Ultra-thin holographic endo-microscopes exploiting multi-mode optical fibres (MMF) have recently emerged as powerful tools, allowing us to explore the deepest regions inside living mouse brains with negligible disruption and diffraction-limited resolution [1,2,3]. Recently published work representing current state-of-the-art holographic endo-microscopy deals with detailed imaging of neuronal structures, functional monitoring of red blood cell velocity and calcium dynamics located in the deepest regions of anaesthetized mice [4]. In addition to technological development, the transition to imaging awake animals is a natural step forward. Here, we present our successful imaging sessions of neuronal activity in awake head-fixed mice using a custom-engineered tip of our optical fibre-based probes. This feature, combined with a novel protocol for awake animals imaging, allows us to obtain sub-cellularly resolved calcium transients correlated with behavior or stimuli.ReferencesOhayon, S., et al. Minimally invasive multimode optical fiber microendoscope for deep brain fluorescence imaging. Biomedical Optics Express 9, 1492–1509 (2018).Vasquez-Lopez, S. A. et al. Subcellular spatial resolution achieved for deep-brain imaging in vivo using a minimally invasive multimode fiber. Light-Science & Applications 7, 110 (2018).Turtaev, S. et al. High-fidelity multimode fibre-based endoscopy for deep brain in vivo imaging. Light-Science & Applications 7, 92 (2018).Stibůrek, M. et al. 110μm thin endo-microscope for deep-brain in-vivo observations of neuronal connectivity, activity and blood flow dynamics. Nature Communications, (2023).