Meng W, Ma J, Ayata C, Hara H, Huang P L, Fishman M C, Moskowitz M A
Department of Neurosurgery and Neurology, Massachusetts General Hospital, Charlestown, USA.
Am J Physiol. 1996 Sep;271(3 Pt 2):H1145-50. doi: 10.1152/ajpheart.1996.271.3.H1145.
We used mice with deletions in either the endothelial nitric oxide synthase (eNOS) or neuronal NOS (nNOS) gene to investigate the role of eNOS and nNOS in acetylcholine (ACh)-induced relaxation of pial arterioles (20-30 microns). Pial arteriolar diameter was measured by intravital microscopy through a closed cranial window, and NOS activity was determined by the conversion of [3H]arginine to [3H]citrulline in subjacent cortex. ACh superfusion (1, 10 microM) caused atropine-sensitive dose-dependent arteriolar dilation in all three mouse strains. At 10 microM, increases of 20 +/- 2, 31 +/- 3, and 23 +/- 3% were recorded in wild-type (n = 25), nNOS mutant (n = 15), and eNOS mutant (n = 20) mice, respectively. NG-nitro-L-arginine (L-NNA, 1 mM) superfusion inhibited cortical NOS activity by > 70% and abrogated the response in wild-type mice while blocking the dilation by approximately 50% in eNOS mutant and nNOS mutant mice. Only in the eNOS mutant did tetrodotoxin (TTX) superfusion (1 microM) attenuate ACh-induced dilation (n = 6). The residual dilation after L-NNA in eNOS mutant mice could be blocked completely by TTX-plus L-NNA. Our findings indicate that 1) ACh dilates pial arterioles of wild-type mice by NOS-dependent mechanisms as reported in other species, 2) the response in nNOS mutant mice resembles the wild-type response except for enhanced dilation to ACh and reduced L-NNA sensitivity, and 3) surprisingly, the response in eNOS mutant mice is partially NOS dependent and attenuated by both TTX and L-NNA. Because nNOS is constitutively expressed in eNOS mutants, these findings coupled with the TTX results suggest that an nNOS-dependent mechanism may compensate for the chronic loss of eNOS activity after targeted gene disruption.
我们使用内皮型一氧化氮合酶（eNOS）或神经元型一氧化氮合酶（nNOS）基因缺失的小鼠，来研究eNOS和nNOS在乙酰胆碱（ACh）诱导的软脑膜小动脉（20 - 30微米）舒张中的作用。通过闭合颅骨视窗利用活体显微镜测量软脑膜小动脉直径，并通过将[3H]精氨酸转化为[3H]瓜氨酸来测定相邻皮质中的一氧化氮合酶（NOS）活性。ACh灌注（1、10微摩尔）在所有三种小鼠品系中均引起对阿托品敏感的剂量依赖性小动脉舒张。在10微摩尔时，野生型（n = 25）、nNOS突变型（n = 15）和eNOS突变型（n = 20）小鼠的小动脉舒张分别增加了20±2%、31±3%和23±3%。NG - 硝基 - L - 精氨酸（L - NNA，1毫摩尔）灌注使皮质NOS活性抑制>70%，消除了野生型小鼠的反应，同时在eNOS突变型和nNOS突变型小鼠中使舒张反应阻断约50%。仅在eNOS突变型小鼠中，河豚毒素（TTX）灌注（1微摩尔）减弱了ACh诱导的舒张（n = 6）。L - NNA处理后eNOS突变型小鼠的残余舒张可被TTX加L - NNA完全阻断。我们的研究结果表明：1）如在其他物种中所报道的，ACh通过依赖于NOS的机制使野生型小鼠的软脑膜小动脉舒张；2）nNOS突变型小鼠的反应类似于野生型反应，只是对ACh的舒张增强且对L - NNA的敏感性降低；3）令人惊讶的是eNOS突变型小鼠的反应部分依赖于NOS，且被TTX和L - NNA均减弱。由于nNOS在eNOS突变体中组成性表达，这些发现与TTX结果表明，一种依赖于nNOS的机制可能补偿了靶向基因破坏后eNOS活性的慢性丧失。
