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Absence of Entourage: Terpenoids Commonly Found in Cannabis sativa Do Not Modulate the Functional Activity of ?9-THC at Human CB1 and CB2 Receptors.

ABSTRACT: Introduction: Compounds present in Cannabis sativa such as phytocannabinoids and terpenoids may act in concert to elicit therapeutic effects. Cannabinoids such as ?9-tetrahydrocannabinol (?9-THC) directly activate cannabinoid receptor 1 (CB1) and cannabinoid receptor 2 (CB2); however, it is not known if terpenoids present in Cannabis also affect cannabinoid receptor signaling. Therefore, we examined six common terpenoids alone, and in combination with cannabinoid receptor agonists, on CB1 and CB2 signaling in vitro. Materials and Methods: Potassium channel activity in AtT20 FlpIn cells transfected with human CB1 or CB2 receptors was measured in real time using FLIPR® membrane potential dye in a FlexStation 3 plate reader. Terpenoids were tested individually and in combination for periods up to 30?min. Endogenous somatostatin receptors served as a control for direct effects of drugs on potassium channels. Results: ?-Pinene, ?-pinene, ?-caryophyllene, linalool, limonene, and ?-myrcene (up to 30-100??M) did not change membrane potential in AtT20 cells expressing CB1 or CB2, or affect the response to a maximally effective concentration of the synthetic cannabinoid CP55,940. The presence of individual or a combination of terpenoids did not affect the hyperpolarization produced by ?9-THC (10??M): (CB1: control, 59%±7%; with terpenoids (10??M each) 55%±4%; CB2: ?9-THC 16%±5%, with terpenoids (10??M each) 17%±4%). To investigate possible effect on desensitization of CB1 responses, all six terpenoids were added together with ?9-THC and signaling measured continuously over 30?min. Terpenoids did not affect desensitization, after 30?min the control hyperpolarization recovered by 63%±6% in the presence of the terpenoids recovery was 61%±5%. Discussion: None of the six of the most common terpenoids in Cannabis directly activated CB1 or CB2, or modulated the signaling of the phytocannabinoid agonist ?9-THC. These results suggest that if a phytocannabinoid-terpenoid entourage effect exists, it is not at the CB1 or CB2 receptor level. It remains possible that terpenoids activate CB1 and CB2 signaling pathways that do not involve potassium channels; however, it seems more likely that they may act at different molecular target(s) in the neuronal circuits important for the behavioral effect of Cannabis.

SUBMITTER: Santiago M

PROVIDER: S-EPMC6757242 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Absence of Entourage: Terpenoids Commonly Found in Cannabis sativa Do Not Modulate the Functional Activity of Δ9-THC at Human CB1 and CB2 Receptors.

Santiago Marina M Sachdev Shivani S Arnold Jonathon C JC McGregor Iain S IS Connor Mark M

Cannabis and cannabinoid research 20190923 3

Introduction: Compounds present in Cannabis sativa such as phytocannabinoids and terpenoids may act in concert to elicit therapeutic effects. Cannabinoids such as Δ9-tetrahydrocannabinol (Δ9-THC) directly activate cannabinoid receptor 1 (CB1) and cannabinoid receptor 2 (CB2); however, it is not known if terpenoids present in Cannabis also affect cannabinoid receptor signaling. Therefore, we examined six common terpenoids alone, and ...[more]

