engleski

The effect of acute and chronic treatment of mouse thymoma cell lines with kappa-opioid agonist

The results described in this communication were obtained during the first author's one- year stay in USA as an INVEST/NIDA postdoctoral fellow 1995/1996 in Dr. Jean M. Bidlack's laboratory. It was demonstrated by Dr. Bidlack and co-workers that mouse thymoma cell lines R1.1, R1.G1 and R1EGO express only k-opioid receptors which are negatively coupled to adenylyl cyclase through a G-inhibitory protein which is sensitive to pertussis toxin. R1.G1 cells express three times, while R1EGO cells six times more k-opioid receptors than the parent cell line R1.1. Chronic treatment of neuronal cells with k-agonists leads to down-regulation of the receptors (decreased receptor number) and to the receptor desensitization (i.e. the loss of receptor responsiveness). Two derivative cell lines R1.G1 and R1EGO cells were tested for the effect of acute (15 min) and chronic (24 hr) treatment with a k-opioid agonist (-)U50,488 (100 nM). (^3H)U69,593 was used for the saturation binding experiments to determine Kd (dissociation constant) and Bmax (receptor number). Adenylyl-cyclase activity was measured by a modified RIA kit using the membranes of the cells treated with (-) U50,488. Bmax values for the R1.G1 cell line were 266 fmol/mg protein (15 min) and 142 fmol/mg protein (24 hr). For the R1EGO cell line the Bmax values were 250 fmol/mg protein (15 min) and 149 fmol/mg protein (24 hrs). The Kd value for (3H)U69,593 binding to the R1G1 cell line memranes was 2.00 nM for the 15 min pretreatment and 2.13 nM for the 24 hr pretreatment with 100 nM of (-)U50,488. For the R1EGO cell line the Kd values were 1.90 nM (15 min) and 2.40 nM (24 hrs). The rate of inhibition of the adenylyl-cyclase activity by (-)U50,488 was not affected by pretreatment of either cell line with 100 nM (-)U50,488 for 24 hrs. Thus, chronic opioid treatment of R1.G1 and R1EGO mouse thymoma cell lines resulted in down-regulation of the k-opioid receptor without desensitization, which is consistent with the results obtained with the R1.1 parent cell line.

kappa-opioid receptors; thymomas; adenylyl cyclase

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