Steffensen Annette B, Sword Jeremy, Croom Deborah, Kirov Sergei A, MacAulay Nanna
Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, DK-2200 Copenhagen, Denmark, and.
Brain and Behavior Discovery Institute and.
J Neurosci. 2015 Sep 2;35(35):12172-87. doi: 10.1523/JNEUROSCI.0400-15.2015.
Spreading depolarizations (SDs) are waves of sustained neuronal and glial depolarization that propagate massive disruptions of ion gradients through the brain. SD is associated with migraine aura and recently recognized as a novel mechanism of injury in stroke and brain trauma patients. SD leads to neuronal swelling as assessed in real time with two-photon laser scanning microscopy (2PLSM). Pyramidal neurons do not express aquaporins and thus display low inherent water permeability, yet SD rapidly induces focal swelling (beading) along the dendritic shaft by unidentified molecular mechanisms. To address this issue, we induced SD in murine hippocampal slices by focal KCl microinjection and visualized the ensuing beading of dendrites expressing EGFP by 2PLSM. We confirmed that dendritic beading failed to arise during large (100 mOsm) hyposmotic challenges, underscoring that neuronal swelling does not occur as a simple osmotic event. SD-induced dendritic beading was not prevented by pharmacological interference with the cytoskeleton, supporting the notion that dendritic beading may result entirely from excessive water influx. Dendritic beading was strictly dependent on the presence of Cl(-), and, accordingly, combined blockade of Cl(-)-coupled transporters led to a significant reduction in dendritic beading without interfering with SD. Furthermore, our in vivo data showed a strong inhibition of dendritic beading during pharmacological blockage of these cotransporters. We propose that SD-induced dendritic beading takes place as a consequence of the altered driving forces and thus activity for these cotransporters, which by transport of water during their translocation mechanism may generate dendritic beading independently of osmotic forces.
Spreading depolarization occurs during pathological conditions such as stroke, brain injury, and migraine and is characterized as a wave of massive ion translocation between intracellular and extracellular space in association with recurrent transient focal swelling (beading) of dendrites. Numerous ion channels have been demonstrated to be involved in generation and propagation of spreading depolarization, but the molecular machinery responsible for the dendritic beading has remained elusive. Using real-time in vitro and in vivo two-photon laser scanning microscopy, we have identified the transport mechanisms involved in the detrimental focal swelling of dendrites. These findings have clear clinical significance because they may point to a new class of pharmacological targets for prevention of neuronal swelling that consequently will serve as neuroprotective agents.
扩散性去极化（SDs）是持续的神经元和胶质细胞去极化波，它在大脑中传播离子梯度的大规模破坏。SD与偏头痛先兆有关，最近被认为是中风和脑外伤患者损伤的一种新机制。如用双光子激光扫描显微镜（2PLSM）实时评估，SD会导致神经元肿胀。锥体神经元不表达水通道蛋白，因此表现出低固有水渗透性，但SD通过未知分子机制迅速诱导沿树突轴的局灶性肿胀（串珠状）。为解决这个问题，我们通过局部微量注射氯化钾在小鼠海马切片中诱导SD，并通过2PLSM观察随后表达EGFP的树突的串珠状变化。我们证实，在大的（100 mOsm）低渗刺激期间树突串珠状变化未出现，这突出表明神经元肿胀并非简单的渗透事件。对细胞骨架的药理学干扰并不能阻止SD诱导的树突串珠状变化，这支持了树突串珠状变化可能完全由过多的水流入导致的观点。树突串珠状变化严格依赖于Cl(-)的存在，因此，联合阻断Cl(-)偶联转运体可导致树突串珠状变化显著减少，而不干扰SD。此外，我们的体内数据显示，在对这些共转运体进行药理学阻断期间，树突串珠状变化受到强烈抑制。我们提出，SD诱导的树突串珠状变化是这些共转运体驱动力和活性改变的结果，这些共转运体在其转运机制中通过水的转运可能独立于渗透力产生树突串珠状变化。
扩散性去极化发生在中风、脑损伤和偏头痛等病理状态下，其特征是细胞内和细胞外空间之间大量离子转运的波，伴有树突反复短暂的局灶性肿胀（串珠状）。许多离子通道已被证明参与扩散性去极化的产生和传播，但负责树突串珠状变化的分子机制仍然难以捉摸。使用实时体外和体内双光子激光扫描显微镜，我们确定了参与树突有害局灶性肿胀的转运机制。这些发现具有明确的临床意义，因为它们可能指向一类新的预防神经元肿胀的药理学靶点，从而将作为神经保护剂。
