Our study demonstrates that C. chinensis significantly relaxes the LES. Previous research has shown that C. chinensis affects gastrointestinal tract motility and function. It demonstrates its effectiveness in alleviating visceral pain in irritable bowel syndrome [8], improving gastrointestinal function in combination with Dolomiaea souliei [13], and impacting immune health and metabolism in various processed forms [14]. Additionally, it enhances intestinal barrier function in treatments for ulcerative colitis [15].

Berberine, extracted from C. chinensis, is a bright yellow compound renowned for its antibacterial and anti-inflammatory properties. Historically utilized as a fabric dye [16], it has also shown potential in combatting atherosclerosis through mechanisms such as lipid regulation, reduction of blood pressure and blood sugar, inflammation control, and inhibition of vascular smooth muscle cell proliferation [17]. Furthermore, berberine demonstrates vasorelaxant and antiproliferative effects [18], highlighting its versatility and broad therapeutic potential.

The impact of berberine on gut microbiota is a major area of research, with significant implications for the management of diabetes, hyperlipidemia, atherosclerosis, and liver diseases [19]. Its role extends to metabolic regulation, underlining its potential in treating obesity and neurodegenerative diseases [20].

In addition to its relaxant impact on the LES observed in our study, berberine also significantly influences gastrointestinal motility. It modulates this motility by relaxing rat gastric fundus muscle through the inhibition of calcium entry [21], reduces the contractility of gastric intestinal smooth muscle [9], and demonstrates therapeutic effects on irritable bowel syndrome by inhibiting colonic smooth muscle neurotransmission [22]. Furthermore, berberine suppresses intestinal myoelectric activity and transit, potentially via opioid and α-adrenergic receptors [23], and obstructs both extracellular and intracellular Ca2+ flow in colon smooth muscle cells [24]. Among these effects, the antidiarrheal properties of berberine are particularly noteworthy, with a systematic review and meta-analysis affirming its efficacy and safety in treating diarrhea among both children and adults [25]. This evidence highlights berberine’s potential as a versatile treatment option for a range of gastrointestinal disorders.

Smooth muscle relaxation is a crucial physiological process mediated by multiple signaling pathways. To elucidate the mechanisms behind berberine-induced relaxation of porcine LES smooth muscle, we used various pharmacological agents targeting potential neural mediation and pathways involving cyclic nucleotides, nitric oxide, and potassium channels [26]. Our comprehensive approach to investigating the potential mechanisms provides valuable insights into berberine’s mode of action.

The assessment of potential neural mediation in berberine-induced relaxation of the porcine LES involved the use of TTX and CTX. The absence of inhibition with TTX and CTX is remarkable, suggesting that the relaxation induced by berberine in the porcine LES is not neurally mediated.

We employed rolipram and vardenafil, which are inhibitors of PDE-4 and PDE-5, respectively. By inhibiting these enzymes, rolipram and vardenafil increase the levels of cAMP and cGMP, potentially enhancing smooth muscle relaxation [7, 27]. However, these inhibitors did not enhance berberine’s relaxing effect on the porcine LES. Similarly, KT5720 and KT5823, which are selective inhibitors of cAMP and cGMP, had no impact on the berberine-induced relaxation in the LES. This indicates that the mechanisms of berberine-induced relaxation do not involve the cAMP, cGMP, or NO pathways [10]. Furthermore, the use of L-NNA, a competitive inhibitor of nitric oxide synthase, did not impede the berberine-induced relaxation of the porcine LES. This finding suggests that NO production is not associated with the relaxation induced by berberine.

Furthermore, we evaluated the role of potassium channels in the berberine-induced relaxation of the porcine LES using specific inhibitors. These included IbTX, apamine, TEA and glibenclamide. Notably, the blockade of potassium channels with TEA led to a significant reduction in the relaxation of porcine LES muscle [10, 12]. However, neither apamine, IbTX, nor glibenclamide significantly affected the berberine-induced relaxation in the porcine LES. These results suggest that the berberine-induced relaxation of the porcine LES is primarily mediated through potassium channels. Future research could further explore the exact molecular identity of the TEA-sensitive channels involved and investigate potential upstream signaling pathways that lead to the activation of these channels.

These findings underscore the distinct mechanism of action of berberine compared to conventional achalasia therapies. Conventional treatments such as nifedipine function by inhibiting calcium influx into smooth muscle cells, while nitrates work by increasing nitric oxide levels [1]. In contrast, berberine primarily induces LES relaxation via potassium channel activation, independent of cAMP, cGMP, and NO pathways. Consequently, berberine may offer an advantage over conventional treatments in terms of reduced systemic side effects.

Research highlights the effects of TCM and natural substances on the LES and esophageal function. The Modified Xiaochaihu Decoction enhances LES pressure and reduces ineffective swallowing, showing potential in treating gastroesophageal reflux disease [28]. Arecae pericarpium extracts, particularly arecoline, induce dose-dependent LES contractions [29]. Research indicates ginger does not alter LES resting pressure or esophageal contractions, but increases LES relaxation and reduces contraction velocity, potentially aiding gas expulsion [30]. 6-Gingerol, found in ginger, is known to increase LES tone [31]. Curcumae longae Rhizoma extract has been shown to lessen esophageal tissue damage and lower biochemical markers in acute reflux esophagitis [32]. Additionally, peppermint oil, a smooth muscle relaxant, effectively treats diffuse esophageal spasm [33]. These findings enhance our understanding of traditional medicines’ impact on LES motility.

Our findings on C. chinensis and berberine’s relaxant effects on the LES offer new perspectives for treating gastrointestinal disorders, bridging TCM with modern pharmacology. The identified potassium channel-mediated relaxation mechanism suggests potential for more targeted therapies. While potassium channel modulators are not yet widely used for gastrointestinal disorders, research has shown their promise in cardiovascular and neurological conditions [34,35,36]. This mechanism may have applications in other smooth muscle disorders [37] and could lead to personalized treatments for LES disorders, opening new directions in gastrointestinal medicine research.

Our pilot study provides valuable initial insights into the potential of C. chinensis and berberine for treating LES disorders. However, our research primarily relies on ex vivo porcine LES models, which limits full understanding of the compounds’ effects. To address these limitations and enhance the study’s impact, we propose incorporating in vitro cell line models and conducting animal studies. These additions will provide crucial insights into molecular mechanisms, toxicity, pharmacokinetics/pharmacodynamics, and safety profiles. Clinical trials are essential to confirm the relaxant effects on human LES and assess efficacy in treating LES motility disorders. Future research should prioritize optimizing dosage and administration methods, evaluating long-term safety and efficacy, exploring effects on other gastrointestinal motility disorders, and investigating potential synergies. Additionally, further studies could explore the effects of other compounds within C. chinensis and their potential contributions to LES relaxation and underlying mechanisms. By addressing these areas, we can work towards developing improved treatments for gastrointestinal motility disorders, offering alternative hope to patients with conditions like achalasia.