Exploring the mechanism of action of huoermai essential oil for plateau insomnia based on the camp/CREB/BDNF/gabaergic pathway.

ETHNOPHARMACOLOGICAL RELEVANCE

The traditional Huoermai therapy is a treatment for insomnia used by the Tibetan people living on the Tibetan plateau in China. This therapy involves the use of Myristica fragrans Houtt. and Carum carvi L., along with fomentation and massage, and has shown significant clinical effects. However, the mechanism of how Huoermai therapy treats plateau insomnia needs further clarification.

AIM OF THE STUDY

This study aimed to investigate the mechanism of action of Huoermai essential oil (HEO) in treating plateau insomnia, focusing on the cAMP/CREB/BDNF/GABAergic pathway.

METHODS

The major components of Huoermai essential oil were identified by Gas chromatography-mass spectrometry (GC-MS) for subsequent network pharmacology analysis. Proteomics techniques were employed to pinpoint disparities in brain tissue protein expression in a mouse model of plateau insomnia following Huoermai therapy administration, in conjunction with network pharmacology to forecast pathways related to hypoxia and insomnia. Plateau insomnia mouse model was established and the therapeutic impact of Huoermai essential oil was evaluated. Hematoxylin & Eosin staining(HE) was conducted to observe pathological damage to the cortex, hippocampus, thalamus and hypothalamus structures. Changes in serotonin (5-HT), melatonin (MT), adenosine (AD), cyclic adenosine monophosphate (cAMP) and malondialdehyde (MDA) levels in mouse brain tissue were gauged through enzyme-linked immunosorbent assay (ELISA) to assess sleep status and oxidative stress levels in mice. Molecular docking was employed to anticipate the target binding energy of Huoermai essential oil constituents. ELISA and Western Blot (WB) were used to ascertain the expression of cAMP/CREB/BDNF/GABAergic pathway.

RESULTS

The results indicated that HEO positively impacted intermittent hypobaric hypoxia-induced plateau insomnia in mice. Histological examination results showed that HEO ameliorated neuronal damage in specific regions of the brain affected by plateau insomnia, such as the cortex, hippocampus, thalamus, and hypothalamus. Through GC-MS analysis, 56 volatile oil components were identified. Subsequently, a combined network pharmacology and proteomics analyses led to selecting the cAMP/CREB/BDNF/GABAergic pathway for further study. ELISA experiments demonstrated that HEO treatment increased GABA and MT levels while significantly reducing 5-HT and adenosine levels in brain tissue of mice with plateau insomnia. WB results revealed that HEO ameliorated plateau insomnia by suppressing the hyperactivation of the cAMP pathway, increasing brain-derived neurotrophic factor (BDNF) levels and B-cell lymphoma-2 (BCL-2) expression, and alleviating hypoxia-induced oxidative stress. Moreover, molecular docking results showed strong binding affinity of all pharmacological components to their targets and proteins in the brain.

CONCLUSION

These results indicate that HEO significantly prolongs sleep duration in plateau insomniac mice and treats plateau insomnia by modulating levels of sleep-related regulators, modulating the cAMP pathway, increasing GABA receptor expression, and improving neuronal survival and anti-apoptosis.