Species can have broad altitudinal distributions and their characteristics may vary throughout distribution ranges. Ploidy level can vary along environmental gradients, such as altitudinal ones. However, the relationships between ploidy level and altitude may be diverse. The karyotype allows us to know the structural and quantitative characteristics of the chromosomal pairs and to relate these characteristics to the environment where the species occur. Species of the Cactaceae family can be found from sea level to 4500 m above sea level (asl). The greatest species richness is found in mountain areas, where species occupy wide altitudinal ranges. Gymnocalycium belongs to the subfamily Cactoideae; it is a genus endemic to southern South America and its main center of diversity is found in the mountain ranges of central and northern Argentina. The basic chromosome number for Cactaceae is x = 11, with polyploidy being the main existing variation. The aim of this work was to analyze cytogenetic variables in populations of four species of the genus Gymnocalycium (G. andreae, G. erinaceum, G. monvillei, and G. mostii) along their altitudinal distribution. Idiograms were constructed using HCl/Giemsa, CMA/DAPI fluorescent chromosomal banding and fluorescent in situ hybridization (FISH). The karyotypes of the analyzed populations of each species were symmetric and showed little variation in size and symmetry. All the populations of G. andreae, G. erinaceum and G. mostii were found to be diploid and had the greatest distribution; G. monvillei was found to be tetraploid in all the populations analyzed. CMA+/DAPIbands associated with secondary constrictions (NORs) were detected in all the populations of all the species. The cytogenetic characteristics of the studied species were constant along the altitudinal gradients, showing that they can occur at different altitudes without major cytogenetic modifications.