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pubmed:abstractText |
1. The effects of cannabinoid (CB) receptor stimulation on membrane currents in single cells from the Syrian hamster vas deferens cell line DDT1MF-2 were investigated using the whole cell patch clamp technique. 2. The CB receptor agonist CP55,940 evoked a concentration-dependent transient outward current. The selective CB1 receptor ligand SR141716 (1 microM), but not the selective CB2 receptor ligand SR144528 (1 microM), inhibited the outward current. Pertussis toxin (100 ng ml-1 for 20 h) completely abolished the outward current. 3. Western blotting with an antibody against the rat (r)CB1 receptor showed a band characteristic for the CB1 receptor around 63 kDa in DDT1MF-2 cells. 4. The reversal potential for the outward current measured using a voltage ramp protocol was -84 +/- 5 mV. The current was inhibited by the Ca2+-dependent K+ channel blockers iberiotoxin (10 nM) and charybdotoxin (10 nM). 5. Removal of Ca2+ from the bathing solution, or the addition of 0.1 mM Cd2+ completely abolished the outward current evoked by 10 microM CP55,940. 6. The sarcoplasmic Ca2+ pump inhibitor thapsigargin reduced the outward current evoked by 10 microM CP55,940 in a concentration-dependent manner. 7. The mitogen-activating protein kinase (MAP kinase) inhibitor PD98059, but not the phospholipase C inhibitor U73122, inhibited the outward current evoked by 10 microM CP55,940. 8. The adenylyl cyclase inhibitor SQ22,536 (100 microM) and 8-Br-cyclic AMP (10 microM) significantly reduced the outward current evoked by 10 microM CP55,940. 9. Our data suggest that CB1 receptor stimulation in DDT1MF-2 cells leads to activation of a large conductance Ca2+-dependent K+ channel through a Gi/Go protein-mediated rise in [Ca2+]i, for which both inhibition of adenylyl cyclase and activation of MAP kinase are required. In addition, the cannabinoid-induced increase in [Ca2+]i is likely to arise from capacitive Ca2+ entry.
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pubmed:affiliation |
Department of Biosciences, University of Hertfordshire, C. P. Snow Building, College Lane, Hatfield AL10 9AB, UK.
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