Ulcerative colitis (UC), a chronic inflammatory condition that the gastrointestinal tract (Tripathi and Feuerstein, 2019), causes impairment in quality of life and is a predictor of colorectal cancer (Shah and Itzkowitz, 2022; Zhang et al., 2023). Acetic acid is widely used to induce intestinal damage in rodents, as it promotes the migration of neutrophils in the intestinal mucosa, causing an increase in the production and release of pro-inflammatory cytokines such as IL-1, IL-6 and TNF-α, reactive oxygen species (ROS) and nitrogen metabolites (Magalhães et al., 2021), thus simulating the pathophysiological characteristics of UC in humans (Porter et al., 2020; Le Berre et al., 2023).

The reduction in parasympathetic activity that has been implicated in the pathophysiology of UC may be related to immune dysregulation and gastrointestinal motility (Gunterberg et al., 2016; Mogilevski et al., 2019). Addind to this, Clinical studies show that patients with UC tend to have decreased parasympathetic activity and sympathetic predominance, reflecting autonomic dysfunction, with the increase of the clinical symptoms involved in the UC (Aghdasi-Bornaun et al., 2018). This dysfunction is associated with the downregulatioin of the M1 muscarinic receptor which is expressed in the human colon, in myenteric and submucosal neurons (Harrington et al., 2010).

The M1 muscarinic acetylcholine receptor (M1 mAChR) belongs to the superfamily of G protein-coupled receptors (GPCRs), being expressed in the intestinal mucosa (Greig et al., 2020; Uwada et al., 2023) and responsible for controlling various functions at the central and peripheral nervous system (Liu et al., 2022). A previous study carried out by our research group showed that this receptor attenuated intestinal inflammation and oxidative stress during colitis experimentally induced by peripheral administration of McN-A-343 (Magalhães et al., 2021); corroborating this finding, central administration of this agonist also reduced the severity of colitis (Ji et al., 2014; Munyaka et al., 2014).

Another important endogenous system in the regulation of intestinal inflammation refers to the endocannabinoid system (ECS), which comprises cannabinoid receptors (CB1 and CB2) and endocannabinoids such as 2-araquidonoyl glycerol (2-AG) and anandamide (Meletis, 2019; Picardo et al., 2019). These receptors are of the GPCR type and are found in the gastrointestinal tract. Treatment with CB1 and/or CB2 agonists attenuates colon inflammation, while the use of antagonists or genetic manipulations for these receptors accentuates the severity of intestinal inflammation (Lima et al., 2022; Kiran et al., 2022). Furthermore, it has been observed that 2-AG can attenuate intestinal inflammation by activating CB1 and/or CB2 receptors (Bochenek et al., 2024).

GPCR signaling has been associated with the cross-regulation of several pathways, including cross-talks between different GPCRs, leading to different physiological effects (Zhang et al., 2024). An example is the fact that the kappa opioid receptor affects the aversive properties of the cannabinoid THC (tetrahydrocannabinol) (Clasen et al., 2017). Recent research points to the possibility of allosteric modulation of M1 receptors by the kappa antagonist, norbinaltorphimine (nor-BNI) (Sousa et al., 2023). Furthermore, interaction between the muscarinic system and the ECS has also been demonstrated in the prefrontal cortex (Martin et al., 2015), hippocampus (Kim et al., 2002), cerebellum (Rinaldo and Hansel, 2013), and small intestine (jejunum) (DiPatrizio et al., 2015).

Evidence suggests that 2-AG mediates M1 mAChR-dependent depression in the prefrontal cortex (Martin et al., 2015). Furthermore, the interaction between these systems was shown during food deprivation, where the muscarinic system was found to regulate the release of 2-AG, which in turn activates cannabinoid receptors, boosting feeding (DiPatrizio et al., 2015). Thus, these findings demonstrate the cross-talk between the muscarinic system and endocannabinoids in different locations.

Considering that the muscarinic system and ECS reduced the severity of colitis, our study aimed to assess whether the attenuation of intestinal damage in the UC is dependent on the crosstalk between the M1 mAChR and the endocannabinoid system.