  Structural microvascular alterations and dysfunction serve as key disease
indicators of cancer, diabetes, ischemic stroke, neurodegenerative disorders
and many other conditions. In vivo visualization of the microvasculature has
traditionally been restricted to millimeter scale depths accessible with
optical microscopy. Optoacoustic imaging has enabled breaking through the
barrier imposed by light diffusion to map functional hemodynamic parameters in
deep-seated vessels, but diffraction and dispersion of ultrasound waves in
heterogeneous living tissues prevents reaching capillary resolution. Herein, we
demonstrate three-dimensional microangiography of deep mouse brain beyond the
acoustic diffraction limit (<20$\mu$m resolution) through the intact scalp and
skull via optoacoustic localization of sparsely-distributed highly absorbing
microparticles. This was enabled by devising 5$\mu$m sized extremely absorbing
dichloromethane microdroplets exhibiting four orders of magnitude higher
optical absorption than red blood cells at near-infrared wavelengths, thus
facilitating in vivo compatibility and single particle sensitivity in the
presence of highly absorbing blood background. Accurate mapping of the blood
flow velocity within microvascular structures was also facilitated by the high
3D frame rate of the optoacoustic tomography system. We further show that the
detected optoacoustic signal intensities from the localized particles may serve
for estimating the light fluence distribution within optically heterogeneous
tissues, a long-standing quantification challenge in biomedical optics. Given
the intrinsic sensitivity of optoacoustics to various functional, metabolic and
molecular events in living tissues, this new approach paves the way for
non-invasive deep-tissue microscopic observations with unrivaled resolution,
contrast and speed.
