Leahy J C, Luo Y, Kent C S, Meiri K F, Vallano M L
Department of Pharmacology, State University of New York Health Science Center, Syracuse 13210-1605.
Neuroscience. 1993 Feb;52(3):563-74. doi: 10.1016/0306-4522(93)90406-6.
Pharmacological and biochemical evidence implicate the Ca2+ and phospholipid-dependent protein kinase C in long-term potentiation. The in vitro hippocampal slice preparation was used to demonstrate redistribution of protein kinase C from cytosol to membrane and protein kinase C-dependent phosphorylation of the presynaptic growth-associated protein-43 substrate following long-term potentiation induction in area CA1. Protein kinase C translocation was assessed using both quantitative immunoblotting with a monoclonal antibody recognizing a common epitope in the alpha and beta isoforms of protein kinase C and Ca2+ and phospholipid-dependent phosphorylation of exogenous histone substrate. Slices examined 5 min after tetanus-induced spike potentiation showed no change in protein kinase C redistribution, whereas slices examined at 15-, 30- and 60-min intervals all showed a similar degree of protein kinase C translocation to membrane, although only at 15 min was the effect statistically significant. Additionally, an increase in protein kinase C-dependent growth-associated protein 43 phosphorylation was observed 10 min after high-frequency stimulation. The translocation of protein kinase C and phosphorylation of growth-associated protein 43 were dependent upon high-frequency (repetitive 400 Hz) afferent stimulation, as no effects were observed in slices receiving low-frequency (1 Hz) or no stimulation. The N-methyl-D-aspartate receptor antagonist, DL-2-amino-5-phosphonovaleric acid (50 microM), inhibited induction of long-term potentiation, redistribution of protein kinase C and phosphorylation of growth-associated protein 43. A significant redistribution of the predominantly presynaptic protein kinase C isoform, protein kinase C-alpha, was also detected 15 min after induction of long-term potentiation using an alpha-isoform-specific monoclonal antibody. These observations support a presynaptic role for protein kinase C and growth-associated protein 43 in the early maintenance phase of LTP, and further suggest that a retrograde messenger produced postsynaptically following N-methyl-D-aspartate receptor activation mediates these effects.
药理学和生物化学证据表明，钙和磷脂依赖性蛋白激酶C与长时程增强有关。利用体外海马脑片制备技术，证明在CA1区诱导长时程增强后，蛋白激酶C从胞质溶胶重新分布到细胞膜，以及突触前生长相关蛋白-43底物的蛋白激酶C依赖性磷酸化。使用识别蛋白激酶C的α和β亚型中共同表位的单克隆抗体进行定量免疫印迹以及对外源组蛋白底物进行钙和磷脂依赖性磷酸化，来评估蛋白激酶C的转位。在强直刺激诱导的峰电位增强后5分钟检查的脑片显示蛋白激酶C重新分布没有变化，而在15、30和60分钟间隔检查的脑片均显示出类似程度的蛋白激酶C向细胞膜的转位，尽管只有在15分钟时该效应具有统计学意义。此外，在高频刺激后10分钟观察到蛋白激酶C依赖性生长相关蛋白43磷酸化增加。蛋白激酶C的转位和生长相关蛋白43的磷酸化依赖于高频（重复400Hz）传入刺激，因为在接受低频（1Hz）或无刺激的脑片中未观察到效应。N-甲基-D-天冬氨酸受体拮抗剂DL-2-氨基-5-磷酸戊酸（50μM）抑制长时程增强的诱导、蛋白激酶C的重新分布和生长相关蛋白43的磷酸化。使用α亚型特异性单克隆抗体还检测到，在诱导长时程增强后15分钟，主要位于突触前的蛋白激酶C亚型蛋白激酶C-α有显著的重新分布。这些观察结果支持蛋白激酶C和生长相关蛋白43在长时程增强的早期维持阶段发挥突触前作用，并且进一步表明，N-甲基-D-天冬氨酸受体激活后在突触后产生的逆行信使介导了这些效应。
