Comparative effects of perfusate composition on rat brain histology following transcardial perfusion fixation
Background: Transcardial perfusion fixation is widely used to preserve rodent brains for histological and ultrastructural analysis, but protocols can vary in the use of pre-fixation washout, fixative formulation, and osmotic additives. Relatively few studies have directly compared how the chemical composition of the perfusate affects tissue preservation. Methods: Male Sprague-Dawley rats…
The study investigates the impact of different perfusate compositions on the histology of rat brains after transcardial perfusion fixation. Transcardial perfusion is commonly used to preserve rodent brains for histological and ultrastructural analysis, but the effect of variations in perfusate composition has not been extensively studied.
Researchers utilized a range of aldehyde-based fixative formulations, which differed in their composition, utilized a phosphate buffered saline washout, and incorporated either mannitol or polyethylene glycol 35 kDa (PEG35). The outcomes of perfusion were evaluated using gross brain morphology, semi-quantitative grading of vascular blood clearance, detection of residual erythrocytes via whole-slide histological images, and measures of cellular visibility and morphology under light microscopy.
Additionally, perfusate osmolality was measured, and selected specimens were examined using electron microscopy.
The results indicated that substantial vascular blood clearance could be achieved through perfusion alone, suggesting that a pre-fixation washout was not necessary for achieving high levels of blood clearance under the tested conditions. Light microscopy-based measures of cellular visibility and morphology displayed a general similarity among the various treatment groups.
Incorporating mannitol and PEG35 into the fixative solution resulted in concentration-dependent gross tissue shrinkage, particularly with PEG35, yet did not produce significant or consistent changes on light microscopy. The study also highlighted the occurrence of considerable case-to-case variation in perfusion quality among animals undergoing similarly performed procedures.
The findings emphasize that while variations in perfusate composition can have noticeable effects on the gross brain morphology, their impact on light microscopy outcomes is relatively modest. Furthermore, the study highlights the importance of procedural factors on perfusion quality beyond the composition of the perfusate itself.
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