Department of Biomedical Engineering, Cornell University, Ithaca, New York, United States of America.
PLoS One. 2013 May 28;8(5):e65663. doi: 10.1371/journal.pone.0065663. Print 2013.
Although microhemorrhages are common in the brain of the elderly, the direct impact of these lesions on neural function remains unclear. In this work, we used femtosecond laser irradiation to rupture the wall of single arterioles in the brain of anesthetized rodents, producing a hematoma of ∼100-µm diameter. Our objective was to study the impact of these microhemorrhages on cortical activity using cell-resolved two-photon imaging of bulk-loaded calcium-sensitive dye. We monitored peripheral sensory stimulus-induced calcium transients from individual neuronal cell bodies, regions of neuropil, and astrocytes at different distances from the microhemorrhage before and 0.5, 2, and 4 hours after the creation of the lesion. We found that immediately after the hemorrhage the average amplitude of the stimulus-induced calcium response was reduced to about half within 150 µm from the hematoma. Beyond 300 µm, there was little effect on cell response, with a smooth increase in response amplitude from 150 µm to 300 µm from the lesion. Cortical function gradually improved with time and by four hours after the lesion the response from neurons and astrocytes had recovered to baseline everywhere but within 150 µm from the hematoma. To assess whether the cells closest to the microhemorrhage recovered over a longer timeframe, we developed a re-openable chronic cranial window preparation that allowed reinjection of calcium-sensitive fluorescent dye. We found that the response largely recovered by one day after the microhemorrhage even within 150 µm from the hematoma. This work suggests that neuronal and astrocyte function is transiently lost near a microhemorrhage, but recovers within one day after the lesion.
尽管老年人的大脑中普遍存在微出血，但这些病变对神经功能的直接影响尚不清楚。在这项工作中，我们使用飞秒激光照射麻醉啮齿动物大脑中的单个动脉壁，产生直径约为 100-µm 的血肿。我们的目的是使用批量加载钙敏染料的细胞分辨双光子成像来研究这些微出血对皮质活动的影响。我们在创建病变之前和之后 0.5、2 和 4 小时，监测来自单个神经元细胞体、神经胶质区域和距微出血不同距离的星形胶质细胞的外周感觉刺激诱导的钙瞬变。我们发现，在出血后立即，血肿 150 µm 范围内刺激诱导的钙反应的平均幅度降低到约一半。超过 300 µm 后，对细胞反应的影响很小，响应幅度从血肿处的 150 µm 到 300 µm 逐渐增加。皮质功能随着时间的推移逐渐改善，并且在病变后四个小时，神经元和星形胶质细胞的反应除了血肿 150 µm 范围内之外，已恢复到基线。为了评估距离微出血最近的细胞是否在更长的时间内恢复，我们开发了一种可重新打开的慢性颅窗制备方法，该方法允许重新注射钙敏荧光染料。我们发现，即使在血肿 150 µm 范围内，微出血后一天内反应也基本恢复。这项工作表明，神经元和星形胶质细胞的功能在微出血附近暂时丧失，但在病变后一天内恢复。
