Infarct volume measurements

Brains were sliced coronally into 2 mm slices and incubated at 37°C for 20 min in 2,3,5-triphenyltetrazolium chloride (TTC; Liu et al., 2009). Brains were then fixed in formalin and then paraffin was embedded. Brain slices were photographed and infarct volume was measured using Image J (NIH.gov). Infarct area was calculated by tracing unstained regions in each section (both sides) and multiplied by slice thickness to calculate infarct volume. For measurements of cortical and subcortical infarction sizes, hematoxylin and eosin (H&E) slides were made from paraffin embedded brain sections. Cortical and subcortical structures were consistently identified using the Rat Brain Atlas as a reference (Swanson, 2018).