The Andean forests are facing severe fragmentation and loss of connectivity due to agricultural expansion, infrastructure development, and urbanization. By integrating remote sensing data and analyzing forest connectivity from 1985 to 2022 across diverse spatial extents, we employed a graph theory-based Equivalent Connected Area (ECA) index and its relative variation (dECA) to assess habitat connectivity based on accessibility for species with varying dispersal abilities. The extent of forest cover decreased by 43% over the period from 1985 to 2022, while forest connectivity declined by 85%. These findings indicate a concerning reduction in connectivity, particularly in ecologically and socially significant regions such as the Eje Cafetero, the southern Andes, and the Cundiboyacense Plateau. The fragmentation of Andean forests has a detrimental impact on species such as the Andean bear and puma, restricting their mobility and hindering their conservation. The utilization of remote sensing technology could facilitate the identification of specific regions that require urgent attention for the maintenance of Andean ecosystems in Colombia. Furthermore, this approach can assist in the delineation of crucial areas where restoration or conservation initiatives may be most effective in enhancing forest connectivity.