Figure 5.

5-HT_1B receptors couple preferentially to G_iα2. HeLa cells co-transfected with cDNA for the human 5-HT_1B receptor and pertussis toxin resistant forms of either (A) G_iα1, (B) G_iα2, or (C) G_iα3 were treated overnight in the presence (lanes 3 and 4) or absence (lanes 1 and 2) of 20 ng ml⁻¹ pertussis toxin before treatment with 10 μM 5-HT (lanes 2 and 4) for 5 min, and subsequent lyses. Total lysate was analysed by immunoblotting with antibody to phospho-ERK1/ERK2 (p-ERK). Membranes were then stripped and analysed with antibody to total ERK1/ERK2 (Total). Net intensities of bands were calculated from three separate experiments, performed in duplicate, and expressed as the means±s.e.mean (×10³). *P<0.05; **P<0.01; ***P<0.001 vs absence of 5-HT, two-sided paired Student t-test calculated separately for both the presence and absence of pertussis toxin. Representative immunoblots from one of the three experiments are shown to demonstrate that treatment with pertussis toxin does not alter the levels of total ERK.