This paper studies the impacts of Madrid-Seville High-Speed Rail (HSR) on population growth and land cover change in the five HSR connected cities - Madrid, Ciudad Real, Puertollano, Cordoba, and Seville - at both regional and local level. The analysis period ranges from 1991 to 2006. The study finds that, at regional level, the population growth and land development process concentrate mostly towards the two largest cities, Madrid and Seville, while other smaller HSR served cities are also benefited by HSR. At local level, the impacts of HSR are more diverse. The process of population redistribution and land development in each city varies largely. Among all evidences, HSR contributes the most to Ciudad Real, with booming population increases and urban development. In addition, younger people are also attracted to reside in this area. To study the accessibility impacts of HSR on the development of new urban areas in each HSR city, binary probit models incorporating the change of accessibility, population, and neighborhood situations are adopted. The results suggest that, the land development in smaller cities can be majorly ascribed to the improvement of regional accessibility and population growth. However, to explain the urban development process in Madrid and Seville, the inputs with only accessibility and population are not sufficient.