Significance We custom synthesize a fluorescent ligand and use highly sensitive microscopy methods to show that receptors may be targeted to distinct sites of a cell. We provide microscopic images of the localization of β ₁ ‐adrenergic receptors (β ₁ -ARs) and β ₂ ‐adrenergic receptors (β ₂ ‐ARs), main drivers of the contractility of the heart, in heart muscle cells. We show that β ₂ ‐ARs are exclusively targeted to specific structures in these cells, the T-tubules, while β ₁ -ARs are found at the entire cell surface and in T-tubules. Such differential localization may explain their different physiological functions, even though they trigger the same biochemical signal, i.e. an increase in cyclic adenosine monophosphate (cAMP). Thus, receptor function may be determined not only by their biochemical signals, but also by their specific cellular localization.