Molecular cloning, mutations and effects of NK1 receptor antagonists reveal the human-like pharmacology of gerbil NK1 receptors

Biochem Pharmacol. 2007 Jan 15;73(2):259-69. doi: 10.1016/j.bcp.2006.09.030. Epub 2006 Oct 12.

Abstract

The present study investigates the pharmacology of the cloned neurokinin 1 receptor from the gerbil (gNK(1)R), a species claimed to have human-like NK(1)R (hNK(1)R) pharmacology. The amino acid sequence of NK(1)R was cloned. The hNK(1)R, rat NK(1)R (rNK(1)R), gNK(1)R and mutants of the gNK(1)R were expressed in CHO cells. The affinity and potency of NKR agonists and the NK(1)R antagonists CP99994 and RP67580 (NK(1)R-selective) and ZD6021 (NK1/2R) were assessed in vitro by monitoring [(3)H]-SarMet SP binding and substance P-evoked mobilization of intracellular Ca(2+). The gerbil foot tap (GFT) method was used to assess the potency of the antagonists in vivo. The gNK(1)R coding sequence displayed an overall 95% and 97% homology with hNK(1)R and rNK(1)R, respectively. The affinity of the NK(1)R-selective agonist (3)H-SarMet SP for human and gerbil NK(1)R was similar (2.0 and 3.1 nM) but lower for rNK(1)R (12.4 nM). The rank order potency of the agonists for NK(1)R was SP > or = ASMSP > or = NKA >>> pro7NKB in all species. The NK(1)R antagonists, ZD6021 and CP99994, had comparable affinity and potency for gerbil and human NK(1)R, but were 1000-fold less potent for rNK(1)R. In contrast, RP67580 had comparable affinity and potency for all three species. Mutations in positions 116 and 290 did not affect agonist potency at the gNK(1)R while the potency of the antagonists ZD6021 and CP99994 were markedly decreased (10-20-fold). It is concluded that gNK(1)R has similar antagonist pharmacology as the human-like orthologue and that species differences in antagonist function depend on key residues in the coding sequence and antagonist structure.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • CHO Cells
  • Cloning, Molecular
  • Cricetinae
  • Cricetulus
  • DNA Primers
  • Gerbillinae
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Neurokinin-1 Receptor Antagonists*
  • Rats
  • Receptors, Neurokinin-1 / chemistry
  • Receptors, Neurokinin-1 / genetics
  • Sequence Homology, Amino Acid

Substances

  • DNA Primers
  • Neurokinin-1 Receptor Antagonists
  • Receptors, Neurokinin-1