Cannabinoid Receptor 2 (CB₂) Signals via G-alpha-s and Induces IL-6 and IL-10 Cytokine Secretion in Human Primary Leukocytes

Abstract

Cannabinoid receptor 2 (CB 2 ) is a promising therapeutic target for immunological modulation. There is, however, a deficit of knowledge regarding CB 2 signaling and function in human primary immunocompetent cells. We applied an experimental paradigm which closely models the in situ state of human primary leukocytes (PBMC; peripheral blood mononuclear cells) to characterize activation of a number of signaling pathways in response to a CB 2 -selective ligand (HU308). We observed a "lag" phase of unchanged cAMP concentration prior to development of classically expected Gα i -mediated inhibition of cAMP synthesis. Application of G protein inhibitors revealed that this apparent lag was a result of counteraction of Gα i effects by concurrent Gα s activation. Monitoring downstream signaling events showed that activation of p38 was mediated by Gα i , whereas ERK1/2 and Akt phosphorylation were mediated by Gα i -coupled βγ. Activation of CREB integrated multiple components; Gα s and βγ mediated ∼85% of the response, while ∼15% was attributed to Gα i . Responses to HU308 had an important functional outcomesecretion of interleukins 6 (IL-6) and 10 (IL-10). IL-2, IL-4, IL-12, IL-13, IL-17A, MIP-1α, and TNF-α were unaffected. IL-6/IL-10 induction had a similar G protein coupling profile to CREB activation. All response potencies were consistent with that expected for HU308 acting via CB 2 . Additionally, signaling and functional effects were completely blocked by a CB 2 -selective inverse agonist, giving additional evidence for CB 2 involvement. This work expands the current paradigm regarding cannabinoid immunomodulation and reinforces the potential utility of CB 2 ligands as immunomodulatory therapeutics.

Cannabinoids as Key Regulators of Inflammasome Signaling: A Current Perspective

“…They also can activate phosphatidylinositol-3-kinase-protein kinase B (PI3K-AKT), ceramide production, and the expression of various genes (24,41,43,44). Interestingly, CB1 and CB2 receptors interact with G s proteins as well to induce cAMP production under specific circumstances (45,46). CB1 receptor activation specifically stimulates G-protein-gated inwardly rectifying potassium (GIRK) channels and inhibits voltagegated (N-type) Ca 2+ channels.…”

Cannabinoids and Cannabinoid Receptors: The Story so Far

“…CB 2 agonists have been explored for slowing neurodegenerative disorders such as Huntingtons chorea and Alzheimer disease (Maccarrone et al, 2007). CB 2 receptors function by inhibiting the activity of adenylyl cyclase through their G I /G oa subunits (Bouaboula et al, 1996;Soethoudt et al, 2017) by coupling to stimulatory Ga i/o subunits, causing a rise in intracellular cAMP levels, and by influencing the Ras-Ref-MEK-ERK pathway, which has a profound effect on mature and neoteric tissues (Slipetz et al, 1995;Saroz et al, 2019).…”

Optical Control of Cannabinoid Receptor 2-Mediated Ca2+ Release Enabled by Synthesis of Photoswitchable Probes

“…16,49,50 More recent studies suggest that CB2 can also couple to other G proteins and effector pathways. 18,51,52 CB2 shows pronounced biased signaling on activation by different ligands, and the effects of CB2 activation on intracellular Ca 2+ concentration ([Ca 2+ ]i) have only been described on few occasions. 22,23,[53][54][55][56] The study of [Ca 2+ ]i To the best of our knowledge, CB2-mediated modulation of [Ca 2+ ]i has not been reported for HU308 derivatives, thus we were intrigued to study Ca 2+ signaling in living cells using HU308 and its photoswitchable derivatives.…”

“…[15][16][17] Functional understanding is required to fully realize the receptor's therapeutic potential. CB2 is linked to a large number of cellular signaling pathways 18 and exhibits pronounced biased signaling in response to different ligands and tissue-specific physiology. 19 Ca 2+ ions are ubiquitous intracellular messengers in cellular signaling, and they are of emerging interest in connection with Alzheimer's and other neurodegenerative diseases linked to CB2.…”