The Role Of ?CaMK? In The Medial Refrontal Cortex Related Cognitive Functions And Synaptic Plasticity

The medial prefrontal cortex(mPFC)is known to regulate a variety of higher cognitive functions,such as working memory and behavioral flexibility.As a major downstream molecule in the NMDA receptor signaling,alpha-Ca2+/calmodulin-dependent protein kinase II(aCaMKII)is crucial for hippocampus-related memory and hippocampal long-term potentiation(LTP).aCaMKII is also expressed abundantly in the mPFC,but the role of aCaMKII in mPFC-related cognitive functions and mPFC synaptic plasticity remains elusive.In the present study,using aCaMKII-F89G transgenic mice created by employing chemical-genetic technique,in which exogenous aCaMKII-F89G was overexpressed in the forebrain,and aCaMKII-shRNA interfering mice created by using shRNA interference technique,in which aCaMKII was decreased in the mPFC,we examined the effects of up-regulated and down-regulated aCaMKII on mPFC-related cognitive functions and mPFC synaptic plasticity,respectively.The main results are as follows:1.The effects of up-regulated ?CaMKII on mPFC-related cognitive functions and mPFC synaptic plasticity1)The working memory was normal,but behavioral flexibility was impaired in?CaMKII-F89G transgenic mice.aCaMK?-F89G transgenic mice showed normal working memory in T-maze delayed non-match to place task and modified water maze task,but exhibited deficit in behavioral flexibility in Y-water maze arm reversal task.2)The NMDAR-dependent LTP and depotentiation were normal,but NMDAR-dependent long-term depression(LTD)was impaired in the mPFC of?CaMKII-F89G transgenic mice.Golgi staining and in vitro electrophysiological analysis showed normal spine density,normal basal synaptic transmission,normal sEPSC\mEPSC,normal NMDAR/AMPAR ratio,normal LTP and depotentiation,but selectively impaired NMDAR-dependent LTD in the mPFC of aCaMKII-F89G transgenic mice.3)aCaMKII-F89G transgenic mice showed deficits in the GluA1-Ser845 dephosphorylation and AMPAR internalization during LTD,and these deficits were caused by impaired PP1 activity and increased stargazin expression in synaptic membrane.To further investigate the mechanisms underlying aCaMK? up-regulation impairing NMDAR-dependent LTD,using Western blotting,we examined the expression of relevant molecules in mPFC membrane protein after Chem-LTD induction.We found that the GluAl-Ser845 dephosphorylation and AMPAR internalization were impaired in aCaMKII-F89G transgenic mice and these deficits were due to the decreased PP1 activity and increased stargazin expression during NMDAR-dependent LTD expression.2.The effects of down-regulated aCaMKII on mPFC-related cognitive functions and mPFC synaptic plasticity1)The expression of ?CaMKII in the mPFC was significantly reduced in aCaMKII-shRNA interfering mice.Real-time PCR and Western blotting results displayed respectively that,compared with control group,the aCaMKII mRNA expression and protein expression in aCaMKII-shRNA mice were significantly down-regulated.2)The working memory and behavioral flexibility were both impaired in aCaMKII-shRNA interfering mice.Modified water maze and Y-maze spontaneous alternation task showed that the working memory in aCaMKII-shRNA mice was impaired.Furthermore,attentional set-shifting task showed that the behavioral flexibility of aCaMKII-shRNA mice was also impaired.3)Both the NMDAR-dependent LTP and LTD in the mPFC were impaired in ?CaMKII-shRNA interfering miceIn vitro field potential recording analysis showed that,compared with that in control mice,the NMDAR-dependent LTP and LTD in aCaMKII-shRNA mice were both significantly impaired,suggesting that down-regulation of aCaMKII in the mPFC impairs bidirectional synaptic plasticity.4)Both the AMPAR function and NMDAR function were impaired in ?CaMKII-shRNA interfering miceTo further investigate the mechanisms underlying aCaMKII down-regulation impairing bidirectional synaptic plasticity,by using whole cell patch clamp recording technique,we found that the resting membrane potential(RP),input resistance,number of spikes,and the frequency of sEPSC\mEPSC were all normal,but the amplitude of sEPSC\mEPSC and the NMDAR/AMPAR ratio were significantly decreased in aCaMKII-shRNA mice,suggesting that the aCaMKII down-regulation impairs the function of AMPAR and NMDAR.Taken together,we found that(1)Forebrain up-regulation of aCaMKII did not affect the working memory,basal synaptic transmission,NMDAR-dependent LTP and depotentiation in the mPFC,but selectively impaired the behavioral flexibility and NMDAR-dependent LTD,suggesting that there is a close relationship between NMDAR-dependent LTD and behavioral flexibility.To further study the mechanisms of these deficits in flexibility and NMDAR-dependent LTD,we found that up-regulation of aCaMKII impaired GluAl-Ser845 dephosphorylation and AMPAR internalization during LTD,and these impairments may be due to the decreased PP1 activity and increased stargazin expression caused by aCaMKII up-regulation.(2)Down-regulation of aCaMKII in the mPFC impaired the working memory and behavioral flexibility,impaired the NMDAR-dependent LTP and LTD in the mPFC.To further study the mechanisms of these impairments,we found that down-regulation of aCaMKII decreased the function of AMPAR and NMDAR.Our results for the first time indicate that aCaMKII plays an important role in behavioral flexibility and NMDAR-dependent LTD in the mPFC,and provide a new evidence for "NMDAR-dependent LTD may be the cellular mechanism of behavioral flexibility".