Target delivery of large foreign materials to cells requires transient permeabilization of the cell membrane without toxicity. Giant unilamellar vesicles (GUVs) mimic the phospholipid bilayer of the cell membrane and are also useful drug delivery vehicles. Controlled increase of the permeability of GUVs is a delicate balance between sufficient perturbation for the delivery of the GUV contents and damage to the vesicles. Here we show that photoacoustic waves can promote the release of FITC-dextran or GFP from GUVs without damage. Real-time interferometric imaging offers the first movies of photoacoustic wave propagation and interaction with GUVs. The photoacoustic waves are seen as mostly compressive half-cycle pulses with peak pressures of ~ 1 MPa and spatial extent FWHM ~ 36 µm. At a repetition rate of 10 Hz, they enable the release of 25% of the FITC-dextran content of GUVs in 15 min. Such photoacoustic waves may enable non-invasive targeted release of GUVs and cell transfection over large volumes of tissues in just a few minutes.