Abstract In the study, rat brain synaptosomes were used, which are an adequate and convenient model for studying presynaptic processes. L Glutamate activity was judged by the change in the intensity of the fluorescent signal, by the change in the cytoplasmic levels of free calcium [Ca2+]in. The study was conducted according to the Weiler method. Synaptosomes were isolated from rat brain using two-stage centrifugation. The entire isolation procedure was carried out at 4°C. To measure the amount of cytosolic Ca2+ synaptosomes were calculated according to the Grinkevich equation. An increase in the concentration of [Ca2+]in caused by L glutamate, primarily due to the activation of membrane permeability, the movement of Ca2+ into the cell, and the release of Ca2+ from intracellular depots. SP-6 competes with L glutamate for the glutamate binding site of NMDA-receptors. L Glutamate partially reduces the effect of SP-6, which may indicate that part of the external calcium also comes under the influence of SP-6 through the open glutamate binding site and in place calcium channels of the NMDA-receptors. It was revealed that SP-6 increases fluorescence and the level of [Ca2+]in, respectively, in synaptic membranes compared to the control. This data gives, about a possible competition between SP-6 and L glutamate for the site of regulation of the opening of ion channels of NMDA-receptors.