Laser scanning and electron microscopic evidence for rapid and specific invivo labelling of cholinergic neurons in the rat basal forebrain with fluorochromated antibodies

Recently developed methods for the selective labelling of cholinergic basal forebrain neurons containing the low-affinity neurotrophin receptor p75 (p 75(NTR)) in vivo and in vitro are based on carbocyanine 3 (Cy3)-tagged anti bodies directed against p75(NTR) The present study focuses on the maintenan ce of this neuronal label after injection of such fluorescent antibodies in to the cerebral ventricle. One, 3, and 10 days after injection this marker exclusively stains neurons immunoreactive for the cholinergic markers choli ne acetyltransferase and vesicular acetylcholine transporter in the rat med ial septum, diagonal band and nucleus basalis. Thirty clays after injection the in vivo labelling was nearly abolished. Predominant labelling of lysos omes was shown by electron microscopic analysis following photoconversion o f the Cy3-label to an electron-dense reaction product. The pre-labelling of cholinergic neurons might facilitate pharmacological and electrophysiologi cal approaches in living slices and cell culture systems as well as detaile d investigations focused on the transport of neurotrophins in vivo and in a nimals with experimentally altered p75(NTR) expression. (C) 2000 Elsevier S cience B.V. All rights reserved.