Localisation of Cannabinoid and Cannabinoid-Related Receptors in the Horse Ileum

The endocannabinoid system (ECS) is composed of endogenous ligands (N-arachidonylethanolamine [anandamide, AEA] and 2-arachidonoyl glycerol [2-AG]), G-protein-coupled receptors (cannabinoid receptors 1 and 2 [CB1R and CB2R]) and enzymes to degrade and recycle the ligands [1]. Phytochemicals produced by the Cannabis sativa plant can interact with the ECS as ligands.

In addition, phytocannabinoids may exert their effects by binding not only with the canonical CB1R and CB2R, but also with other receptors, such as G-protein coupled receptors, nuclear receptors, transient receptor potential channels, serotonin receptors and glycine receptors [2].

Therefore, the definition of the ECS is expanding, and includes endocannabinoid-like mediators, such as palmitoylethanolamide (PEA) in addition to the classical cannabinoid receptors and endocannabinoids, [3].

The ECS regulates different gastrointestinal functions, including motility, secretion, maintenance of the epithelial barrier integrity, emesis, and appetite or satiety, under both physiological and pathological conditions [4], [5], [6].

Both the CB1R and the CB2R can be found throughout the gastrointestinal tract (GIT). Cannabinoid receptor 1 is predominantly located in the enteric nervous system (ENS) and the epithelial lining [7,8]. In the ENS, the CB1R is expressed in cholinergic neurons, explaining why CB1R receptor activation can reduce intestinal motility [9]. In the epithelium, the CB1R participates in epithelial wound healing during intestinal inflammation [10]. Cannabinoid receptor 2 is mainly expressed in intestinal lamina propria cells, suggesting a role in immunomodulation [11].

Colic, a common digestive disorder in horses, includes different forms of abdominal pain [12]. It is one of the most dangerous emergency problems in equine medicine, and one of the principal causes of death in horses; therefore, it is a primary health concern of owners [13].

Colic syndrome can include volvulus [14], impaction [15], and idiopathic muscular hypertrophy of the ileum as well as other conditions [16], all of which can involve the distal ileum and the ileocaecal region.

To these significant naturally occurring syndromes of equine intestinal dysmotility, which are still poorly understood, the induced condition of postoperative ileus should also be added as it is a very frequent disorder after laparotomy and intestinal manipulation/resection [17].

Due to the common occurrence of gastrointestinal disorders in the horse, efforts have been made to investigate new therapeutic molecules directed principally against visceral and somatic pain in this species.

In recent years, several companies have been producing and/or marketing medical marijuana and cannabinoid receptor agonists to be used in equine medicine. Early studies and anecdotal experience has suggested that cannabinoids can be very useful as anxiolytics and for analgesia in horses [18,19,20,21]. It is interesting to note that, even in the early 1900s, the U.S. Department of War recommended the use of Cannabis extracts for colic in horses [22].

Cannabidiol (CBD), a non-psychoactive compound extracted from Cannabis sativa, seems to be one of the most promising therapeutic substances due to its analgesic, anti-inflammatory and anti-spasmodic benefits [23], [24], [25].

Knowing the cellular distribution of the specific receptors is fundamental in understanding the action of a drug. To date, reliable histological studies regarding the cellular distribution of cannabinoid and cannabinoid-related receptors in the GIT of domestic mammals have been published only for dogs and cats [26,27] whereas studies in the horse intestinal tract are still lacking.

In order to help in filling these histological gaps, the present study was designed to identify the cellular distribution of two canonical cannabinoid receptors, i.e. CB1R and CB2R, and of three different putative cannabinoid receptors, that is nuclear peroxisome proliferator-activated receptor alpha (PPARα), transient potential ankyrin receptor 1 (TRPA1), and serotonin receptor 1a (5-HT1aR) in the equine distal ileum.

The latter represents, together with the ileocaecal junction (Papilla ilealis), the highly specialised intestinal region in which the flow of ingesta is dramatically slowed. This region is often involved in colic syndrome in association with small intestinal obstruction and/or strangulation, and undifferentiated enteritis [28], [29], [30], [31].