Nissl stain
In order to quantify the total number of neurons in the dorsal
hippocampus, Nissl staining was performed on brain
sections prepared using the same fixation and cutting
protocol described for immunohistochemical procedures
(see below). Sections were thoroughly rinsed with PBS,
put on gelatin-treated slides and air-dried overnight.
Then, slides were briefly washed in distilled water and
then immersed in a 0.5% Cresyl Violet solution at 37 C
for 10 min. Afterward, they were washed rapidly in distilled water and dehydrated through a series of graded
ethanol (95% 2 30 min and 100% 2 5 min each) and
cleared in xylene (2 5 min each). Slides were
coverslipped using Vectamount (Vector Labs, Inc.,
Burlingame, CA, USA).