Abstract
Cannabinoid receptor (CB) expression was characterized in immunological cell and tissue preparations. Northern analysis revealed ∼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 ∼4-kb transcript in mouse spleen and thymus and as ∼2.4-kb transcripts in rat spleen. Quantitation of CB2 transcripts in mouse spleen and thymus revealed ∼4 × 103and ∼4 × 102molecules/100 ng RNA, respectively, with no quantifiable CB2 in mouse brain. Conversely, CB1 was expressed in mouse brain (∼2 × 105molecules/100 ng RNA) with lower expression in mouse spleen (∼2 × 102molecules/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, Δ9-THC, and cannabinol all competed for receptor binding with3H-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 approximateKdof 200–700 pmin purified splenic T cells which also exhibited cannabinoid-induced inhibition of adenylate cyclase. Northern analysis of human T-cell lines revealed ∼2.4-kb CB2 mRNA transcripts but no CB1 in HPB-ALL cells, a cell line which also exhibited inhibition of adenylate cyclase by Δ9-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 Δ9-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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