The goal of our studies was to compare different measuring methods of glutamine, glutamate and GABA of rat hippocampus used for study of pathogenesis of autism. The methods under consideration were: in vivo MRS and two in vitro ones, NMR and HPLC. Univariate statistical analysis of ratios of tested amino acids with respect to glutamate concentration was performed using General Linear Model. This demonstrated statistically significant differences between the results from three methods for both, glutamine and GABA ratios. OPLS-DA analysis allowed build models for differentiation of two animal models of disease and control group in NMR and HPLC.