PMID: 31559333

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Project description:Introduction: Cannabis sativa produces hundreds of bioactive compounds, including cannabinoids and terpenoids. It has been proposed that cannabinoids act in synergy with terpenoids to produce the entourage effect, a concept used to explain the therapeutic benefits of medicinal cannabis. One molecular explanation for the entourage effect is that the terpenoids augment the actions of cannabinoids at their molecular drug targets in cells. We recently reported that terpenoids commonly found in cannabis do not influence the functional effects of Δ9-tetrahydrocannabinol (Δ9-THC) on cannabinoid 1 and cannabinoid 2 receptors. The present study aimed to extend on this research by examining whether terpenoids influence the effects of phytocannabinoids and endocannabinoids on human transient receptor potential ankyrin 1 (hTRPA1) and human transient receptor potential vanilloid 1 (hTRPV1) channels heterologously expressed in mammalian cells. Materials and Methods: The activity of terpenoids, phytocannabinoids, and endocannabinoids was assessed in inducible HEK Flp-In T-Rex cells transfected with hTRPA1 and hTRPV1 channels, respectively. Real-time changes in intracellular calcium ([Ca]i) were measured using the Calcium 5 dye and a FlexStation 3 plate reader. Results: α-pinene, β-pinene, β-caryophyllene, linalool, limonene, β-myrcene or α-humulene did not affect [Ca]i in hTRPA1 and hTRPV1 overexpressing cells. Cinnamaldehyde (CA), Δ9-THC, and 2-arachidonoylglycerol (2-AG) activated TRPA1 receptors with high efficacy and similar potency (EC50s of ∼10 μM). Capsaicin and anandamide (AEA) activated TRPV1 receptors with an EC50 of 61 nM and 4.3 μM, respectively, but TRPV1 showed no response to Δ9-THC, cannabidiol, and other minor cannabinoids. Terpenoids did not significantly affect the responses of TRPA1 and TRPV1 receptors to submaximal and maximal concentrations of CA and Δ9-THC or the endocannabinoids AEA and 2-AG. Discussion: We could not find any evidence that the terpenoids tested here activate TRPA1 and TRPV1 channels or modulate their activation by Δ9-THC and other agonists, including endocannabinoids.

Project description:BackgroundGlandular capitate trichomes which form on bract tissues of female inflorescences of high THC-containing Cannabis sativa L. plants are important sources of terpenes and cannabinoids. The influence of plant age and cannabis genotype on capitate trichome development, morphology, and maturation has not been extensively studied. Knowledge of the various developmental changes that occur in trichomes over time and the influence of genotype and plant age on distribution, numbers, and morphological features should lead to a better understanding of cannabis quality and consistency.MethodsBract tissues of two genotypes-"Moby Dick" and "Space Queen"-were examined from 3 weeks to 8 weeks of flower development using light and scanning electron microscopy. Numbers of capitate trichomes on upper and lower bract surfaces were recorded at different positions within the inflorescence. Observations on distribution, extent of stalk formation, glandular head diameter, production of resin, and extent of dehiscence and senescence were made at various time points. The effects of post-harvesting handling and drying on trichome morphology were examined in an additional five genotypes.ResultsTwo glandular trichome types-bulbous and capitate (sessile or stalked)-were observed. Capitate trichome numbers and stalk length were significantly (P = 0.05) greater in "Space Queen" compared to "Moby Dick" at 3 and 6 weeks of flower development. Significantly more stalked-capitate trichomes were present on lower compared to upper bract surfaces at 6 weeks in both genotypes, while sessile-capitate trichomes predominated at 3 weeks. Epidermal and hypodermal cells elongated to different extents during stalk formation, producing significant variation in length (from 20 to 1100 μm). Glandular heads ranged from 40 to 110 μm in diameter. Maturation of stalked-capitate glandular heads was accompanied by a brown color development, reduced UV autofluorescence, and head senescence and dehiscence. Secreted resinous material from glandular heads appeared as droplets on the cuticular surface that caused many heads to stick together or collapse. Trichome morphology was affected by the drying process.ConclusionCapitate trichome numbers, development, and degree of maturation were influenced by cannabis genotype and plant age. The observations of trichome development indicate that asynchronous formation leads to different stages of trichome maturity on bracts. Trichome stalk lengths also varied between the two genotypes selected for study as well as over time. The variability in developmental stage and maturation between genotypes can potentially lead to variation in total cannabinoid levels in final product. Post-harvest handling and drying were shown to affect trichome morphology.

Dataset Information

Absence of Entourage: Terpenoids Commonly Found in Cannabis sativa Do Not Modulate the Functional Activity of ?9-THC at Human CB1 and CB2 Receptors.

Publications

Absence of Entourage: Terpenoids Commonly Found in <i>Cannabis sativa</i> Do Not Modulate the Functional Activity of Δ<sup>9</sup>-THC at Human CB<sub>1</sub> and CB<sub>2</sub> Receptors.

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