A: overview of a frontal section at the
level of the medulla oblongata showing a typical iontophoretic injection
site in the nucleus reticularis parvicellularis (case K120) using a 2-µA
pulsed positive current for 30 min.
D: adjacent section to that shown in
B at a higher magnification, counterstained with Cresyl violet and immunostained
with serotonin, showing the presence of counterstained cell bodies and
serotonin immunoreactive fibers in the site proving the absence of tissue
C: photomicrograph illustrating retrogradely
labeled cells in the medial (left part) and lateral (right part) divisions
of the nucleus of the solitary tract. Note also the presence of punctate
anterograde labeling in the same areas.
E: photomicrograph showing anterogradely
labeled fibers organized as a bundle running dorsally to the facial nucleus
approximately 2 mm rostral to the site shown in A
and B. Varicose fibers inside the facial nucleus
sometimes formed basket-like structures around motoneuron somata (arrow).
To optimize the CTb immunostaining shown in C, E and F, the sections were incubated 3-4 days in CTb antiserum (1: 40,000) at 4 °C followed by sequential incubations overnight at 4 °C in the biotinylated donkey anti-goat IgG (1:2,000) and the streptavidin-HRP (1:40,000). Note that with such maximal conditions, we obtained a weak counterstaining of the sections ideally suited for accurate localization of CTb immunostained material.