用于缺血和复苏期间脑成像和脑生理学监测的小鼠心脏骤停模型
Summary
该方案展示了一种独特的窒息性心脏骤停小鼠模型，该模型不需要胸外按压进行复苏。该模型可用于监测和成像心脏骤停和复苏期间的大脑生理动力学。
Abstract
大多数心脏骤停 （CA） 幸存者会出现不同程度的神经功能缺损。为了了解支撑 CA 诱导的脑损伤的机制并随后开发有效的治疗方法，实验性 CA 研究至关重要。为此，已经建立了一些小鼠CA模型。在大多数这些模型中，将小鼠置于仰卧位，以便进行胸外按压以进行心肺复苏（CPR）。然而，这种复苏程序使得 CA 和复苏期间大脑生理学的实时成像/监测具有挑战性。为了获得这些关键知识，本方案提出了一种不需要胸外按压CPR步骤的小鼠窒息CA模型。该模型可以研究血流、血管结构、电位和脑组织氧的动态变化，从 CA 前基线到 CA 再灌注后早期。重要的是，该模型适用于老年小鼠。因此，该小鼠CA模型有望成为破译CA对大脑生理学影响的关键工具。
Introduction
心脏骤停 （CA） 仍是一场全球公共卫生危机1.仅在美国，每年就报告超过 356,000 例院外 CA 病例和 290,000 例住院 CA 病例，大多数 CA 患者年龄超过 60 岁。值得注意的是，冠状动脉后神经损伤在幸存者中很常见，这对冠状动脉管理构成了重大挑战2,3,4,5。为了了解 CA 后脑病理变化及其对神经系统结果的影响，已在患者6、7、8、9、10、11、12 中应用了各种神经生理学监测和脑组织监测技术。使用近红外光谱，还对 CA 大鼠进行了实时脑部监测，以预测神经系统结果13。
然而，在小鼠 CA 模型中，由于需要胸外按压以恢复自发循环，这种成像方法变得复杂，这总是需要大量的身体运动，因此阻碍了精细的成像程序。此外，CA模型通常在小鼠仰卧位的情况下进行，而对于许多脑成像方式，小鼠必须转向俯卧位。因此，在许多情况下，需要在手术过程中具有最小身体运动的小鼠模型，以便在从CA前到复苏后的整个CA过程中对大脑进行实时成像/监测。
此前，Zhang 等人报道了一种可用于脑成像的小鼠 CA 模型14。在他们的模型中，CA是通过推注维库溴铵和艾司洛尔诱导的，然后停止机械通气。他们表明，在CA5分钟后，可以通过输注复苏混合物来实现复苏。然而，值得注意的是，在他们的模型中，循环停滞仅发生在艾司洛尔注射后约 10 秒。因此，该模型没有概括患者窒息诱导的 CA 的进展，包括逮捕前期间的高碳酸血症和组织缺氧。
目前外科手术的总体目标是模拟小鼠的临床窒息性CA，然后在没有胸外按压的情况下进行复苏。因此，该 CA 模型允许使用复杂的成像技术来研究小鼠的大脑生理学15。
Protocol
这里描述的所有程序都是根据美国国立卫生研究院 （NIH） 关于在研究中护理和使用动物的指南进行的，并且该协议已获得杜克大学动物护理和使用委员会研究所 （IACUC） 的批准。本研究采用8-10周龄的C57BL/6雄性和雌性小鼠。 1. 手术准备 在数字秤上称量小鼠，并将其放入 4 英寸 x 4 英寸 x 7 英寸的有机玻璃麻醉诱导盒中。 将麻醉汽化器调整为5%异氟…
Representative Results
为了诱导CA，用1.5%异氟烷麻醉小鼠，并用100%氮气通气。这种情况导致 45 秒内出现严重的心动过缓（图 1）。缺氧2分钟后，心率急剧降低（图2），血压降至20mmHg以下，脑血流完全停止（图1）。当异氟醚被关闭时，体温不再被控制，并在CA结束时缓慢下降到约32°C（图1）。 CA 8 分钟后，立…
Discussion
在实验性 CA 研究中，窒息、氯化钾注射或电流衍生的心室颤动已被用于诱导 CA 16、17、18、19、20、21、22、23。通常，在这些 CA 模型中，复苏需要心肺复苏术，尤其是在小鼠中。我们配制了一种复?…
Divulgations
The authors have nothing to disclose.
Acknowledgements
作者感谢凯西·盖奇（Kathy Gage）的编辑支持。这项研究得到了麻醉学系（杜克大学医学中心）、美国心脏协会资助 （18CSA34080277） 和美国国立卫生研究院 （NIH） 资助（NS099590、HL157354、NS117973 和 NS127163）的资金支持。
Materials
| Adrenalin | Par Pharmaceutical | NDC 42023-159-01 | |
| Alcohol swabs | BD | 326895 | |
| Animal Bio Amp | ADInstruments | FE232 | |
| BP transducer | ADInstruments | MLT0699 | |
| Bridge Amp | ADInstruments | FE117 | |
| Heparin sodium injection, USP | Fresenius Kabi | NDC 63323-540-05 | |
| Isoflurane | Covetrus | NDC 11695-6777-2 | |
| Laser Doppler perfusion monitor | Moor Instruments | moorVMS-LDF1 | |
| Laser speckle imaging system | RWD | RFLSI III | |
| Lubricant eye ointment | Bausch + Lomb | 339081 | |
| Micro clip | Roboz | RS-5431 | |
| Mouse rectal probe | Physitemp | RET-3 | |
| Needle electrode | ADInstruments | MLA1213 | 29 Ga, 1.5 mm socket |
| Nitrogen | Airgas | UN1066 | |
| Optic plastic fibre | Moor Instruments | POF500 | |
| Otoscope | Welchallyn | 728 | 2.5 mm Speculum |
| Oxygen | Airgas | UN1072 | |
| PE-10 tubing | BD | 427401 | Polyethylene tubing |
| Povidone-iodine | CVS | 955338 | |
| PowerLab 8/35 | ADInstruments | ||
| Rimadyl (carprofen) | Zoetis | 6100701 | Injectable 50 mg/ml |
| Small animal ventilator | Kent Scientific | RoVent Jr. | |
| Temperature controller | Physitemp | TCAT-2DF | |
| Triple antibioric & pain relief | CVS | NDC 59770-823-56 | |
| Vaporizer | RWD | R583S | |
| 0.25% bupivacaine | Hospira | NDC 0409-1159-18 | |
| 0.9% sodium chroride | ICU Medical | NDC 0990-7983-03 | |
| 1 mL plastic syringe | BD | 309659 | |
| 4-0 silk suture | Look | SP116 | Black braided silk |
| 6-0 nylon suture | Ethilon | 1698G | |
| 8.4% sodium bicarbonate Inj., USP | Hospira | NDC 0409-6625-02 | |
| 20 G IV catheter | BD | 381534 | 20GA 1.6 IN |
| 30 G PrecisionGlide needle | BD | 305106 | 30 G X 1/2 |
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Citer Cet Article
Li, R., Duan, W., Zhang, D., Hoffmann, U., Yao, J., Yang, W., Sheng, H. Mouse Cardiac Arrest Model for Brain Imaging and Brain Physiology Monitoring During Ischemia and Resuscitation. J. Vis. Exp. (194), e65340, doi:10.3791/65340 (2023).