Mechanisms of cannabinoid tolerance

Abstract

Cannabis has been used recreationally and medically for centuries, yet research into understanding the mechanisms of its therapeutic effects has only recently garnered more attention. There is evidence to support the use of cannabinoids for the treatment of chronic pain, muscle spasticity, nausea and vomiting due to chemotherapy, improving weight gain in HIV-related cachexia, emesis, sleep disorders, managing symptoms in Tourette syndrome, and patient-reported muscle spasticity from multiple sclerosis. However, tolerance and the risk for cannabis use disorder are two significant disadvantages for cannabinoid-based therapies in humans. Recent work has revealed prominent sex differences in the acute response and tolerance to cannabinoids in both humans and animal models. This review will discuss evidence demonstrating cannabinoid tolerance in rodents, non-human primates, and humans and our current understanding of the neuroadaptations occurring at the cannabinoid type 1 receptor (CB₁R) that are responsible tolerance. CB₁R expression is downregulated in tolerant animals and humans while there is strong evidence of CB₁R desensitization in cannabinoid tolerant rodent models. Throughout the review, critical knowledge gaps are indicated and discussed, such as the lack of a neuroimaging probe to assess CB₁R desensitization in humans. The review discusses the intracellular signaling pathways that are responsible for mediating CB₁R desensitization and downregulation including the action of G protein-coupled receptor kinases, β-arrestin2 recruitment, c-Jun N-terminal kinases, protein kinase A, and the intracellular trafficking of CB₁R. Finally, the review discusses approaches to reduce cannabinoid tolerance in humans based on our current understanding of the neuroadaptations and mechanisms responsible for this process.

Keywords: Cannabinoid; Cannabis use disorder; Desensitization; Downregulation; THC; Tolerance.

Figure 1.. Roles of ECS in physiology and human health.

Endocannabinoids, such as the AEA and 2-AG, have been implicated in the modulation and maintenance of many human organ systems. These effects may be induced (+), suppressed (−), or variable (+/−) following endocannabinoid activation.

Figure 2.. CB₁R modulation of intracellular signaling pathways.

Agonist-induced activation by CB₁R causes inhibition of VGCCs and adenylyl cyclase and production of cAMP while stimulating GIRK potassium channels and MAPK signaling pathways such as extracellular-regulated kinase (ERK) 1/2, JNK, and p38. Activation of CB₁R also increases PKC and PI3K signaling pathways. A major overall effect of CB₁R activation is that it inhibits neuronal signaling by preventing neurotransmitter release (inhibition of VGCCs) and hyperpolarization of the membrane potential (activation of GIRKs).

Figure 3.. CB₂R-mediated signaling pathways.

While the targets of CB₂R signaling are largely similar to that of CB₁R, differences in biases for these pathways account for significant differences in physiology and behaviors when activated. Additionally, CB₂R activation has substantial anti-inflammatory components, which are produced through stimulation of ceramide and phospholipase C (PLC) pathways.

Figure 4.. Wildtype mice display faster tolerance to Δ⁹-THC compared to CP55,940, a strongly internalizing cannabinoid agonist.

Wild-type mice were treated with once-daily intraperitoneal injections of 0.3 mg/kg CP55,940 (red circles and line) or 30 mg/kg Δ⁹-THC (THC; black squares and line) once daily and tested for tail-flick antinociception and body temperature 1 hour after drug treatment. To measure antinociception, a Columbus Instruments TF-1 tail-flick analgesia meter (Columbus, OH) was calibrated to an intensity of 5. To avoid potential tissue damage to the tail, the instrument was programmed to a 10 s cut-off time. Tail-flick measurements were recorded between 2–5 times for each time point. The recorded measurements were used to calculate the antinociceptive response as a percent of the maximum possible effect (%MPE) using the following equation: %MPE = [(post-drug latency)-(pre-drug latency)]/[pre-determined cut-off time (10 s)-(pre-drug latency)]x100. Hypothermia was assessed by taking each subject’s body temperature using a mouse rectal thermometer (Physiotemp Instruments, Clifton, NJ) prior to and 60 minutes following injection. Recorded values, in °C, were used to calculate the percent change in body temperature (%Δ) = [(post-body temperature)-(pre-body temperature)/(pre-body temperature)]x100. Data represent mean values ± SEM and were analyzed by mixed two-way ANOVA with Bonferroni post-hoc tests (*p<0.05, **P<0.01, ***p<0.001, ****p<0.0001). For this experiment, tolerance is defined as the decrease in antinociceptive response each day that occurs with repeated dosing. There were significant effects of drug treatment (F_1,46=28.56, p<0.0001), time (F_4.417,195.8=11.74, p<0.0001), and a time × drug treatment interaction effect (F_6,266=4.530, p<0.001) for the antinociceptive effects of CP55,940 versus Δ⁹-THC. There were significant effects of drug treatment (F_1,50=112.3, p<0.0001), time (F_{3.858,187.1=94.36}, p<0.0001), and a time × drug treatment interaction effect (F_6,291=9.038, p<0.0001) for the hypothermic effects of CP55,940 versus Δ⁹-THC. The sample size of tested animals is designated in parentheses.

Figure 5.. CB₁R desensitization.

Upon activation of CB₁R by an agonist (A), the coupled G protein disassociates. While the alpha subunit participates in signaling mechanisms, the beta and gamma complex recruits GRK. Phosphorylation of the C-terminus of CB₁R by GRK results in the recruitment of β–arrestin2 (βArr2) which causes steric inhibition and prevents the G protein from interacting with CB₁R. The association of β–arrestin2 with CB₁R also causes recruitment of adaptor protein-2 (AP-2) and clathrin which facilitate endocytosis of the receptor. This mechanism of desensitization reduces response to CB₁R activation and signaling, and repetitions of this process leads to downregulation and tolerance.

Figure 6.. CB₁R downregulation.

Clathrin-mediated endocytosis results in internalization of CB₁Rs, which is sorted to be degraded or recycled. Increased bioavailability of resensitized receptors may reduce tolerance, while degradation of the receptor and reduction of receptor bioavailability promotes tolerance.