Dynamic Changes in Neurexins' Alternative Splicing: Role of Rho-Associated Protein Kinases and Relevance to Memory Formation

The three neurexins genes (NRXN1/2/3) encode polymorphic synaptic membrane proteins that are involved in cognitive functioning. Neurexins' selectivity of function is presumably conferred through differential use of 2 promoters and 5 alternative splicing sites (SS#1/2/3/4/5). In day-old rat brain neurons grown in culture, activation (depolarization) induces reversible, calcium dependent, repression of NRXN2 alpha SS#3 insert. The effects of depolarization on NRXN1/2/3 alpha splicing and biochemical pathways mediating them were further studied in these neurons. NRXN1/2/3 alpha splicing in the course of memory formation in vivo was also explored, using fear conditioning paradigm in rats in which the animals were trained to associate an aversive stimulus (electrical shock) with a neutral context (a tone), resulting in the expression of fear responses to the neutral context. In the cultured neurons depolarization induced, beside NRXN2a SS#3, repression of SS#3 and SS#4 exons in NRXN3 alpha but not NRXN1 alpha. The repressions were mediated by the calcium/protein kinase C/Rho-associated protein kinase (ROCK) pathway. Fear conditioning induced significant and transient repressions of the NRXN1/2/3 alpha SS#4 exons in the rat hippocampus. ROCK inhibition prior to training attenuated the behavioral fear response, the NRXN1/2/3 alpha splicing repressions and subsequent recovery and the levels of excitatory (PSD95) and inhibitory (gephyrin) synaptic proteins in the hippocampus. No such effects were observed in the prefrontal cortex. Significant correlations existed between the fear response and hippocampal NRXN3 alpha and NRXN2 alpha SS#4 inserts as well as PSD95 protein levels. Hippocampal NRXN1 alpha SS#4 insert and gephyrin levels did not correlate with the behavioral response but were negatively correlated with each other. These results show for the first time dynamic, experience related changes in NRXN1/2/3 alpha alternative splicing in the rat brain and a role for ROCK in them. Specific neurexins' transcripts may be involved in synaptic remodeling occurring at an intermediate (hours) time scale in the course of memory formation.