Synaptic Localization of NMDA Receptor and Shank Protein in Hippocampal Neuron in Vitro / 대한해부학회지

ABSTRACT

The existence of NMDA receptor and a new organizer protein, Shank, in the postsynaptic density was studied with the cultured hippocampal neurons using by double immunofluorescence method. The hippocampi from embryonic 18 days were dissected and hippocampal neurons were obtained from dissociated hippocampi with 0.25% trypsin and 0.1% DNase in PBS. The hippocampal neurons were plated with density 3,600/cm2 on the poly-L-lysine coated coverglass and cultured 37degrees C, 5% CO2 incubator for 5 weeks. The N2 supplemented MEM was used as a culture medium. Following results are obtained from experiments 1. The 3~5 minor processes from the cell body of hippocampal neurons were observed at 20 hr in vitro. One of the minor processes was elongated and looked like an axon, and another minor processes showed dendritic branching pattern with slender in thickness. 2. The excitatory NMDA receptor colocalized with PSD-95 which is the postsynaptic density protein. The presynaptic protein, synapsin 1, was closely apposed with PSD-95. 3. Shank which is an organizer protein colocalize with NMDA receptor/PSD-95 complex in the postsynaptic density. Shank proteins may be concerned with the cluster formation of NMDA receptor/PSD-95 in the postsynaptic membrane.