Opioid receptor ligands in the neonatal rat spinal cord: binding and in vitro depression of the nociceptive responses.
1. Opioid receptors in the neonatal rat spinal cord have been characterized by measurements of ligand binding to crude membrane fractions and by functional tests on the nociceptive spinal response in a spinal cord-tail preparation in vitro. 2. There were high affinity binding sites for [3H]-[D-Ala2, MePhe4, Gly(ol)5]enkephalin (DAGOL), [3H]-U69593, and [3H]-ethylketocyclazocine (EKC) on spinal cord membranes from neonatal rats. Hill slopes for binding of [3H]-DAGOL and [3H]-U69593 were close to unity. The Hill slope for binding of [3H]-EKC was less than unity, even after its interactions at mu-receptors had been blocked with 100 nM unlabelled DAGOL. Binding sites for [3H]-[D-Pen2, D-Pen5]enkephalin (DPDPE) could not be detected. 3. In competition assays U50488 was as potent as PD117302 and U69593 in competition for either [3H]-U69593 or [3H]-EKC binding sites. Hill slopes for a range of competing ligands at [3H]-DAGOL or [3H]-U69593 sites were close to unity. Hill slopes for competition at [3H]-EKC sites were less than one. 4. In the spinal cord-tail preparation from neonatal rats, opioid receptor agonists depressed spinal nociceptive responses evoked by application of capsaicin or heat to the tail. The order of potency was DAGOL greater than U69593 = PD117302 greater than morphine greater than U50488 = [D-Pen2, L-Pen5]enkephalin (DPLPE). 5. The antagonist naloxone was about equally potent against DAGOL, morphine and DPLPE, and about ten times less potent against U69593 and PD117302. The effects of U50488 were much less sensitive to blockade by naloxone than the effects of PD11703 or U69593. The Kappa antagonist, nor-binaltorphimine was equipotent against all three Kappa agonists. 6. The absence of delta-binding sites, and the low potency and relatively high sensitivity to naloxone suggest that DPLPE could be working at mu-receptors in the neonatal rat spinal cord. 7. The binding assays show that U50488 has the same affinity as PD1 17302 and U69593 for Kappa-receptors, yet it was less effective in the depression of nociceptive responses. This may be because U50488 has a relatively low efficacy at Kappa-receptors. It is possible that at high concentrations U50488 activates receptors not affected by other Kappa-ligands. These additional receptors may be non-opioid receptors (hence the insensitivity to naloxone), or they could be a subtype of Kappa-opioid receptor.
James
I F
IF
Sandoz Institute for Medical Research, London.
Bettaney
J
J
Perkins
M N
MN
Ketchum
S B
SB
Dray
A
A
D000818:Animals; D000831:Animals, Newborn; D044482:Benzeneacetamides; D003496:Cyclazocine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000378:metabolism / Q000494:pharmacology; D016314:Ethylketocyclazocine; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D009020:Morphine / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D009270:Naloxone / Q000494:pharmacology; D009294:Narcotics / Q000494:pharmacology; D009619:Nociceptors / Q000187:drug effects*; D011759:Pyrrolidines / Q000494:pharmacology; D051381:Rats; D011957:Receptors, Opioid / Q000187:drug effects* / Q000378:metabolism; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D013116:Spinal Cord / Q000378:metabolism*
D044482:Benzeneacetamides; D004745:Enkephalins; D008024:Ligands; D009294:Narcotics; D011759:Pyrrolidines; D011957:Receptors, Opioid; D017465:Receptors, Opioid, delta; D017473:Receptors, Opioid, kappa; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D009270:Naloxone; D016314:Ethylketocyclazocine; D009020:Morphine; D020881:Enkephalin, D-Penicillamine (2,5)-; C045444:U 69593; D003496:Cyclazocine