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Signal transduction mechanisms of recombinant bovine neurokinin‐2 receptor stably expressed in Baby hamster kidney cells
Author(s) -
Eistetter Harald R.,
Mills Ann,
Arkinstall Stephen J.
Publication year - 1993
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240520112
Subject(s) - neurokinin a , neurokinin b , arachidonic acid , chinese hamster ovary cell , receptor , inositol phosphate , microbiology and biotechnology , second messenger system , biology , inositol trisphosphate , endocrinology , medicine , chemistry , inositol , biochemistry , substance p , neuropeptide , enzyme
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 [ 125 I]neurokinin A (NKA) binding sites/cell characterized further. [ 125 I]NKA binding was displaced by unlabeled NKA with an IC 50 of 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 (IP 3 ) levels (EC 50 = 32 ± 10 nM; n = 7) which was paralleled by a transient biphasic rise in intracellular free calcium concentration [Ca 2+ ] i (EC 50 = 35 ± 20 nM; n = 3). In addition to phosphoinositide (PI) hydrolysis and Ca 2+ mobilization, NKA was found to stimulate both cyclicAMP formation (EC 50 = 1.02 ± 0.26 μM; n = 7) and [ 3 H]arachidonic acid mobilization (EC 50 = 0.65 ± 0.45 μM; n = 4). Interestingly, cyclicAMP levels also rose after addition of an exogenous arachidonic acid metabolite, prostaglandin E 2 (PGE 2 ) (EC 50 = 11.5 ± 2 μM). Similar observations of NKA‐induced IP 3 production, Ca 2+ 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.