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体内 正电子发射断层扫描揭示大鼠深部脑刺激诱导的活动模式

Published: March 23, 2022
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我们描述了一种临床前实验方法,以评估由 体内 FDG-PET急性深部脑刺激诱导的代谢神经调节。这份手稿包括所有实验步骤,从立体定位手术到刺激治疗的应用以及 PET 图像的采集、处理和分析。

Abstract

深部脑刺激(DBS)是一种侵入性神经外科技术,基于将电脉冲应用于涉及患者病理生理学的大脑结构。尽管DBS的历史悠久,但其作用机制和适当的协议仍不清楚,这突出了旨在解决这些谜团的研究的必要性。从这个意义上说,使用功能成像技术评估DBS的 体内 效应代表了确定刺激对大脑动力学影响的有力策略。在这里,描述了临床前模型(Wistar大鼠)的实验方案,结合纵向研究[18F]-氟脱氧正电子发射断层扫描(FDG-PET),以评估DBS对脑代谢的急性后果。首先,动物接受立体定向手术,将电极植入前额叶皮层。获取每只动物的术后计算机断层扫描(CT)以验证电极位置。恢复一周后,在没有刺激的情况下获得了每只手术动物的第一个静态FDG-PET(D1),两天后(D2),在刺激动物的同时获得了第二个FDG-PET。为此,在向动物施用FDG后,将电极连接到隔离的刺激器。因此,在FDG摄取期(45分钟)刺激动物,记录DBS对大脑代谢的急性影响。鉴于本研究的探索性,FDG-PET图像基于D1和D2研究之间的配对T检验,通过体素方法进行分析。总体而言,DBS和成像研究的结合可以描述神经网络的神经调节后果,最终有助于解开围绕DBS的难题。

Introduction

术语神经刺激包括许多不同的技术,旨在刺激神经系统,其治疗目标为1。其中,深部脑刺激(DBS)是临床实践中最普遍的神经刺激策略之一。DBS包括用神经刺激器传递的电脉冲刺激深部脑核,通过放置在大脑靶标中的电极直接植入患者体内,通过立体定向手术进行调节。评估DBS在不同神经和精神疾病中的可行性的文章数量不断增加2,尽管其中只有一些获得了食品和药物协会(FDA)的批准(即特发性震颤,帕金森病,肌张力障碍,强迫症和医学难治性癫痫)3.此外,大量的大脑靶标和刺激方案正在研究中,用于DBS治疗比官方批准的更多的病理,但没有一个被认为是确定的。DBS研究和临床程序中的这些不一致可能部分是由于缺乏对其作用机制的充分理解4。因此,人们正在付出巨大的努力来破译DBS对大脑动力学的 体内 影响,因为每一次进步,无论多么小,都将有助于完善DBS方案以获得更大的治疗成功。

在这种情况下,分子成像技术为观察DBS的 体内 神经调节作用打开了直接窗口。这些方法不仅提供了确定DBS应用时的影响的机会,而且还提供了揭示其后果的性质,防止不良副作用和临床改善的机会,甚至使刺激参数适应患者的需求5。在这些方法中,使用2-脱氧-2-[18F]氟-D-葡萄糖(FDG)的正电子发射断层扫描(PET)特别令人感兴趣,因为它提供了有关不同大脑区域激活状态的特定和实时信息6。具体而言,FDG-PET成像基于神经元和神经胶质细胞之间代谢耦合的生理原理提供了神经激活的间接评估6。从这个意义上说,一些临床研究已经报道了使用FDG-PET调节DBS的大脑活动模式(见3 )。然而,临床研究在关注患者时容易出现一些缺点,例如异质性或招募困难,这极大地限制了他们的研究潜力6。这种背景导致研究人员使用人类状况的动物模型在临床转化之前评估生物医学方法,或者,如果已经在临床实践中应用,则解释治疗益处或副作用的生理起源。因此,尽管人类病理学与实验动物的模拟状况之间存在很大距离,但这些临床前方法对于安全有效地过渡到临床实践至关重要。

这份手稿描述了小鼠模型的实验性DBS方案,并结合纵向FDG-PET研究,以评估DBS对大脑代谢的急性后果。通过该协议获得的结果可能有助于解开DBS诱导的大脑活动的复杂性调节模式。因此,提供了一种合适的实验策略来检查 体内 刺激的后果,使临床医生能够预测特定情况下的治疗效果,然后根据患者的需求调整刺激参数。

Protocol

实验动物程序根据欧洲共同体理事会指令2010/63 / EU进行,并得到格雷戈里奥·马拉尼翁医院动物实验伦理委员会的批准。实验方案的图形摘要如图 1A所示。 1. 通过体内神经成像定位脑靶标 动物制备注意:使用~300g的雄性Wistar大鼠。将动物放入麻醉诱导盒中并密封顶部。 打开七氟醚蒸发器(5%用于100%O2中的诱?…

Representative Results

在研究结束时或当动物的福利受到损害时,使用CO2处死动物。 图3显示了来自手术动物的完整PET / CT研究示例。因此,插入大鼠大脑的电极可以在 图3A所示的CT图像中清楚地观察到。这种成像模式提供了良好的解剖学信息,并有助于配准FDG-PET图像,因为功能模式往往比结构图像更模糊(图3A,B)。此外,同一?…

Discussion

鉴于对脑功能和神经精神疾病病理生理学所涉及的神经网络的理解取得了进展,越来越多的研究认识到DBS在广泛的基于神经的病理学中的潜力2。然而,这种疗法的作用机制尚不清楚。一些理论试图解释在特定病理和刺激情况下获得的效果,但所提出的研究的异质性使得很难得出明确的结论4。因此,尽管付出了巨大的努力,但没有真正的共识,但接受DBS干预的?…

Açıklamalar

The authors have nothing to disclose.

