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Published on: January 27, 2014
Cannabinoid receptors CB1 and CB2: a characterization of expression and adenylate cyclase modulation within the
A R Schatz1, M Lee, R B Condie
1Department of Pharmacology & Toxicology, Michigan State University, East Lansing, USA.
Insights
Cannabinoid receptor type 2 (CB2) is the primary cannabinoid receptor in the immune system, particularly on T cells. CB2 expression was detected in mouse spleen and thymus, while CB1 was mainly in the brain.
Area of Science:
- Immunology
- Neuroscience
- Pharmacology
Background:
- Cannabinoid receptors (CB) play roles in various physiological processes.
- Previous studies indicated cannabinoids can inhibit T-cell-dependent immune responses.
Purpose of the Study:
- To characterize the expression and function of cannabinoid receptors (CB1 and CB2) in immune cells.
- To determine the predominant cannabinoid receptor subtype involved in immune cell responses.
Main Methods:
- Northern blot analysis to detect CB1 and CB2 mRNA transcripts in various tissues and cell lines.
- Radioligand binding assays to quantify receptor binding sites and affinity.
- Adenylate cyclase activity assays to assess receptor-mediated signaling.
Main Results:
- CB2 receptor mRNA was detected in mouse spleen and thymus, and as 2.4-kb transcripts in rat spleen.
- CB1 receptor mRNA was found in mouse brain and rat cerebellum but not in mouse thymus or rat spleen.
- T cells exhibited cannabinoid receptor binding sites and showed inhibition of adenylate cyclase activity upon cannabinoid treatment.
- Human T-cell lines (HPB-ALL) expressed CB2 mRNA and showed inhibited adenylate cyclase, while Jurkat E6-1 cells showed unusual CB2 mRNA patterns and no response.
Conclusions:
- CB2 is the predominant cannabinoid receptor expressed in the immune system, particularly on T cells.
- The findings support a role for CB2 in modulating immune cell function.
- Differential expression and signaling of cannabinoid receptors in T-cell lines warrant further investigation.
Abstract:
Cannabinoid receptor (CB) expression was characterized in immunological cell and tissue preparations. Northern analysis revealed approximately 6-kb transcripts for CB1 (brain-type) in mouse spleen and brain and in rat cerebellum. CB1 was not detected in mouse thymus or rat spleen RNA by Northern analysis. CB2 (peripheral) was detected as a approximately 4-kb transcript in mouse spleen and thymus and as approximately 2.4-kb transcripts in rat spleen. Quantitation of CB2 transcripts in mouse spleen and thymus revealed approximately 4 x 10(3) and approximately 4 x 10(2) molecules/100 ng RNA, respectively, with no quantifiable CB2 in mouse brain. Conversely, CB1 was expressed in mouse brain (approximately 2 x 10(5) molecules/100 ng RNA) with lower expression in mouse spleen (approximately 2 x 10(2) molecules/100 ng RNA) and was not quantifiable in mouse thymus. Competition binding in intact mouse splenocytes demonstrated that nonradiolabeled cannabinoids CP-55940, Win-55212-2, CP-56667, delta 9-THC, and cannabinol all competed for receptor binding with 3H-CP-55940, a high-affinity nondiscriminating CB1 and CB2 receptor ligand. Based on previous findings which demonstrated a marked inhibition of T-cell-dependent immune responses by cannabinoids, primary T cells and several T-cell lines were characterized. Radioligand binding analysis identified 100-300 cannabinoid receptor binding sites/cell with an approximate Kd of 200-700 pM in purified splenic T cells which also exhibited cannabinoid-induced inhibition of adenylate cyclase. Northern analysis of human T-cell lines revealed approximately 2.4-kb CB2 mRNA transcripts but no CB1 in HPB-ALL cells, a cell line which also exhibited inhibition of adenylate cyclase by delta 9-THC. Conversely, Jurkat E6-1 cells expressed an unusual mRNA banding pattern for CB2 expressing three distinct transcript sizes, none of which were 2.4 kb, the size for human CB2. Jurkat also did not express CB1 mRNA and did not exhibit inhibition of adenylate cyclase when treated with delta 9-THC. Collectively, these results provide further evidence that CB2 is the predominant cannabinoid receptor within the immune system and that this form of the receptor is expressed on T cells.
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