Behavioral and Physiological Significance of PKC-mediated Phosphorylation of the GluN2A C-terminus

Activity-dependent plasticity in N-methyl-D-aspartate receptor containing synapses could be regulated by phosphorylation of specific amino acids in the C-terminal domain of the receptor subunits. Previous experiments done in our lab established that the phosphorylation of the serines (S1291 and S1312) directly by PKC and tyrosines (Y1292 and Y1387) indirectly via PKC activation of Src positively modulate the receptor currents. To understand the behavioral and physiological significance of those sites in vivo, the Grin2aDeltaPKC mouse expressing GluN2A with those four mutated amino acids (S1291A, S1312A, Y1292F and Y1387F) was generated using homologous recombination. The Grin2aDeltaPKC and WT mice were similar in their body weight, and general activity. They also have similar expression levels of Grin2a mRNA, and GluN2A protein. The Grin2aDeltaPKC mice alternated above chance levels in a four-arm and Y maze spontaneous alternation tasks, while they alternated at levels lower than WT mice only in the Y maze. The mutants also have decreased alternation in a non-reinforced T maze alternation task. Thus, the mutant mice may have a mild spatial memory deficit. When these mutant mice were subjected to anxiety-associated tasks, they exhibited reduced anxiety-related behaviors such as increased time spent in the center of an open field, increased time in lit side of light/dark box, and increased entries and dwell times in the open arms of an elevated plus maze. Immunostaining for Fos in the hippocampus after exposure of the animals to novel environments shows region specific differences between the mutants and WT mice. There was no increase in Fos levels in mutants after exposure to novel environments in CA1 and CA3 compared to home-cage Fos levels, in contrast, Fos levels increased in the WT mice in CA1, CA3 and dentate gyrus. When the Schaffer collateral-CA1 synapses of mutant hippocampal synapses were stimulated using a theta-burst protocol, it was found that there was no impairment in LTP. Also, the mutant mice showed no significant differences in input-output curves and paired-pulse facilitation. Taken together, these results suggest that PKC-mediated phosphorylation of at least one of those four sites regulates NMDAR-mediated signaling that modulates anxiety, and spatial working memory.