kappa-Opioid binding sites on a murine lymphoma cell line.
As a first step in determining whether any subset of lymphocytes expresses opioid receptors, membranes prepared from mouse lymphoma cell lines were screened for [3H]naloxone binding sites. Membranes from the R1.1 cell line specifically bound [3H]naloxone. The Hill coefficient for [3H]naloxone binding was 0.93 +/- 0.18, and nonlinear regression analysis indicated that a one-site model was the best fit of the [3H[naloxone saturation binding data. Low concentrations of kappa-selective opioids, but neither mu nor delta opioids, inhibited [3H]naloxone binding. Saturation binding studies with the kappa-selective compound [3H]U69,593 revealed a single binding site with a KD value of 0.204 +/- 0.039 nM and a Bmax value of 31.7 +/- 3.1 fmol/mg of membrane protein. The Hill coefficient for [3H]U69,593 binding was 1.03 +/- 0.11, indicative of a single site. Time courses for the association and dissociation of [3H]U69,593 binding at 25 degrees C exhibited properties consistent with a single class of binding sites. Low concentrations of kappa-selective opioids, including dynorphin peptides, inhibited [3H]U69,593 binding, while high concentrations of mu opioids were needed to inhibit binding, and the delta-selective ligands were ineffective at concentrations up to 10 microM. Stereoselectivity of the binding site was demonstrated by the finding that the Ki value for (-)-pentazocine in inhibiting [3H]U69,593 binding was 25 times less than for the (+)-isomer. Based on its high affinity for U69,593, alpha-neo-endorphin, and dynorphin B, the kappa opioid binding site on R1.1 cell membranes belongs to the kappa 1b subtype. As observed with brain kappa opioid binding sites, sodium inhibited [3H]U69,593 binding to R1.1 cell membranes in a concentration-dependent manner. These data demonstrate that the murine lymphoma cell line R1.1 expresses kappa opioid binding sites that are very similar to brain kappa opioid binding sites.
Bidlack
J M
JM
Department of Pharmacology, University of Rochester, School of Medicine and Dentistry, NY 14642.
Saripalli
L D
LD
Lawrence
D M
DM
D000818:Animals; D044482:Benzeneacetamides; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D008214:Lymphocytes / Q000378:metabolism; D008223:Lymphoma / Q000378:metabolism; D051379:Mice; D009270:Naloxone / Q000378:metabolism; D011759:Pyrrolidines / Q000378:metabolism; D011957:Receptors, Opioid / Q000378:metabolism*; D014316:Tritium; D014407:Tumor Cells, Cultured / Q000378:metabolism
D044482:Benzeneacetamides; D011759:Pyrrolidines; D011957:Receptors, Opioid; D014316:Tritium; D009270:Naloxone; C045444:U 69593