The Phenotype Evaluation Of PKC Rat Model And The Study Of The Electrophysiological Mechanism Of Prrt2 Regulating Neuronal Excitability

It's well known that PRRT2 mutants underlie many nerve diseases such as seizures,paroxysmal kinesigenic choreoathetosis.Most mutations lead to abnormal protein expression of PRRT2,indicating a loss of function mechanism.But the physiologic function of PRRT2 and pathogenesis of PKC remain obscure.In this research,we measured the time-space expression pattern of PRRT2 in SD rats by Western blotting,finding that it is selectively expressed in the nerve system like cerebral cortex,cerebellum and hippocampus.And it's barely expressed in the embryo stage,but shows a rapid rise in the nervous system of postnatal-7-days rats,and reaches peak level at PD-14-28,after which remains a stable expression level.Firstly,to uncover the molecular mechanism of PKC,we established prrt2 truncated mutant rat model by CRISPR-Cas9 gene editing technology.After that,we videotaped prrt2-/-rats of different ages and successfully recorded spontaneous attacks.These prrt2-/-rats not only showed apparently decline in motor coordination assessed by beam balance test and coat hanger test,but also have poor tolerance to PTZ.While other behaviors of prrt2-/-rats are normal,which agrees with the clinical data.Next,the results from transmission electron microscope showed that the readily release pool size of prrt2-/-rats was larger than that of prrt2+/+ cohorts.To figure out if the electrophysiology of prrt2-/-rats changed,we recorded mEPSC,mIPSC of the pyramidal cell in the cerebral cortex,finding that both frequency and amplitude of mEPSC were significantly increased in prrt2-/-rats,while amplitude of mIPSC was decreased and the ratio of mEPSC/mIPSC was significantly increased.In hippocampus,eEPSC amplitude was increased due to the larger RRP size,mIPSC frequency was increased.In conclusion,PRRT2 plays an important role of transmitter release machine,the loss-of-function mutations could lead to larger RRP size and then abnormal synaptic transmission,resulting in the imbalance between excitatory and inhibitory neurons and finally triggering PKC.