Holst B D, Vanderklish P W, Krushel L A, Zhou W, Langdon R B, McWhirter J R, Edelman G M, Crossin K L
Department of Neurobiology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Proc Natl Acad Sci U S A. 1998 Mar 3;95(5):2597-602. doi: 10.1073/pnas.95.5.2597.
To study regulation in vivo of the promoter for the neural cell adhesion molecule, N-CAM, we have used homologous recombination to insert the bacterial lacZ gene between the transcription and translation initiation sites of the N-CAM gene. This insertion disrupts the gene and places the expression of beta-galactosidase under the control of the N-CAM promoter. Animals homozygous for the disrupted allele did not express N-CAM mRNA or protein, but the pattern of beta-galactosidase expression in heterozygous and homozygous embryos was similar to that of N-CAM mRNA in wild-type animals. The homozygotes exhibited many of the morphological abnormalities observed in previously reported N-CAM knockout mice, with the exception that hippocampal long-term potentiation in the Schaffer collaterals was identical in homozygous, heterozygous, and wild-type animals. Heterozygous mice were used to examine the regulation of the N-CAM promoter in response to enhanced synaptic transmission. Treatment of the mice with an ampakine, an allosteric modulator of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors that enhances normal glutamate-mediated synaptic transmission, increased the expression of beta-galactosidase in vivo as well as in tissue slices in vitro. Similar treatments also increased the expression of N-CAM mRNA in the heterozygotes. The effects of ampakine in slices were strongly reduced in the presence of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), an AMPA receptor antagonist. Taken together, these results indicate that facilitation of AMPA receptor-mediated transmission leads to activation of the N-CAM promoter and provide support for the hypothesis that N-CAM synthesis is regulated in part by synaptic activity.
为了研究神经细胞黏附分子（N-CAM）启动子在体内的调控机制，我们利用同源重组技术，将细菌lacZ基因插入到N-CAM基因的转录起始位点和翻译起始位点之间。这种插入破坏了该基因，并使β-半乳糖苷酶的表达受N-CAM启动子的控制。纯合缺失等位基因的动物不表达N-CAM mRNA或蛋白质，但杂合和纯合胚胎中β-半乳糖苷酶的表达模式与野生型动物中N-CAM mRNA的表达模式相似。纯合子表现出许多在先前报道的N-CAM基因敲除小鼠中观察到的形态学异常，但不同的是，纯合子、杂合子和野生型动物的海马体中沙费尔侧支的长时程增强效应是相同的。杂合小鼠被用于研究N-CAM启动子对增强的突触传递的响应调控。用一种安帕金（一种α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）受体的变构调节剂，可增强正常的谷氨酸介导的突触传递）处理小鼠，可增加体内以及体外组织切片中β-半乳糖苷酶的表达。类似的处理也增加了杂合子中N-CAM mRNA的表达。在存在AMPA受体拮抗剂6-氰基-7-硝基喹喔啉-2,3-二酮（CNQX）的情况下，安帕金在切片中的作用被大大减弱。综上所述，这些结果表明，AMPA受体介导的传递促进导致N-CAM启动子的激活，并为N-CAM合成部分受突触活动调控这一假说提供了支持。
