Lohof A M, Quillan M, Dan Y, Poo M M
Department of Biological Sciences, Columbia University, New York, New York 10027.
J Neurosci. 1992 Apr;12(4):1253-61. doi: 10.1523/JNEUROSCI.12-04-01253.1992.
The possible role of cyclic nucleotides as second messengers mediating growth cone turning was studied by producing an asymmetric distribution of cyclic nucleotides across the growth cone. A repetitive pulse application method was developed to produce microscopic chemical gradients near the growth cone of embryonic Xenopus neurons in cell culture. When picoliters of a solution containing 20 mM dibutyryl cAMP (dB-cAMP), a membrane-permeable analog of cAMP, were repetitively ejected from a micropipette near the growth cone, neurite growth was consistently directed toward the pipette. Theoretical analysis of the diffusion gradient showed that the neurite is capable of detecting a 10% difference in dB-cAMP concentration across the growth cone. Similar responses were also observed using gradients of the phosphodiesterase inhibitor isobutylmethylxanthine, or of forskolin, which activates adenylate cyclase. Dibutyryl cGMP, however, produced no significant turning. These results suggest that a cytoplasmic gradient of cAMP across the growth cone is sufficient to initiate its turning response, and that cAMP in the growth cone could serve as a second messenger in mediating the action of extracellular guidance cues.
通过在生长锥上产生环核苷酸的不对称分布，研究了环核苷酸作为介导生长锥转向的第二信使的可能作用。开发了一种重复脉冲施加方法，以在细胞培养的非洲爪蟾胚胎神经元生长锥附近产生微观化学梯度。当从生长锥附近的微量移液器中重复喷出皮升含有20 mM二丁酰环磷酸腺苷（dB-cAMP，一种环磷酸腺苷的膜通透性类似物）的溶液时，神经突生长始终指向移液器。对扩散梯度的理论分析表明，神经突能够检测到生长锥上dB-cAMP浓度10%的差异。使用磷酸二酯酶抑制剂异丁基甲基黄嘌呤或激活腺苷酸环化酶的福司可林的梯度也观察到了类似的反应。然而，二丁酰环磷酸鸟苷（dB-cGMP）并未产生明显的转向。这些结果表明，生长锥上cAMP的细胞质梯度足以启动其转向反应，并且生长锥中的cAMP可以作为第二信使来介导细胞外引导信号的作用。
