BACKGROUND Inhalational anesthetics are known to disrupt PDZ2 domain-mediated protein-protein interactions of the postsynaptic density (PSD)-95 protein. The aim of this study is to investigate the underlying mechanisms in response to early isoflurane exposure on synaptic PSD-95 PDZ2 domain disruption that altered spine densities and cognitive function. The authors hypothesized that activation of protein kinase-G by the components of nitric oxide (NO) signaling pathway constitutes a mechanism that prevents loss of early dendritic spines and synapse in neurons and cognitive impairment in mice in response to disruption of PDZ2 domain of the PSD-95 protein. METHODS Postnatal day 7 mice were exposed to 1.5% isoflurane for 4 h or injected with 8 mg/kg active PSD-95 wild-type PDZ2 peptide or soluble guanylyl cyclase activator YC-1 along with their respective controls. Primary neurons at 7 days in vitro were exposed to isoflurane or PSD-95 wild-type PDZ2 peptide for 4 h. Coimmunoprecipitation, spine density, synapses, cyclic guanosine monophosphate-dependent protein kinase activity, and novel object recognition memory were assessed. RESULTS Exposure of isoflurane or PSD-95 wild-type PDZ2 peptide relative to controls causes the following. First, there is a decrease in PSD-95 coimmunoprecipitate relative to N-methyl-d-aspartate receptor subunits NR2A and NR2B precipitate (mean ± SD [in percentage of control]: isoflurane, 54.73 ± 16.52, P = 0.001; and PSD-95 wild-type PDZ2 peptide, 51.32 ± 12.93, P = 0.001). Second, there is a loss in spine density (mean ± SD [spine density per 10 µm]: control, 5.28 ± 0.56 vs. isoflurane, 2.23 ± 0.67, P < 0.0001; and PSD-95 mutant PDZ2 peptide, 4.74 ± 0.94 vs. PSD-95 wild-type PDZ2 peptide, 1.47 ± 0.87, P < 0.001) and a decrease in synaptic puncta (mean ± SD [in percentage of control]: isoflurane, 41.1 ± 14.38, P = 0.001; and PSD-95 wild-type PDZ2 peptide, 50.49 ± 14.31, P < 0.001). NO donor or cyclic guanosine monophosphate analog prevents the spines and synapse loss and decline in the cyclic guanosine monophosphate-dependent protein kinase activity, but this prevention was blocked by soluble guanylyl cyclase or protein kinase-G inhibitors in primary neurons. Third, there were deficits in object recognition at 5 weeks (mean ± SD [recognition index]: male, control, 64.08 ± 10.57 vs. isoflurane, 48.49 ± 13.41, P = 0.001, n = 60; and female, control, 67.13 ± 11.17 vs. isoflurane, 53.76 ± 6.64, P = 0.003, n = 58). Isoflurane-induced impairment in recognition memory was preventable by the introduction of YC-1. CONCLUSIONS Activation of soluble guanylyl cyclase or protein kinase-G prevents isoflurane or PSD-95 wild-type PDZ2 peptide-induced loss of dendritic spines and synapse. Prevention of recognition memory with YC-1, a NO-independent activator of guanylyl cyclase, supports a role for the soluble guanylyl cyclase mediated protein kinase-G signaling in countering the effects of isoflurane-induced cognitive impairment. EDITOR’S PERSPECTIVE

中文翻译：

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异氟烷通过规范的 NO 介导的蛋白激酶 G 信号传导破坏幼年小鼠突触后 Density-95 蛋白相互作用，导致神经元突触损失和认知障碍。

背景已知吸入麻醉剂会破坏PDZ2结构域介导的突触后密度(PSD)-95蛋白的蛋白质-蛋白质相互作用。本研究的目的是调查早期异氟醚暴露对突触 PSD-95 PDZ2 结构域破坏的潜在机制，从而改变脊柱密度和认知功能。作者假设，一氧化氮 (NO) 信号通路成分激活蛋白激酶-G 构成了一种机制，可防止神经元中早期树突棘和突触的丢失以及小鼠因 PSD 的 PDZ2 结构域破坏而导致的认知障碍-95蛋白质。方法 出生后第 7 天的小鼠暴露于 1.5% 异氟烷 4 小时，或注射 8 mg/kg 活性 PSD-95 野生型 PDZ2 肽或可溶性鸟苷酸环化酶激活剂 YC-1 及其各自的对照。体外第 7 天的原代神经元暴露于异氟烷或 PSD-95 野生型 PDZ2 肽 4 小时。评估了共免疫沉淀、棘密度、突触、环鸟苷单磷酸依赖性蛋白激酶活性和新物体识别记忆。结果 相对于对照，异氟烷或 PSD-95 野生型 PDZ2 肽的暴露会导致以下结果。首先，相对于 N-甲基-d-天冬氨酸受体亚基 NR2A 和 NR2B 沉淀，PSD-95 免疫共沉淀有所减少（平均值 ± SD [对照百分比]：异氟烷，54.73 ± 16.52，P = 0.001；PSD- 95 野生型 PDZ2 肽，51.32 ± 12.93，P = 0.001)。其次，脊柱密度有所损失（平均值±SD [每 10 µm 的脊柱密度]：对照，5.28 ± 0.56 对比异氟烷，2.23 ± 0.67，P < 0.0001；PSD-95 突变 PDZ2 肽，4.74 ± 0.94 对比PSD-95 野生型 PDZ2 肽，1.47 ± 0.87，P < 0.001）和突触斑点减少（平均值 ± SD [对照百分比]：异氟烷，41.1 ± 14.38，P = 0.001；PSD-95 野生型-型 PDZ2 肽，50.49 ± 14.31，P < 0.001)。NO供体或环单磷酸鸟苷类似物可防止棘和突触损失以及环单磷酸鸟苷依赖性蛋白激酶活性的下降，但这种预防被原代神经元中的可溶性鸟苷酸环化酶或蛋白激酶-G抑制剂所阻断。第三，5周时物体识别存在缺陷（平均值±SD[识别指数]：男性，对照，64.08±10.57，异氟烷，48.49±13.41，P = 0.001，n = 60；女性，对照，67.13± 11.17 与异氟烷，53.76 ± 6.64，P = 0.003，n = 58）。通过引入 YC-1 可以预防异氟烷引起的识别记忆损伤。结论 可溶性鸟苷酸环化酶或蛋白激酶-G 的激活可防止异氟烷或 PSD-95 野生型 PDZ2 肽诱导的树突棘和突触损失。使用 YC-1（一种不依赖于 NO 的鸟苷酸环化酶激活剂）预防识别记忆，支持可溶性鸟苷酸环化酶介导的蛋白激酶-G 信号传导在对抗异氟烷引起的认知障碍的影响中的作用。编辑的观点