CUMS literature, including the open field test (OFT), which assesses voluntary motor behavior, exploratory tendencies, and tension, thereby gauging the severity of depression19. Other tests such as the elevated plus maze (EPM) evaluate anxiety-like behaviors, the Morris water maze test (MWM) examines cognitive functioning20, and the FST assesses susceptibility to negative emotions and behavioral despair20. Furthermore, the majority of stressors that impact humans are social in nature. Individuals with suboptimal social relationships, characterized by limited social activities, networks, and support, are at a heightened risk for various diseases21,22. This is also relevant in the case of rodents, who are social animals living in groups. For example, rats that are housed in isolation exhibit characteristics of what is termed isolation syndrome, which induces social stress and expedites the onset of depression23.
Mongolian medicine, a significant branch of Chinese medicine, posits that the onset of disease is a complex interplay between intrinsic and external factors. These external factors, referred to as the four auxiliary conditions, encompass climate change, diet, lifestyle, and sudden events such as infections, startling incidents, and psychological disorders. The disease process is conceptualized as an ongoing interaction between three elements-referred to as the three types of homors-and the Seven Bodily Constituents in concert with the four auxiliary conditions24. Mongolian medicine holds that the human body functions as an integrated entity, maintained by a relative balance among the three homors. A disruption in this balance is considered to be a precursor to illness24. Given the pivotal role of animal experimentation in bridging traditional and modern medicine, it is crucial to develop animal models that are relevant for research in the field of Mongolian medicine. Accordingly, we employed a 28-day isolation methodology coupled with CUMS to simulate these physiological and psychological stressors. We selected nine specific unpredictable stressors and sought to underpin this modeling method through the Three homors theory of Mongolian medicine. Establishing a robust animal model is fundamental for advancing basic research in Mongolian medicine and will contribute significantly to its foundational studies.

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chronic unpredictable mild stress in rats based on the mongolian medicine

Depression is a prevalent affective disorder and constitutes a leading cause of global disability. The limitations of current pharmacological interventions contribute to the substantial health burden attributed to this condition. There is a pressing need for a deeper understanding of the underlying mechanisms of depression, making pre-clinical models with translational potential highly valuable. Mongolian medicine, a subset of traditional medicine, posits that disease occurrence is closely tied to the equilibrium of wind, bile, and Phlegm. In this study, we introduce a protocol for the chronic unpredictable mild stress (CUMS) method in rats. Within this framework, rats are subjected to a series of fluctuating, mild stressors to induce a depression-like phenotype, mimicking the pathogenesis of human depression. Behavioral assays employed in this protocol include the sucrose preference test (SPT), indicative of anhedonia-a core symptom of depression; the open field test (OFT), which measures anxiety levels; and the Morris water maze test (MWM), which evaluates spatial memory and learning abilities. The CUMS method demonstrates the capability to induce anhedonia and to cause long-term behavioral deficits. Furthermore, this protocol is more aligned with Mongolian medical theory than other animal models designed to elicit depression-like behavior. The development of this animal model and subsequent research provide a robust foundation for future innovative studies in the realm of Mongolian medicine.

Author Spotlight: Establishing a Rodent Model for Investigating Depression Factors in Traditional Mongolian Medicine

Chronic Unpredictable Mild Stress in Rats based on the Mongolian medicine

The scope of our research is Mongolian medical research. In this study, we are investigating the factors contributing to depression in Mongolian medicine, with a particular emphasis on the interaction between internal factors, such as wind, biofilm, and external factors. The problem of conforming to the lack of traditional Mongolian medical models was solved.This protocol addresses the establishment of an animal model based on Mongolian medical theory and then provides some suggestions for future research in Mongolian medicine. In a depressive disorder study, establishing an animal model is crucial for understanding new drugs'pathological mechanisms and effects. Compared with other techniques, this protocol is more in line with the pathogenesis and treatment principles of depression in traditional Mongolian medicine, which will help future research on the treatment of depression in Mongolian medicine.The protocol can demonstrate the relationship between external and internal factors interacting to produce disease and is in accordance with the theoretical system of Mongolian medicine. It will advance the basic research of traditional medicine on depression and provide some recommendations that will help subsequent researchers to research, develop, and utilize traditional medicines.

Results from the behavioral tests in the CUMS-induced rat depression model 
To corroborate the efficacy of the CUMS procedure for inducing depression-like behaviors, a manipulation check was performed. Male Sprague-Dawley (SD) rats were randomly allocated to either the MOD or CON group for a period of 4 weeks, as outlined in step 1.2.3. Subsequently, the rats were sacrificed, and their hippocampi were completely dissected for assessment of 5-HT, a neurotransmitter strongly associated with depression...

Watch this Scientific Journal Video about Chronic Unpredictable Mild Stress in Rats based on the Mongolian medicine at JoVE.com

This protocol outlines a chronic unpredictable mild stress (CUMS) model for depression based on Mongolian medical theory, along with methods for validating behavioral tests.