Figure 7. Involvement of Ca²⁺/CaMKIIα in 5-HT₃R-mediated ERK activation.

A) Time-course of 2-Me-5-HT-induced ERK1/2 activation in the least shrew brainstem. Least shrews were injected with 5 mg/kg (i.p.) 2-Me-5-HT and their brainstems were collected at 5, 10, 20 and 30 min (n = 3 per group). Phosphorylated (pERK1/2) and total ERK1/2 of the same sample from different shrews were determined by immunoblot with the antibodies to pERK1/2 and to total ERK1/2. The ratios of pERK1/2 (42 kD/44 kD) to ERK1/2 were calculated and expressed as fold change of vehicle-treated control (0 min). Graph A represents the summarized data and the insets show the representative Western blot. *P<0.05 vs. 0 min. Graphs B–D) Immunoblot analyses of ERK1/2 phosphorylation were performed on brainstems collected from the experimental shrews 10 min after 2-Me-5-HT treatment (5 mg/kg, i.p.) in the absence (vehicle) or presence of antagonists. B) Selective blockade of 5-HT₃Rs with palonosetron (5 mg/kg, s.c.) 30 min prior to 2-Me-5-HT injection. *P<0.05 vs. vehicle/vehicle control and ^#P<0.05 vs. vehicle + 2-Me-5-HT. C) Either vehicle (Veh, i.p.), the inositol-1, 4, 5-triphosphate receptor blocker 2-APB (10 mg/kg. i.p.), L-type Ca²⁺ channel blocker amlodipine (Aml, 10 mg/kg, s.c.), ryanodine receptor blocker dantrolene (Dan, 20 mg/kg, i.p.) or a combination (Aml+Dan) of less effective doses of amlodipine (5 mg/kg, s.c.) and dantrolene (10 mg/kg, i.p.) were administered to different groups of shrews 30 min prior to 2-Me-5-HT injection. *P<0.05 vs. Veh/Veh control (Ctl). ^#P<0.05 vs. Veh + 2-Me-5-HT. ^aP<0.05 vs. 2-APB + 2-Me-5-HT. D) Inhibition of CaMKII with KN93 (10 mg/kg, i.p.) blocked 2-Me-5-HT-evoked ERK1/2 phosphorylation in brainstem. n = 3 per group. Graphs show the summarized data and insets show representative Western blots. *P<0.05 vs. vehicle/vehicle control. ^#P<0.05, vs. vehicle + 2-Me-5-HT.