Uruguayan Southern lagoons exhibit high Holocene resolution paleoenvironmentalpaleoclimatic records for inferring long-term regional changes. The multiproxy analysis of three sediment cores allowed to recognize Holocene climatic variability from the paleolimnological record of Peña lagoon over the last 2,458 yr BP. Four main stages were identified throughout the record. The first (2,458-1,500 cal yr BP) was characterized as a shallow mesoeutrophic system with high abundances of aerophilic benthic species (i.e., Hantzschia amphioxys, Nitzschia brevissima, Frustulia sp., Luticola goeppertiana), epiphytic taxa (i.e Epithemia adnata, Eunotia spp., Rophalodia gibba) and planktonic taxa (i.e. Aulacoseira ambigua and A. granulata). The second stage showed a noticeable change in the diatom assemblage dominated by fresh-brackish benthic species Staurosira construens, but also fluctuations in the abundance of Aulacoseira ambigua and A. granulata, which indicates the occurrence of temperate to cold and semiarid climatic conditions, including intervals of high rainfall. The core chronology allowed us to ascribe this stage to the Little Ice Age (LIA). The third stage, post 390 cal yr BP, showed the highest proportion of freshwater planktonic species throughout the entire core, thus indicating the development of a eutrophic system under relatively warm and wet conditions, which were assigned to the Current Warm Period. After ca. 1962 AD, a sharp increase in the abundance of epiphytic species (i.e., Cocconeis placentula, Eunotia spp, Epithemia adnata and Encyonema minutum) highlights the onset of the fourth stage, which was characterized by littoral expansion and consequently, the proliferation of associated macrophytes due to anthropogenic impacts.