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Medicina

使用激光多普勒流测量测量大鼠脑血流自动调节的评估

Published: January 19, 2020
doi:
10.3791/60540
, , ,
1Department of Physiology,Medical College of Wisconsin

Summary

本文演示了使用激光多普勒流量测量来评估大脑循环在动脉血压降低期间自动调节其血流的能力。

Abstract

在研究大脑血流调节机理时，可以使用激光多普勒流量测定仪（LDF）获得对微循环血流的相对测量。本文演示了一种封闭的颅骨制备，允许在不穿透头骨或安装腔室或脑窗的情况下评估脑血流。为了评估自动调节机制，可以同时使用LDF，同时利用通过分级出血控制血压降低的模型。这使得实时跟踪血流的相对变化，以响应循环血量的戒断所产生的动脉血压降低。这种范例是研究脑血流在动脉血压降低期间自动调节的宝贵方法，并且，在协议稍作修改后，作为出血性休克的实验模型也很有价值。除了评估自动调节反应外，LDF 还可用于监测皮质血流，用于调查调节脑血流和各种实验的影响的代谢、造血、内皮、体液或神经机制脑血流的介入和病理状况。

Introduction

大脑循环中的自动调节机制在维持大脑平衡和正常功能方面起着至关重要的作用。脑血流的自动调节受心率、血速、灌注压力、脑阻力动脉直径、微循环阻力等多种因素的影响，这些因素对维持大脑总脑血流在全身血压生理范围内的作用。当动脉压力增加时，这些机制收缩动脉和阻力动脉，以防止颅内压力的危险增加。当动脉血压降低时，局部控制机制使动脉稀释，以保持组织灌注和O2分娩。各种病理状况，如高帽病、创伤性或全球缺氧性脑损伤，以及糖尿病微血管病1、2、3、4、5、6，可能会破坏大脑自动调节其血流的能力。例如，慢性高血压将有效的自动调节范围向高压力7、8、9转移，高盐（HS）饮食不仅干扰脑微循环10的正常内皮依赖性扩张，而且会削弱脑循环中自动调节机制在动脉压力降低时扩张和维持组织灌注的能力。大脑自动调节也损害在达尔盐敏感大鼠时，他们被喂食HS饮食12。

在动脉压力降低期间，尽管灌注压力降低，但脑阻力动脉和动脉的扩张最初使脑血流恢复控制值。随着动脉压力的进一步降低，脑血流在较低压力（自动调节反应的高原阶段）保持恒定，直到血管不再扩张以保持血流量在较低压力下。器官能够维持正常血流的最低压力称为自动调节 （LLA） 的下限。在LLA以下的压力下，脑血流量从静息值中显著减少，并且随着动脉灌注压力的每次降低，脑血流以线性方式减少。如高血压7、8、9所观察到的LLA向上移位，在动脉灌注压力降低（例如心肌梗死、缺血性中风或循环休克）的情况下，可能会增加缺血损伤的风险和严重程度。

LDF已被证明是评估微循环中血流的极有价值的方法，在各种情况下，包括大脑循环11、14、15的血流的自动调节。除了评估自动调节反应，LDF可用于监测皮质血流时，调查代谢，造血，内皮，体液，或神经机制，调节大脑血流和各种实验干预和病理条件对脑血流的影响10，16，17，18，19，20，21。

LDF测量反射激光的变化，以响应运动粒子的数量和速度，在这种情况下，红血球（RBC）。对于脑血管自动调节的研究，动脉血压通过注射α-肾上腺素激动剂来增加动脉压力而改变（因为大脑循环本身对α-阿德雷涅克血管收缩剂激动剂不敏感）12、15或通过控制血量戒断来降低动脉压力11、14。在本研究中，LDF用于演示降压对健康大鼠大脑自动调节的影响。虽然文献中已经描述了开闭头骨方法，但本文展示了一个封闭的头骨准备，允许在不穿透头骨或安装室或脑窗的情况下评估脑血流。

Protocol

威斯康星医学院机构动物护理和使用委员会 （IACUC） 批准了本文中描述的所有协议，所有程序都符合国家卫生研究院 （NIH） 实验室动物福利办公室 （OLAW） 的要求。法规。 1. 实验动物和准备录音 使用重达250~300克的8~12周大雄性斯普拉格-道利大鼠。对于这些实验，喂食大鼠的标准饮食包括0.4%纳氯、200克/千克碱、3克/千克DL-蛋氨酸、497.77克/千克蔗糖、150克/千克玉米淀…

Representative Results

图2总结了10只雄性斯普拉格-道利大鼠喂食标准实验室的试验结果。在这些实验中，平均LCBF在前三次血量戒断后保持在出血前值的20%以内，直到平均动脉压力达到LLA。随后在LLA以下压力下抽取血量导致LCBF逐渐减少，表明脑循环不再能够产生足够的血管扩张水平，以维持在较低的灌注压力下脑血流恒定。 图 3总结了高原相中的平均动…

Discussion

使用激光多普勒流量测定 （LDF） 评估组织血流反应。如上所述，LDF 信号与微循环中移动粒子的数量和速度成正比，在这种情况下为 RBC。不同器官的LDF读数与电磁流量计和放射性微球30等既定方法评估的全器官血流密切相关，与评价可点化动脉制剂10、31、32、33、34和原位微循环制剂<sup cl…

Divulgazioni

The authors have nothing to disclose.

Acknowledgements

作者衷心感谢卡利·科扎克、梅根·斯图普夫和杰克·布力斯在完成这项研究和准备手稿方面给予的出色帮助。资助：NIH #R01-HL128242、#R21-OD018309和#R21-OD024781。

Materials

3-0 braided black silk suture Midwest Vet 193.73000.2
Arterial Pressure Transducer Merit Medical 041516504A
Automated Data Acquisition Systems (WINDAQ & BIOPAC system) DATAQ Instruments
Blood Pressure Display Unit Stoelting 50115
Circulating warm water pump Gaymar Industries T-pump
End-tidal CO2 monitor Stoelting Capstar-100
Heparin Sodium Midwest Vet 191.46720.3
Kimwipe Fisher Scientific 06-666A
Laser Doppler Flow Meter Perimed PeriFlux 5000 LDPM
Laser Doppler Refill Motility Standard Perimed PF1001
Polyethylene Tubing (PE240) (for trachea cannula) VWR 63018-828
Polyethylene Tubing (PE50) (for femoral catheters) VWR 63019-048
Rodent Ventilator Cwe/Stoelting SAR-830/P
Saline Midwest Vet 193.74504.3
Sprague-Dawley Outbred Rats Variable N/A Rats were ordered from various companies
Standard Rat Chow Dyets, Inc. 113755
Stereotaxic Instrument Cwe/Stoelting Clasic Lab Standard
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Tags

Laser Doppler FlowmetryCerebral Blood Flow AutoregulationRatNon-invasiveBlood PressureVascular BedsFemoral ArteryCannulaAnesthesiaShavingDisinfectionSuturesLigation

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Allen, L. A., Terashvili, M., Gifford, A., Lombard, J. H. Evaluation of Cerebral Blood Flow Autoregulation in the Rat Using Laser Doppler Flowmetry. J. Vis. Exp. (155), e60540, doi:10.3791/60540 (2020).
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