Abstractα₁-adrenergic receptors (α₁-ARs) play critical roles in the cardiovascular and nervous systems where they regulate blood pressure, cognition, and metabolism. However, the lack of specific agonists for all α₁ subtypes has limited our understanding of the physiological roles of different α₁-AR subtypes, and led to the stagnancy in agonist-based drug development for these receptors. Here we report cryo-EM structures of α_1A-AR in complex with heterotrimeric G-proteins and either the endogenous common agonist epinephrine or the α_1A-AR-specific synthetic agonist A61603. These structures provide molecular insights into the mechanisms underlying the discrimination between α_1A-AR and α_1B-AR by A61603. Guided by the structures and corresponding molecular dynamics simulations, we engineer α_1A-AR mutants that are not responsive to A61603, and α_1B-AR mutants that can be potently activated by A61603. Together, these findings advance our understanding of the agonist specificity for α₁-ARs at the molecular level, opening the possibility of rational design of subtype-specific agonists.