Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Objective: Sustained beta-adrenergic stimulation is a hallmark of sympathetic hyperactivity in cardiovascular diseases. It is associated with oxidative stress and altered vasoconstrictor tone. This study investigated the beta-adrenoceptor subtype and the signaling pathways implicated in the vascular effects of b-adrenoceptor overactivation. Methods and Results: Mice lacking the beta(1)- or beta(2)-adrenoceptor subtype (beta 1KO, beta 2KO) and wild-type (WT) were treated with isoproterenol (ISO, 15 mu g.g(-1). day(-1), 7 days). ISO significantly enhanced the maximal vasoconstrictor response (Emax) of the aorta to phenylephrine in WT (+34%) and beta 1KO mice (+35%) but not in beta 2KO mice. The nitric oxide synthase (NOS) inhibitor L-NAME abolished the differences in phenylephrine response between the groups, suggesting that ISO impaired basal NO availability in the aorta of WT and beta 1KO mice. Superoxide dismutase (SOD), pertussis toxin (PTx) or PD 98,059 (p-ERK 1/2 inhibitor) incubation reversed the hypercontractility of aortic rings from ISO-treated WT mice; aortic contraction of ISO-treated beta 2KO mice was not altered. Immunoblotting revealed increased aortic expression of Gi alpha-3 protein (+50%) and phosphorylated ERK1/2 (+90%) and decreased eNOS dimer/monomer ratio in ISO-treated WT mice. ISO enhanced the fluorescence response to dihydroethidium (+100%) in aortas from WT mice, indicating oxidative stress that was normalized by SOD, PTx and L-NAME. The ISO effects were abolished in beta 2KO mice. Conclusions: The beta(2)-adrenoceptor/Gi alpha signaling pathway is implicated in the enhanced vasoconstrictor response and eNOS uncoupling-mediated oxidative stress due to ISO treatment. Thus, long-term beta(2)-AR activation might results in endothelial dysfunction.93Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)