The Endocannabinoid System A brief molecular overview

The Endocannabinoid System

A brief molecular overview

Stes de Necker

Cannabis, once a taboo subject, has made a pragmatic shift into mainstream culture. The plant has become a popular topic of conversation worldwide, in part due to changes in public opinion, increased medical access for patients, and a changing approach to drug enforcement focusing on harm reduction rather than criminalization.

Currently, cannabis is the most consumed illicit drug in Western culture (Radhakrishnan et al., 2014), and it has a long history of human use for medicinal purposes (Mechoulam and Parker, 2013).

 However, a turbulent past and restrictive policy have made it difficult to explore the therapeutic mechanisms of cannabis (Pacher et al., 2006).

Despite these major barriers to academic research, our understanding of the molecular mechanisms that mediate the effects of cannabis continues to advance, with the hope that this work may contribute to novel therapies to combat human disease.  The goal of this article is to provide a generalized overview of the intracellular signalling mechanisms that are responsible for the effects of cannabis.

The molecular components of the endocannabinoid system

Originally, it was proposed that the effects of cannabis were mediated by a nonspecific membrane-associated mechanism (Mechoulam and Parker, 2013); however, this hypothesis was later refuted as various components of the endocannabinoid system (ECS) were discovered in the late 1980s and early 90s, revolutionizing our understanding of the effects of cannabis (Howlett et al., 2002).

The ECS is an important and highly conserved cellular signaling pathway found within vertebrates, where it modulates multiple critical physiological processes (Pacher et al., 2006; Russo, 2016). 

Dysregulation of the ECS pathway has been implicated in several human diseases (Mackie, 2006; Pacher et al., 2006).

The ECS is comprised of two cannabinoid receptors (CB), CB1 and CB2, which belong to the G-coupled protein receptor super family, whose members influence various cellular processes by sensing extracellular signalling molecules (Mechoulam and Parker, 2013).  CB1 is most abundantly expressed in the central and peripheral nervous system, with the exception of the brain stem region (Hu and Mackie, 2015).  CB1 is also present, although less abundant, in peripheral organs such as the digestive tract, epidermis, adipose tissue, and skeletal muscle, as well as the cardiovascular, lymphatic, and respiratory systems (Mackie, 2006).  CB2 receptor expression, on the other hand, is mainly restricted to immune cells and tissues, but has recently been described in certain glial and neuronal cell subpopulations, as well as in bone and liver (Mackie, 2006).

CB receptors are endogenously activated by lipid-based ligands, collectively termed the endocannabinoids (listed in Table 1), which bind to CB1 and CB2 with differing affinities and potency (De Petrocellis et al., 2004).  Unlike most other neurotransmitters, which are stored in synaptic vesicles and released upon appropriate cellular stimuli, endocannabinoids are made on demand by postsynaptic neurons and cross the synapse to inhibit presynaptic activity (Mechoulam and Parker, 2013).  Endocannabinoids are also subject to regulation at these synapses.  For example, endocannabinoids are regulated by a known set of enzymes that control ECS activation by modulating ligand levels in response to cellular demand.

Phytocannabinoids

In addition to the endocannabinoids, exogenous plant derived phytocannabinoids also activate the ECS pathway via CB binding (see Table 1).  Polypharmaceutical cannabis produces hundreds of metabolites, including terpenes, flavonoids, phenolic compounds and various cannabinoids (Russo, 2011).  Most notable are the two major cannabinoids found in cannabis: psychoactive ∆-9-tetrahydrocannabinol (THC) and non-psychoactive cannabidiol (CBD).  THC is a partial CB1 and CB2 activator, while CBD has little affinity for CB1 and CB2 but can inhibit THC-receptor binding at low concentrations (Russo and Guy, 2006).

In addition to THC and CBD, other less abundant phytocannabinoids can bind the CB receptors with differing affinities, stimulating or inhibiting downstream ECS activation.   Moreover, terpenoids and other cannabis metabolites are thought to synergistically contribute to the therapeutic effects of phytocannabinoids in a phenomenon known as the ‘entourage effect’ (Russo, 2011).  The entourage effect hypothesis, originally coined by R. Mechoulam’s group (Ben-Shabat et al., 1998), states that complex, multiple compound mixtures show synergistic biological activity, where isolated compounds fail to achieve similar effects, thus providing the rationale for whole plant extracts versus individual compounds in treating disease.

Downstream ECS signalling mechanisms

What do we know about the downstream ECS signalling events?  CB receptors activate a number of signal transduction pathways through the Gi/o family of G-coupled protein receptors in response to extracellular ligand binding, which in turn influence key cellular processes (Howlett et al., 2002) 

For example, ECS activation influences several types of ion channels and it is hypothesized that short, versus sustained, CB activation may underlie differences in downstream intracellular signalling, where pathway stimulation for seconds alters ion channels and sustained activation modulates intracellular enzymes activity, such as protein kinase A (PKA) and Mitogen-activated protein kinase (MAPK) (Stella, 2009).

