Pharmacological profile of NOP receptors coupled with calcium signaling via the chimeric protein Gαqi5

Abstract

In this study, the Gαqi5 protein was used to force the human nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor to signal through the Ca2+ pathway in CHO cells. [Ca2+]i levels were monitored using the fluorometer FlexStation II and the Ca2+ dye Fluo 4AM. Concentration response curves were generated with a panel of full and partial agonists, while NOP antagonists
were assessed in inhibition-response curves. The following rank order of potency of antagonists was measured: ${\text{SB}} - {\text{612111}} >{\text{J}} - 113397 = {\text{Trap}} - 101 \geqslant {\text{UFP}} - 101 >\left[ {{\text{Nphe}}^{\text{1}} } \right]{{\text{N}} \mathord{\left/ {\vphantom {{\text{N}} {{\text{OFQ}}}}} \right. \kern-\nulldelimiterspace} {{\text{OFQ}}}}\left( {1 - 13} \right){\text{NH}}_{\text{2}} >>${\text{SB}} - {\text{612111}} >{\text{J}} - 113397 = {\text{Trap}} - 101 \geqslant {\text{UFP}} - 101 >\left[ {{\text{Nphe}}^{\text{1}} } \right]{{\text{N}} \mathord{\left/ {\vphantom {{\text{N}} {{\text{OFQ}}}}} \right. \kern-\nulldelimiterspace} {{\text{OFQ}}}}\left( {1 - 13} \right){\text{NH}}_{\text{2}} >> naloxone, which is superimposable to literature findings. The rank order of potency of full and partial agonists is also
similar to that obtained in previous studies with the exception of a panel of ligands (UFP-112, Ro 64-6198, ZP120, UFP-113)
whose potency was relatively low in the Gαqi5–NOP receptor calcium assay. Interestingly, these NOP ligands are characterized by slow kinetic of interaction with the NOP
receptor, as demonstrated by bioassay experiments. These results demonstrated that the FlexStation II–Gαqi5–NOP receptor calcium assay represents an adequate and useful screening for NOP receptor ligands, particularly for antagonists.