FREEZE-BLOWING - NEW TECHNIQUE FOR STUDY OF BRAIN IN-VIVO
FREEZE-BLOWING - NEW TECHNIQUE FOR STUDY OF BRAIN IN-VIVO
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DOI：
10.1111/j.1471-4159.1973.tb12115.x
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发表时间：
1973-01-01
影响因子：
4.7
通讯作者：
VEECH, EH
中科院分区：
文献类型：
--
作者：
VEECH, RL;HARRIS, RL;VEECH, EH
描述了一种新的装置，它可以比以前更快地取出并冷冻清醒大鼠的大脑。该装置由两个探针组成，这两个探针被同时驱动到固定在一个特殊构造的限制笼中的大鼠的颅穹窿中。就位后，加压空气通过一个探针进入，将脑组织的幕上部分（位于嗅球和上级丘之间）吹出另一个探针，进入预先在液氮中冷却的薄室。这种方法比先前描述的微波辐射、断头进入液氮或整个动物浸入液氮的方法更快地停止脑组织代谢，如通过测量不稳定代谢物和氧化还原状态所证明的。因此，与通过其他方法获得的脑组织相比，冷冻吹制脑样品的a-酮戊二酸、磷酸肌酸、丙酮酸、葡萄糖和葡萄糖-6-磷酸水平较高，乳酸、苹果酸和AMP水平较低。在冷冻吹制样品中，游离细胞质[NAD+]/[NADH 2]、[NADP+]/[NADPH 2]和[ATP]/[ADP] [HPO 42-]比值较高。这些数据表明，当使用除冷冻吹制以外的方法时，发生了更广泛的缺氧代谢。我们得出结论，采用冷冻吹扫技术获得的脑中测量的代谢物水平更接近体内代谢物水平。
A new apparatus is described which removes and freezes brains of conscious rats more rapidly than was heretofore possible. The apparatus consists of two probes which are driven simultaneously into the cranial vault of the rat immobilized in a specially constructed restraining cage. When in position, air under pressure enters through one probe and blows the supratentorial portion of the brain tissue (situated between the olfactory bulbs and the superior colliculi) out the other probe and into a thin chamber previously cooled in liquid N2. This method stops brain tissue metabolism more rapidly than the previously‐described methods of microwave irradiation, decapitation into liquid N2, or whole‐animal immersion into liquid N2, as evidenced by the measurement of labile metabolites and redox states. Thus, samples of freeze‐blown brain had higher levels of a‐oxoglutarate, creatine phosphate, pyruvate, glucose and glucose‐6‐phosphate and lower levels of lactate, malate and AMP than brain tissue obtained by the other methods. The free cytoplasmic [NAD+]/[NADH2], [NADP+]/[NADPH2] and [ATP]/[ADP] [HPO42‐] ratios were higher in freeze‐blown samples. These data indicate that more extensive anoxic metabolism occurred when methods other than freeze‐blowing were used. We conclude that the levels of metabolites measured in brain obtained with the freeze‐blowing technique more closely resemble those which occurin vivo.