Acknowledgements

我们感谢Christine Winter教授,Julia Klein,Alexandra de Francisco和Yolanda Sierra在优化本文所述方法方面的宝贵支持。MLS得到了科学和创新部,卡洛斯三世健康研究所(项目编号PI17/01766和赠款号BA21/0030)的支持,由欧洲区域发展基金(ERDF)共同资助,“创造欧洲的方式”;CIBERSAM(项目编号CB07/09/0031);德罗加斯国家计划(项目编号2017/085);马夫雷基金会;和艾丽西亚·科普洛维茨基金会。 MCV得到了塔蒂亚娜·佩雷斯·德·古兹曼·布埃诺基金会的支持,作为该机构的奖学金获得者,以及欧盟联合计划 – 神经退行性疾病研究(JPND)。DRM得到了马德里社区教育与调查委员会的支持,由欧洲社会基金“投资你的未来”(资助号PEJD-2018-PRE/BMD-7899)共同资助。NLR得到了格雷戈里奥·马拉尼翁卫生研究所的支持,“2019年I+D+I壁内计划”。医学博士的工作得到了 科学和创新部(MCIN)和卡洛斯三世健康研究所(ISCIII)的支持(PT20/00044)。CNIC得到了卡洛斯三世健康研究所(ISCIII),科学与创新部(MCIN)和Pro CNIC基金会的支持,并且是Severo Ochoa卓越中心(SEV-2015-0505)。

Materials

7-Tesla Biospec 70/20 scanner Bruker, Germany SN0021 MRI scanner for small animal imaging
Betadine Meda Pharma S.L., Spain 644625.6 Iodine solution (iodopovidone)
Beurer IL 11 Beurer SN87318 Infra-red light
Bipolar cable 50 cm w/50 cm mesh covering up to 100 cm Plastics One, USA 305-305 (CM)
Bipolar cable TT2  50 cm up to 100 cm Plastics One, USA 305-340/2 Bipolar cable TT2  50 cm up to 100 cm
Buprex Schering-Plough, S.A 961425 Buprenorphine (analgesic)
Ceftriaxona Reig Jofré 1g IM Laboratorio Reig Jofré S.A., Spain 624239.1 Ceftriaxone (antibiotic)
Commutator Plastics One, USA SL2+2C 4 Channel Commutator for DBS
Concentric bipolar platinum-iridium electrodes Plastics One, USA MS303/8-AIU/Spc Electrodes for DBS
Driller Bosh T58704 Driller
FDG Curium Pharma Spain S.A., Spain —– 2-[18F]fluoro-2-deoxy-D-glucose (PET radiotracer)
Heating pad DAGA, Spain 23115 Heating pad
Ketolar Pfizer S.L., Spain 776211.9 Ketamine (anesthetic drug)
Lipolasic 2 mg/g Bausch & Lomb S.A, Spain 65277 Ophthalmic lubricating gel
MatLab R2021a The MathWorks, Inc Support software for SPM12
MRIcro McCausland Center for Brain Imaging,  University of South Carolina, USA v2.1.58-0 Software for imaging preprocessing and analysis
Multimodality Workstation (MMWKS) BiiG, Spain Software for imaging processing and analysis
Omicrom VISION VET RGB Medical Devices, Spain 731100 ReV B Cardiorrespiratory monitor for small imaging
Prevex Cotton buds Prevex, Finland —– Cotton buds
Sevorane AbbVie Spain, S.L.U, Spain 673186.4 Sevoflurane (inhalatory anesthesia)
Small screws Max Witte GmbH 1,2 x 2 DIN 84 A2 Small screws
Standard U-Frame Stereotaxic Instrument for Rat, 18° Ear Bar Harvard Apparatus, USA 75-1801 Two-arms Stereotactic frame for rat
Statistical Parametric Mapping (SPM12) The Wellcome Center for Human Neuroimaging, UCL Queen Square Institute of Neurology, UK SPM12 Software for voxel-wise imaging analysis
STG1004 Multi Channel Systems GmbH, Germany STG1004 Isolated stimulator
SuperArgus PET/CT scanner Sedecal, Spain S0026403 NanoPET/CT scanner for small animal imaging
Suture thread with needle, 1/º Lorca Marín S.A., Spain 55325 Braided natural silk non-absorbable suture 1/0, with triangle needle
Technovit 4004 (powder and liquid) Kulzer Technique, Germany 64708471; 64708474 Acrylic dental cement for craniotomy tap
Wistar rats (Rattus norvergicus) Charles River, Spain animal facility Animal model used
Xylagesic Laboratorios Karizoo, A.A, Spain 572599-4 Xylazine (anesthetic drug)
Normon S.A., Spain 602910 Mepivacaine in gel for topical use

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Bu Makaleden Alıntı Yapın
Casquero-Veiga, M., Lamanna-Rama, N., Romero-Miguel, D., Desco, M., Soto-Montenegro, M. L. In vivo Positron Emission Tomography to Reveal Activity Patterns Induced by Deep Brain Stimulation in Rats. J. Vis. Exp. (181), e63478, doi:10.3791/63478 (2022).

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