[3H]-[H-D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2] ([3H]CTOP), a potent and highly selective peptide for mu opioid receptors in rat brain.
The cyclic, conformationally restricted octapeptide [3H]-[H-D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2] ([3H]CTOP) was synthesized and its binding to mu opioid receptors was characterized in rat brain membrane preparations. Association rates (k+1) of 1.25 x 10(8) M-1 min-1 and 2.49 x 10(8) M-1 min-1 at 25 and 37 degrees C, respectively, were obtained, whereas dissociation rates (k-1) at the same temperatures were 1.93 x 10(-2) min-1 and 1.03 x 10(-1) min-1 at 25 and 37 degrees C, respectively. Saturation isotherms of [3H]CTOP binding to rat brain membranes gave apparent Kd values of 0.16 and 0.41 nM at 25 and 37 degrees C, respectively. Maximal number of binding sites in rat brain membranes were found to be 94 and 81 fmol/mg of protein at 25 and 37 degrees C, respectively. [3H]CTOP binding over a concentration range of 0.1 to 10 nM was best fit by a one site model consistent with binding to a single site. The general effect of different metal ions and guanyl-5'-yl-imidodiphosphate on [3H]CTOP binding was to reduce its affinity. High concentrations (100 mM) of sodium also produced a reduction of the apparent mu receptor density. Utilizing the delta opioid receptor specific peptide [3H]-[D-Pen2,D-Pen5]enkephalin, CTOP appeared to be about 2000-fold more specific for mu vs. delta opioid receptor than naloxone. Specific [3H]CTOP binding was inhibited by a large number of opioid or opiate ligands.(ABSTRACT TRUNCATED AT 250 WORDS)
Hawkins
K N
KN
Department of Pharmacology, University of Arizona Health Sciences Center, Tucson.
Knapp
R J
RJ
Lui
G K
GK
Gulya
K
K
Kazmierski
W
W
Wan
Y P
YP
Pelton
J T
JT
Hruby
V J
VJ
Yamamura
H I
HI
D000818:Animals; D001921:Brain / Q000378:metabolism; D004723:Endorphins / Q000494:pharmacology; D020881:Enkephalin, D-Penicillamine (2,5)-; D004745:Enkephalins / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011957:Receptors, Opioid / Q000378:metabolism*; D017450:Receptors, Opioid, mu; D012964:Sodium / Q000494:pharmacology; D013004:Somatostatin / Q000031:analogs & derivatives* / Q000378:metabolism; D013696:Temperature; D014316:Tritium
D004723:Endorphins; D004745:Enkephalins; D011957:Receptors, Opioid; D017450:Receptors, Opioid, mu; D014316:Tritium; C049352:phenylalanyl-cyclo(cysteinyltyrosyl-tryptophyl-ornithyl-threonyl-penicillamine)threoninamide; D013004:Somatostatin; D020881:Enkephalin, D-Penicillamine (2,5)-; D012964:Sodium; D008274:Magnesium