Osteocytes interconnect with each other forming an intricate cell network within the mineralized bone matrix. One important function of the osteocyte network is the mechano-regulation of bone remodeling, where a possible mechanism includes the fluid flow through the porosity housing the cell network - the osteocyte lacuno-canalicular network (OLCN). In our study the OLCN in human osteons was three-dimensionally imaged with the aim to obtain a quantitative description of the canalicular density and spatial variations of this quantity within osteons. The topology of the OLCN was determined by first staining the bone samples with rhodamine, then imaging the OLCN with confocal laser scanning microscopy and finally using image analysis to obtain a skeletonized version of the network for further analysis. In total 49 osteons were studied from the femoral cortical bone of four different middle-aged healthy women. The mean canalicular density given as length of the canaliculi in a unit volume was 0.074 ± 0.015 μm/μm³ (corresponding to 74 km/cm³). No correlation was found between the canalicular density and neither the size of the osteon nor the volume fraction occupied by osteocyte lacunae. Within osteons the canalicular density varied substantially with larger regions without any network. On average the canalicular density decreases when moving from the Haversian canal outwards towards the cement line. We hypothesize that a decrease in accessible canaliculi with tissue age as a result of micropetrosis can reduce the local mechanosensitivity of the bone. Systematic future studies on age- and disease-related changes on the topology of the OLCN have to demonstrate the diagnostic potential of the presented characterization method.