Original Article

Ca2+ participates in α1B-adrenoceptor-mediated cAMP response in HEK293 cells1

Yao SONG, Yun-fang LI, Er-dan DONG, Qi-de HAN, You-yi ZHANG


Aim: To investigate the α1B-adrenoceptor (α1B-AR)-mediated cAMP response and
underlying mechanisms in HEK293 cells.
Methods: Full-length cDNA encoding
α1B-AR was transfected into HEK293 cells using the calcium phosphate precipitation
method, and α1B-AR expression and cAMP accumulation were determined
by using the saturation radioligand binding assay and ion-exchange chromatography,
Results: Under agonist stimulation, α1B-AR mediated cAMP
synthesis in HEK293 cells, and blockade by PLC-PKC or tyrosine kinase did not
reduce cAMP accumulation induced by NE. Pretreatment with pertussis toxin
(PTX) had little effect on basal cAMP accumulation as well as norepinephrine
(NE)-stimulated cAMP accumulation. In addition, pretreatment with cholera toxin
(CTX) neither mimicked nor blocked the effect induced by NE. The extracellular
Ca2+ chelator egtazic acid (EGTA), nonselective Ca2+ channel blocker CdCl2 and
calmodulin (CaM) inhibitor W-7 significantly reduced NE-induced cAMP accumulation
from 1.59%±0.47% to 1.00%±0.31%, 0.78%±0.23%, and 0.90%±0.40%,
Conclusion: By coupling with a PTX-insensitive G protein, α1BAR
promotes Ca2+ influx via receptor-dependent Ca2+ channels, then Ca2+ is linked
to CaM to form a Ca2+-CaM complex, which stimulates adenylyl cyclase (AC),
thereby increasing the cAMP production in HEK293 cell lines.