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S100β, a calcium-binding brain specific protein, may affect both brain development and hippocampal long-term potentiation. S100β levels are elevated in Down syndrome (DS), and the gene for S100β is located on chromosome 21, which is duplicated in DS. To test the hypothesis that elevated levels of S100β cause behavioral alterations in a mammalian system, 3 transgenic mouse lines with multiple copies of the human gene for S100β were derived and behaviorally tested. The spontaneous alternation behavior of transgenic and normal littermate mice were compared in a T maze during a 15-trial test. The overall alternation rate was found to be significantly decreased in the transgenic mice compared with their normal littermates. The S100β transgenic mouse model offers one of the first opportunities to investigate the relation between overexpression of a human chromosome 21 gene product and abnormal behavior and brain function.