Toxic heavy metals in air, soil, and water are global problems that are a growing threat to all living kinds in the environment. Exposure to toxic heavy metals can result in cytotoxic, carcinogenic and mutagenic effects. Mutagenesis is an indicative of genetic instability and can be assayed using microsatellite instability (MSI). The aim of the present study is to investigate; based on the rat model, the effects of oral acute and sub-chronic doses of four heavy metals (cadmium chloride, lead acetate trihydrate, arsenic trioxide and copper sulphate pentahydrate)on microsatellite instability at D6mit3, D9mit2 and D15Mgh1 loci, which are located in three different common fragile sites (6q21, 9q32-q33 and 15p14, respectively), within rat genome. In the acute and sub-chronic studies, the MSI were observed in the three studied loci and was in the form of addition or deletion of base pair nucleotides. These finding may indicate the sensitivity of microsatellite sequences located at the fragile sites and the sensitivity of the simple sequence repeats (SSRs) assay for the detection of small variations in DNA sequence.