Activities of mixed NOP and mu-opioid receptor ligands

Abstract

Background and purpose: Compounds that activate both NOP and mu-opioid receptors might be useful as analgesics and drug abuse medications. Studies were carried out to better understand the biological activity of such compounds.

Experimental approach: Binding affinities were determined on membranes from cells transfected with NOP and opioid receptors. Functional activity was determined by [(35)S]GTPgammaS binding on cell membranes and using the mouse vas deferens preparation in vitro and the tail flick antinociception assay in vivo.

Key results: Compounds ranged in affinity from SR14150, 20-fold selective for NOP receptors, to buprenorphine, 50-fold selective for mu-opioid receptors. In the [(35)S]GTPgammaS assay, SR compounds ranged from full agonist to antagonist at NOP receptors and most were partial agonists at mu-opioid receptors. Buprenorphine was a low efficacy partial agonist at mu-opioid receptors, but did not stimulate [(35)S]GTPgammaS binding through NOP. In the mouse vas deferens, each compound, except for SR16430, inhibited electrically induced contractions. In each case, except for N/OFQ itself, the inhibition was due to mu-opioid receptor activation, as determined by equivalent results in NOP receptor knockout tissues. SR14150 showed antinociceptive activity in the tail flick test, which was reversed by the opioid antagonist naloxone.

Conclusions and implications: Compounds that bind to both mu-opioid and NOP receptors have antinociceptive activity but the relative contribution of each receptor is unclear. These experiments help characterize compounds that bind to both receptors, to better understand the mechanism behind their biological activities, and identify new pharmacological tools to characterize NOP and opioid receptors.

Figure 1

Structures of the compounds tested.

Figure 2

Concentration–response curve for N/OFQ inhibition of electrically induced contractions of the mouse vas deferens obtained in the absence and presence of 1 μM SR16430. The values are means±s.e.mean of four separate experiments.

Figure 3

Effect of N/OFQ, SR14148 and SR16476 in the electrically stimulated mouse vas deferens from NOP^+/+ (a) and NOP^−/− (b) mice. N/OFQ had pEC₅₀ of 7.49 in the NOP^+/+ mice and was inactive in the tissue from NOP^−/− animals. SR14148 had pEC₅₀ values of 5.92 and 6.02, and SR16476 had pEC₅₀ values of 6.85 and 7.08 in the NOP^+/+ and NOP^−/− mice, respectively. The values are means±s.e.mean of three separate experiments.

Figure 4

Concentration–response curve for N/OFQ inhibition of electrically induced contractions of the mouse vas deferens obtained in the absence and presence of 1 μM SR14148. Naloxone (1 μM) was added to the buffer. The values are means±s.e.mean of three separate experiments.

Figure 5

Concentration–response curve for SR14150 (a) and SR16476 (b) inhibition of electrically induced contractions of the mouse vas deferens obtained in the absence and presence of UFP-101 (10 μM) and naloxone (1 μM). The values are means±s.e.mean of three separate experiments.

Figure 6

The effect of naloxone (1 mg kg⁻¹) on SR14150-induced antinociception in the tail flick assay in vivo at 60 min post-injection time. Data are mean %MPE (±s.e.mean). An asterisk (^*P<0.05; Student–Newman–Keuls test) represents a significant difference from vehicle control, whereas a plus sign (+P<0.05; Student–Newman–Keuls test) represents a significant difference from SR14150 alone.