NK1R-mediated activation of ERK1/2. (a and
b) Cells were incubated with 10 nM SP (KNRK) or 100 nM
[Sar9MetO211]SP (SM-SP, HDMEC)
for 0–60 min. Phosphorylation of ERK1/2 in KNRK-NK1R cells
(a) and HDMEC cells (b).
(Inset) representative pERK Western blot. *,
P < 0.05, as compared with untreated controls,
n = 4. (c) KNRK-NK1R, KNRK-NK1R +
ARR319–418, and KNRK-NK1Rδ325 cells were treated with 10
nM SP for 5 min and pERK1/2 was determined. (Inset)
pERK Western blots of untreated and SP-treated cells
(Top, NK1R; Middle,
NK1R+ARR319–418; Bottom, NK1R, δ325).
*, P < 0.05 as compared with SP-treated
KNRK-NK1R cells, n = 3. (d)
KNRK-NK1R cells were treated with 10 nM SP for 5 min with vehicle for
inhibitors (veh), or after pretreatment with 100 nM GF103209X (GFX), 20
μM genistein (GEN), or 20 μM tyrphostin 25 (TYR) for 20 min, and
pERK1/2 was determined. *, P < 0.05 for
cells treated with SP-treated controls, n = 3.
(e) KNRK-NK1R and KNRK-NK1Rδ325 cells were treated
with or without 10 nM SP for 5 min and immunoprecipitated with either
PYK2 antibody (Left) or shc antibody
(Right), followed by Western blotting with
phosphotyrosine (pTYR) antibody.