Springs are abundant, ecologically, culturally, and socio-economically productive, and highly ecologically interactive ecosystems, providing baseflow for many rivers on Earth, but are intensively used and threatened by human activities. Assessment of >4,000 springs in western North America highlighted variation and trade-offs of springs biodiversity with their ecosystem structure and function in relation to biogeographic principles and climate change. Springs are often hotspots of rare, endemic and endangered species, but display several patterns contrary to normal biogeographic species-area relationships. 1) Larger, higher-elevation springs support greater richness of vascular wetland plants, but diversity and wetland function decline dramatically with elevation and latitude, contrary to the latitudinal diversity gradient, and partially in response to increasing pH. 2) Springs commonly serve as ambush points for predators, increasing their ecological interactivity. 3) Small, remote springs are sought by actively dispersing and migrating species. 4) Vegetation inventories of 50-100 springs contained 20-25% of the regional vascular plant species. 5) Native and non-native species richness are commonly positively related in springs. 6) Contrary to assemblage nestedness theory, smaller springs do not uniformly support the most common, widely distributed wetland species. Overall, spring ecosystem interactivity is often amplified by environmental severity, with habitat area negatively related to both biodiversity and springs keystone ecosystem functions. Climate change and associated human impacts on groundwater through extractive consumption and pollution widely threaten springs and humanity’s last reserve of fresh water; however, the majority of impacts are local, and therefore relatively easily mitigated if the supporting aquifer is relatively intact. We argue for a pan-scale springs conservation strategy that, along with mitigation of climate change, includes rehabilitation of spring ecological functionality. A relatively minor amount of conservation attention at springs can contribute substantially to regional and global biodiversity conservation.