Reis W L, Biancardi V C, Son S, Antunes-Rodrigues J, Stern J E
Department of Physiology, Georgia Regents University, Augusta, GA, USA; Department of Physiology, School of Medicine of Ribeirao Preto, University of Sao Paulo, Ribeirao Preto, SP, Brazil.
J Neuroendocrinol. 2015 Feb;27(2):111-22. doi: 10.1111/jne.12245.
Nitric oxide (NO) and carbon monoxide (CO) are diffusible gas messengers in the brain. Previously, we have shown their independent involvement in central fluid/electrolyte homeostasis control. In the present study, we investigated a possible functional interaction between NO/CO in the regulation of vasopressin (VP) and oxytocin (OT) magnocellular neurosecretory cells (MNCs) activity in euhydrated (EU) and dehydrated [48-h water-deprived (48WD)] rats. Using brain slices from EU and 48WD rats, we measured, by immunohistochemistry, the expression of neuronal NO synthase (nNOS, which synthesises NO) and haeme-oxygenase (HO-1, which synthesises CO) in the hypothalamic supraoptic nucleus (SON). In addition, we used patch-clamp electrophysiology to investigate whether regulation of SON MNC firing activity by endogenous CO was dependent on NO bioavailability and GABAergic inhibitory synaptic function. We found a proportion of OT and VP SON MNCs in EU rats to co-express both of HO-1 and nNOS (33.2 ± 2.9% and 15.3 ± 1.4%, respectively), which was increased in 48WD rats (55.5 ± 0.9% and 21.0 ± 1.7%, respectively, P < 0.05 for both). Inhibition of endogenous HO activity [chromium mesoporphyrin IX chloride (CrMP) 20 μm] induced MNC membrane hyperpolarisation and decreased firing activity, and these effects were blunted by previous blockade of endogenous NOS activity (l-NAME, 2 mm) or blockade of inhibitory GABA function [Picrotoxin (Sigma-Aldrich, St Louis, MO, USA), 50 μm]. No significant changes in SON NO bioavailability (4,5 diaminofluorescein diacetate fluorescence) were observed after CrMP treatment. Taken together, our results support a state-dependent functional inter-relationship between NO and CO in MNCs, in which CO acts as an excitatory gas molecule, whose effects are largely dependent on interactions with the inhibitory SON signals NO and GABA.
一氧化氮（NO）和一氧化碳（CO）是大脑中的可扩散气体信使。此前，我们已证明它们分别参与中枢液体/电解质稳态控制。在本研究中，我们调查了在正常水合（EU）和脱水[48小时禁水（48WD）]大鼠中，NO/CO在调节血管加压素（VP）和催产素（OT）大细胞神经分泌细胞（MNCs）活性方面可能存在的功能相互作用。使用来自EU和48WD大鼠的脑片，我们通过免疫组织化学测量了下丘脑视上核（SON）中神经元型一氧化氮合酶（nNOS，合成NO）和血红素加氧酶（HO-1，合成CO）的表达。此外，我们使用膜片钳电生理学来研究内源性CO对SON MNC放电活动的调节是否依赖于NO的生物利用度和GABA能抑制性突触功能。我们发现，在EU大鼠中，一定比例的OT和VP SON MNCs共表达HO-1和nNOS（分别为33.2±2.9%和15.3±1.4%），而在48WD大鼠中这一比例增加（分别为55.5±0.9%和21.0±1.7%，两者P<0.05）。抑制内源性HO活性[氯化中卟啉铬（CrMP）20μm]会导致MNC膜超极化并降低放电活动，而这些效应会被先前对内源性NOS活性的阻断（左旋硝基精氨酸甲酯，l-NAME，2mm）或对抑制性GABA功能的阻断[印防己毒素（美国密苏里州圣路易斯西格玛奥德里奇公司），50μm]所减弱。CrMP处理后，SON中NO的生物利用度（4,5-二氨基荧光素二乙酸酯荧光）未观察到显著变化。综上所述，我们的结果支持MNCs中NO和CO之间存在状态依赖性的功能相互关系，其中CO作为一种兴奋性气体分子，其作用在很大程度上依赖于与抑制性SON信号NO和GABA的相互作用。
