Femtosecond laser-produced sub-surface cuts to halt focal epileptic seizures

描述(申请人提供)：局灶性新皮质癫痫是一个在很大程度上难以解决的医学问题，大多数病例对抗惊厥药物反应不佳，目前的手术治疗选择因神经功能障碍的可能性而受到限制。由于许多信息处理是在皮质中垂直组织的，而癫痫的传播主要通过侧向连接进行，因此一系列的切开可以阻止癫痫的扩散或启动 但在很大程度上保留了功能。这些切开不能切断大脑表面的血管，目前还不清楚切断哪些皮质层最好，以实现最佳的癫痫控制和最小的神经影响。紧聚焦的飞秒激光脉冲提供了一种独特的工具，可以在组织的整体内进行微米级的切割，并将附带损害降至最低。我们假设，使用这些切开横断目标皮质层中的神经连接将阻止局灶性癫痫发作的启动或传播。由于这些切割只针对特定皮质层的水平连接，而大脑皮层的大部分神经连接都得到了保留，因此将会有最少的 神经缺陷。这项建议的一个主要目标是确定应该切断哪一层大脑皮层(S)，以何种几何模式最大限度地干扰癫痫发作的启动和传播，同时将对正常功能的影响降至最低。在目标1中，我们测试了飞秒激光切割特定皮质层在预防癫痫的启动和传播方面的急性疗效。采用皮质微量注射4-氨基吡啶的方法复制大鼠癫痫发作模型。局域场电位记录和双光子钙敏感染料成像用于监测神经活动和癫痫的传播。飞秒激光消融是用亚表面切割来包围或细分癫痫发作起始点。首先，我们确定必须横切哪些皮质层，以防止癫痫发作传播到被包围的区域之外。目标是确定为遏制癫痫发作而削减的最低层数。基于最近的数据表明，临床癫痫发作是由微癫痫发作合并引起的，我们接下来研究在癫痫发作起始部位切割的网格图案是否可以通过分离微域来防止癫痫发作。在目标2中，我们通过记录胡须刺激后躯体感觉皮层诱发信号的变化来探索从目标1到最有希望的激光切割的潜在副作用。这些实验将在动物模型中测试一种基于激光的治疗局灶性新皮质癫痫的新手术方法。此外，这项工作将提供有价值的活体数据，关于皮质层特异性癫痫发作的启动和传播。如果这里提出的急性动物模型实验以及未来评估长期有效性的研究被证明是成功的，那么使用最近开发的激光技术在人类身上实施是可行的，并将使除脑沟底部外的所有部位都能产生特定层的切割，从而为癫痫的新手术治疗打开了大门。 与公共卫生相关：这里提出的工作可以为一种潜在的根治手术疗法提供基础，同时将对局灶性新皮质癫痫患者的神经副作用降至最低，其研究方法与癫痫研究中更典型的以分子/细胞为重点的基础研究和基于药理学的治疗开发明显不同。如果我们的工作成功，利用最近开发的激光技术，将我们的基于激光的手术方法扩展到人类是可行的，这可能对癫痫的治疗产生重大的、变革性的影响。这一点尤其正确，因为那些不能用药物很好地控制癫痫发作的患者往往会出现我们建议的治疗方法最好治疗的那种局灶性癫痫发作。

DESCRIPTION (provided by applicant): Focal neocortical epilepsy is a largely intractable medical problem, with most cases responding poorly to anti-convulsive medications and current surgical treatment options limited because of the likelihood of neurological deficits. Because much information processing is vertically organized in cortex, while seizure propagation occurs primarily through lateral connections, a series of incisions could prevent the spread or initiation of seizures but largely preserve function. These incisions must not cut the blood vessels on the brain surface and it remains unclear which cortical layers are best cut to achieve optimal seizure control and minimal neurological impact. Tightly-focused femtosecond laser pulses provide a unique tool to make micrometer-scale cuts several millimeters within the bulk of a tissue with minimal collateral damage. We hypothesize that using these cuts to transect the neural connections in targeted cortical layers will block the initiation or propagation of focally initiatd epileptic seizures. Because these cuts target only horizontal connections in a specific cortical layer and the majority of the neural connectivity of the cortex is preserved, there will be minimal neurological deficit. A primary goal of this proposal is to determine which cortical layer(s) shoul be cut and in what geometric pattern to maximally interfere with seizure initiation and propagation while minimally affecting normal function. In Aim 1, we test the acute efficacy of femtosecond laser cuts to specific cortical layers in preventing epilepsy initiation and propagation. Epileptic seizures are modeled in rats by microinjection of 4-aminopyridine into cortex. Local field potential recordings and two-photon calcium sensitive dye imaging are used to monitor neural activity and seizure propagation. Femtosecond laser ablation is used to encircle or subdivide the seizure initiation site with subsurface cuts. First, we determine which cortical layers must be transected to prevent seizures from propagating outside of the encircled region. The goal is to determine the minimum number of layers to cut for seizure containment. Building on recent data that suggests that clinical seizures result from the coalescence of microseizures, we next investigate whether a grid pattern incised at the seizure initiation site can prevent seizure initiation by separating microdomains. In Aim 2, we explore potential side effects of the most promising laser cuts from Aim 1 by recording changes in evoked signals in somatosensory cortex after whisker stimulation. These experiments will test, in animal models, a new laser-based surgical method for the treatment of focal neocortical epilepsy. In addition, this work will provide valuable, in vivo data on cortical layer- specific initiation and propagatio of seizures. If the acute animal model experiments proposed here as well as future studies that evaluate the longer-term effectiveness prove successful then human implementation is feasible using recently-developed laser technology and would enable layer-specific cuts to be produced in all but the bottom of sulci, opening the door to new surgical treatments for epilepsy. PUBLIC HEALTH RELEVANCE: The work proposed here could provide the basis for a potentially curative surgical therapy with minimized neurological side effects to patients with focal neocortical epilepsy, with a research approach that diverges significantly from the very molecular/cellular focused basic research and pharmacology-based therapy development more typical of epilepsy research. If our work is successful, extension of our laser-based surgical method to humans is feasible with recently-developed laser technology, potentially having a major, transformative impact on epilepsy treatment. This is particularly true because the patients who are not able to control their seizures well with medication tend to have the kind of focally-initiated seizures that the therapy we propose would be best to treat.