Mountain regions are fragile ecosystems and often host remarkably rich biodiversity, and thus they are especially under threat from ongoing global changes. Located in the Eastern Alps, Trentino–South Tyrol is bioculturally diverse but an understudied region from an ethnobotanical perspective. We explored the ethnomedicinal knowledge of the area from a cross-cultural and diachronic perspective by conducting semi-structured interviews with 22 local inhabitants from Val di Sole (Trentino) and 30 from Überetsch–Unterland (South Tyrol). Additionally, we compared the results with ethnobotanical studies conducted in Trentino and South Tyrol over 25 years ago. The historical comparison revealed that about 75% of the plants currently in use were also used in the past in each study region. We argue that the adoption of “new” medicinal species could have occurred through printed and social media and other bibliographical sources but may also be due to limitations in conducting the comparison (i.e., different taxonomic levels and different methodologies). The inhabitants of Val di Sole and Überetsch–Unterland have shared most medicinal plants over the past few decades, yet the most used species diverge (perhaps due to differences in local landscapes), and in South Tyrol, people appear to use a higher number of medicinal plants, possibly because of the borderland nature of the area.