SIGNAL-TRANSDUCTION MECHANISMS OF RECOMBINANT BOVINE NEUROKININ-2 RECEPTOR STABLY EXPRESSED IN BABY HAMSTER-KIDNEY CELLS

The bovine neurokinin-2 (NK-2) receptor gene was stably transfected into Baby hamster kidney (BHK-21) fibroblasts and one recombinant clone expressing 17,700 high-affinity [1251]neurokinin A (NKA) binding sites/cell characterized further. [I-125]NKA binding was displaced by unlabeled NKA with an IC50 8.26 +/- 2 nM (n = 5) and with the rank order of potency NKA > neurokinin B (NKB) > Substance P (SP) confirming pharmacological characteristics of an NK-2 receptor subtype. Stimulation with NKA resulted in a rapid and dose-dependent increase in inositol 1,4,5-trisphosphate (IP3) levels (EC50 = 32 +/- 10 nM; n = 7) which was paralleled by a transient biphasic rise in intracellular free calcium concentration [Ca2+], (EC50 = 35 +/- 20 nM; n = 3). In addition to phosphoinositide (PI) hydrolysis and Ca2+ mobilization, NKA was found to stimulate both cyclicAMP formation (EC50 = 1.02 +/- 0.26 muM; n = 7) and [H-3]arachidonic acid mobilization (EC50 = 0.65 +/- 0.45 muM; n = 4). Interestingly, cyclicAMP levels also rose after addition of an exogenous arachidonic acid metabolite, prostaglandin E2 (PGE2) (EC50 = 11.5 +/- 2 muM). Similar observations of NKA-induced IP3 production, Ca2+ mobilization, arachidonic acid liberation, and cAMP formation have been made previously following expression of the bovine NK-2 receptor in Chinese hamster ovary (CHO) epithelial cells. The present results suggest that activation of NK-2 receptors leads to characteristic and reproducible intracellular second messenger responses in a subclass of cell types which includes fibroblasts and epithelial cells irrespective of their genetic and phenotypic background.