Research Background
Toad venom (Chansu), first documented in the "Yaoxing Lun" (Treatise on the Properties of Medicines), is a treasured traditional Chinese medicinal material. The "Bencao Gangmu" (Compendium of Materia Medica) records its efficacy in "treating boils, carbuncles, and all malignant swellings," with the effects of "opening orifices to awaken the spirit, detoxifying and reducing swelling." Zhang Jingye, a physician of the Ming Dynasty, noted in the "Jingyue Quanshu" that Chansu "excels at opening stagnation and breaking hard accumulations," emphasizing its unique role in promoting qi circulation and resolving stagnation. For millennia, Chansu has served as a core ingredient in classic formulas such as Liu Shen Wan and Meihua Dianshe Dan, demonstrating irreplaceable therapeutic effects in clinical practice.
Traditional Chinese medicine theory categorizes depression as "stagnation syndrome," closely associated with liver qi stagnation and heart-spirit malnutrition. The "Huangdi Neijing·Suwen" states "wood stagnation should be released" and "fire stagnation should be dispersed," emphasizing that treatment should focus on promoting circulation. Chansu, warm in nature and entering the heart and liver meridians, can "disperse wind-fire stagnation" and "open orifices to awaken the spirit," aligning seamlessly with modern antidepressant treatment concepts. However, the mechanisms of traditional Chinese medicines have long remained at the empirical level, constraining the modernization of Chinese medicine. Elucidating the molecular mechanisms underlying Chansu's medicinal properties represents a critical breakthrough for advancing the inheritance and innovation of traditional Chinese medicine.
Research Progress
Building on over a decade of research into the bioactive components of Chansu, the team led by Professors Ma Hongyue and Duan Jin'ao from Nanjing University of Chinese Medicine collaborated with Professor Xu Huaqiang's team at the Shanghai Institute of Materia Medica, Chinese Academy of Sciences. The research team selected key components from natural toad venoms with distinct medicinal properties (spirit-awakening, hallucinogenic, and detoxifying)—namely 5-OH-DMT, 5-MeO-DMT, and 5-OH-TMT—to systematically analyze their structure-target-efficacy-toxicity-property relationships. At atomic resolution, this study revealed the molecular mechanisms by which toad tryptamines selectively activate various serotonin receptor (5-HTR) subtypes under different medicinal property labels. The research provides a solid molecular pharmacological foundation for the modern interpretation of Chansu's traditional properties and profoundly illuminates why component profile variations among different source materials or processing methods of the same medicinal material can produce distinctly different medicinal properties.
The research team first systematically evaluated the agonist activities of these natural toad tryptamines on six depression-related 5-HT receptor subtypes (5-HT1A, 5-HT1B, 5-HT1D, 5-HT2A, 5-HT2C, and 5-HT7) through comprehensive functional experiments. The study revealed that subtle structural modifications in toad tryptamines result in marked selectivity differences toward receptor subtypes. Specifically, 5-MeO-DMT, characterized by "hallucinogenic" properties, exhibits potent agonist activity toward 5-HT2 receptors, particularly demonstrating high-efficiency selective agonism at the hallucinogenic target 5-HT2A. 5-OH-DMT, characterized by "spirit-awakening" properties, shows optimal selectivity and activity toward the key antidepressant target 5-HT1A while displaying significantly reduced activity at 5-HT2A receptors. 5-OH-TMT, characterized by "detoxifying" properties (with previously confirmed significant anti-inflammatory and analgesic activities), exhibits relatively weak agonist activities toward all the aforementioned 5-HTR subtypes.
Utilizing cryo-electron microscopy, the research team successfully resolved high-resolution three-dimensional structures of the three toad tryptamines in complex with the 5-HT1A receptor-Gi protein-scFv16. Structural analysis revealed that all three components occupy the orthosteric binding pocket of the receptor. Their indole rings form π-π stacking interactions with F361 and F362 residues in the TM6 region, the indole nitrogen forms hydrogen bonds with T121 on TM3, and the positively charged nitrogen of the ethylamine side chain forms salt bridge interactions with the highly conserved D116 residue. These key interactions trigger conformational changes in the receptor, including depression of the "molecular switch" W358, rearrangement of the PIF and DRY motifs, ultimately leading to an approximately 13.7 Å outward movement of the TM6 cytoplasmic end, thereby exposing the Gi protein binding interface. Notably, the indole ring of 5-OH-DMT forms a unique hydrophobic contact with the I189 residue in extracellular loop 2, a structural feature not observed in the other two compounds, which may be closely related to its distinctive pharmacological activity.
By comparing the binding modes of toad tryptamines to 5-HT1A and 5-HT2A receptors, the research team identified significant differences at key positions in the binding pockets of the two receptors. The A365 residue on TM6 of the 5-HT1A receptor is replaced by N343 in the 5-HT2A receptor, resulting in an approximately 26.6-degree rotational difference of the ligand indole ring between the two receptors. Additionally, the 5-HT2A receptor lacks the I167 residue on TM4, while V235 on TM5 participates in additional ligand interactions. These structural differences provide a molecular basis for designing non-hallucinogenic drugs with 5-HT1A selectivity and reveal the scientific foundation of Chinese medicine property identification at the molecular level.
In animal behavioral experiments, the research team employed chronic unpredictable mild stress mouse depression model and PCPA-induced depression model to systematically evaluate the antidepressant activities of toad tryptamine compounds. Through hippocampal 5-HT1A receptor knockdown experiments and receptor antagonist treatments, the study confirmed that 5-OH-DMT demonstrates a superior efficacy-toxicity ratio compared to 5-MeO-DMT. This research profoundly elucidates the scientific connotation of the traditional Chinese medicine Chansu's properties of "dispersing wind-fire stagnation" and "opening orifices to awaken the spirit," and particularly helps understand the fundamental reasons why component profile differences among different source materials, origins, or processing methods of the same medicinal material led to distinct pharmacological manifestations.
Prospects
The medicinal properties of Chinese medicines represent a core scientific issue in the discipline of Chinese materia medica, possessing rich connotations and profound substance. Using the toxic Chinese medicine Chansu as an example, this research demonstrates the approach and methodology for investigating the structure-target-efficacy-toxicity-property relationships of representative components with different medicinal property characteristics in treating emotional disorders. This "bottom-up" research strategy of exploring macroscopic medicinal properties from the microscopic world of drug-target interactions helps promote the translation of Chinese medicine property research into innovative drug development and clinical applications, thereby injecting new vitality into the modernization of traditional Chinese medicine.
Sources: https://spj.science.org/doi/10.34133/research.0987
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