Multiplicity of kappa opioid receptor binding in the rat cardiac sarcolemma.
In order to determine the opioid receptor binding in the rat cardiac sarcolemma, direct and displacement binding assays of tritiated bremazocine, a nonselective opioid, agonist and U69593, a specific kappa-receptor agonist, were performed in a sarcolemmal membrane preparation. Saturation studies revealed that there were substantial binding sites for both radioligands. The [3H]bremazocine had a binding capacity of 100 fmol/mg protein with a Kd of 3.4 nM while the binding of [3H]U69593 showed a Kd of 7.4 nM and a Bmax of 139 fmol/mg protein. Displacement binding assay showed that [D-Ala2, MePhe4, Gly-(ol)5]enkephalin (DAGO), a specific mu-agonist, exhibited no inhibitory effect on the binding of either [3H]bremazocine or [3H]U69593. [D-Ala2,D-Leu5]enkephalin (DADLE), a specific delta-agonist, had no inhibition on the binding of [3H]U69593, but displaced the binding of [3H]bremazocine at higher concentrations (10(-5)-10(-6)M). U50488, a specific kappa-agonist, completed with the binding of [3H]U69593 23 times more potently than with the binding of [3H]bremazocine. A residual binding sites of [3H]bremazocine was found with a Kd of 6.4 nM and a Bmax of 76 fmol/mg protein under conditions in which mu, delta and kappa 1 opioid bindings were suppressed by DAGO and DADLE at 200 nM and U50488 at 1 microM. DADLE at 5 microM, a concentration known to block kappa 2 binding sites, abolished the residual U50488-insensitive specific binding of [3H]bremazocine. Met5-Enkephalin-Arg6-Phe7, a specific kappa 2 ligand, displaced the binding of [3H]bremazocine much more readily than the binding of [3H]U69593. The present study provided evidence for the presence of two kappa-receptor subtypes in the cardiac sarcolemma: one U50488-sensitive and DADLE-insensitive, and the other U50488-insensitive and DADLE-sensitive. These two kappa-receptor binding subtypes may correspond to kappa 1 and kappa 2 receptor subtypes. Computer assisted non-linear regression analysis in the competition experiments of U50488 with [3H]U69593 afforded a best fit of the inhibition curves for two displacing components of kappa 1 binding sites: one with high affinity and low Bmax, another with low affinity and high Bmax.
Zhang
W M
WM
Department of Physiology, University of Hong Kong, Hong Kong.
Jin
W Q
WQ
Wong
T M
TM
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000700:Analgesics / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D044482:Benzeneacetamides; D001575:Benzomorphans / Q000378:metabolism* / Q000494:pharmacology; D005952:Glucose-6-Phosphatase / Q000378:metabolism; D008297:Male; D009206:Myocardium / Q000378:metabolism*; D011759:Pyrrolidines / Q000378:metabolism* / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017473:Receptors, Opioid, kappa / Q000378:metabolism*; D012508:Sarcolemma / Q000378:metabolism; D014316:Tritium
D000700:Analgesics; D044482:Benzeneacetamides; D001575:Benzomorphans; D011759:Pyrrolidines; D017473:Receptors, Opioid, kappa; D014316:Tritium; D005952:Glucose-6-Phosphatase; D000251:Adenosine Triphosphatases; C027248:bremazocine; C045444:U 69593