As well, the ECS pathway is implicated in regulating immediate early gene family expression, protein synthesis and nitric oxide production (Howlett et al., 2002).  In cancer and tumour cells, elevated ceramide release by a CB-dependent mechanism has been linked to decreased cellular growth and increased programmed cell death within these cells (Howlett et al., 2002).

Thus, ECS activation appears to influence a number of key, context dependent cellular signaling mechanisms, and the future challenge will be to understand exactly how these ECS-dependent signaling events translate into physiological changes at the tissue level.  We do know, for example, that CB1 activation in the central nervous system suppresses neurotransmitter release at excitatory and inhibitory synapses (Mechoulam and Parker, 2013), and CB2 activation modulates immune system function by inhibiting cytokine release and immune cell migration (Mechoulam and Parker, 2013), which is thought to contribute to the therapeutic benefits reported by patients.

Other signalling pathways influenced by Endocannabinoids

CB receptors have an affinity for endocannabinoids, phytocannabinoids and a number of synthetic compounds that often activate or block one type of receptor more potently than another, but are there non-CB receptor targets for these ligands?  If so, what is the physiological consequence of binding these alternative receptors, and does this mechanism contribute to any disease conditions or therapeutic application of phytocannabinoids?

It is known that canonical ECS ligands, such as endocannabinoids and phytocannabinoids, can influence other major cell signalling pathways.  For example, CBD modulates serotonin signalling (Franklin and Carrasco, 2014; Resstel et al., 2009; Russo et al., 2005), an important chemical messenger in the regulation of emotional state, pain, and behaviour, and this mechanism has been linked to CBD-dependent anxiolytic-like effects (Resstel et al., 2009).

Also, Vanilloid type 1 receptor (TRPV1), a putative cannabinoid receptor, is activated by several endocannabinoids (Pertwee, 2006) and is implicated in the regulation of vasodilation, thermoregulation and pain (Elphick and Egertova, 2001).  Similarly, GPR55, another hypothesized putative cannabinoid receptor, is activated by canonical ECS ligands. And although the exact physiological significance of GPR55 activation is unknown (Brown, 2007), it has been implicated in the regulation of bone development (Whyte et al., 2009), cancer cell proliferation (Pineiro et al., 2011), and inflammation (Balenga et al., 2011).  Evidence also suggests endocannabinoids and phytocannabinoids activate the peroxisome proliferators-activated receptor (PPAR) family, which primarily regulates energy balance and metabolism, cell differentiation and inflammation (O’Sullivan, 2007).

More recently, phtyocannabinoids and endocannabinoids were shown to inhibit the Hedgehog pathway, a signalling system critically important during development and tissue homeostasis, by direct receptor inhibition (Khaliullina et al., 2015).

Taken together, the promiscuity of typical ECS ligands to bind and activate other pathways raises the interesting possibility that some of the effects of cannabis could be a result of ECS-independent signalling, widening the scope of mechanistic investigation beyond simply CB1 and CB2 receptors.

Conclusion

Our understanding of the ECS system and the use of cannabis as a therapeutic tool in pathological and normal conditions is becoming more clear, but there remain significant gaps in our understanding of these processes.  The next major challenge, from a research perspective, will be tying together how downstream intracellular signalling mechanisms influence tissue and organ system function, and how these changes can be exploited to treat human disease.  Currently, access to cannabis for research purposes is cumbersome.  As laws change, it will be increasingly important that policy be updated to better balance the restriction of access to reduce public harm with the need for reasonable access to the plant for research purposes.

Increasing access to cannabis for research by reducing hurdles and maintaining a rational approach to drug policy will allow Canada to become an international leader in the field of cannabis research. This in turn will lead to large economic benefits, in the form of improved or novel therapeutic treatments to combat human disease, and and new or improved agricultural markets.

Table 1. List of Endocannabinoids and major phytocannabinoids.

Endocanabinoids

2-arachidonoyl glycerol (2-AG)

N-arachidonoyl-ethanolamine (anandamide, AEA)

2-arachidonyl-glyeryl ether (Noladin, 2-AGE)

O-arachidonyl-thanolamine (virhodamine)

N-arachidonoyl-dopamine (NADA)

Phytocannabinoids

Cannabigerol (CBG)

∆-9-tetrahydrocannabinol (THC)

Cannabidiol (CBD)

Cannabichromene (CBC)

Cannabinol (CBN)

-Dr. Charles Campbell and Dr. Randy Ringuette

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Dr. Charles Campbell and Dr. Randy Ringuette are experienced life scientists who share a strong passion for improving the healthcare system and treatment of human disease through basic research.

Together,they have over 15 years in a medical research environment and have published multiple original, peer-reviewed publications.

Both Dr. Campbell and Dr. Ringuette hold a Ph.D. degree in Cellular & Molecular Medicine and an honours B.Sc. degree in Biology from the University of Ottawa, and they are joint founders of Apical Science Inc., a scientific consulting and research group focused on the emerging Canadian Cannabis Industry.

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Citations

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