title: Evaluation of three tracers for labeling distal cerebrospinal fluid-contacting neurons.
abstract: It has been reported that distal cerebrospinal fluid-contacting neurons (dCSF-CNs) can be detected by immunohistochemical assay using cholera toxin subunit B-conjugated horseradish peroxidase (CB-HRP). In the present study, another two methods with CB alone or CB-conjugated FITC (CB-FITC) were used, and the results from the three methods were compared. Adult Sprague-Dawley rats were randomly divided into three groups with CB-HRP, CB or CB-FITC. Tracers were diluted to 30% in artificial cerebrospinal fluid and injected separately (in a volume of 3 μL) into the lateral ventricle. Animals from the CB-HRP and CB groups were perfused 48 h after surgery while animals from the CB-FITC group were perfused 1, 3, 6, 12, 24 or 48 h after surgery. The brain was sectioned (40 μm) for immunofluorescence and five sections with positive neurons were selected from each rat for neuron counting. Three clusters of positive neurons in a 'Y-like' distribution were detected ventral to the cerebral aqueduct of rats from the three groups. No significant difference was observed among the quantitative data. In the CB-FITC group, stable staining was detected even at 6 h after injection. Taken together, lateral ventricle injection of CB/CB-FITC is a useful method for labeling dCSF-CNs in rats. The CB-FITC method makes dCSF-CNs labeling much simpler and more convenient.
authors.affiliation: Department of Anesthesiology, The First Clinical College, China Medical University, Shenyang, 110001, China.
authors.forename: Fang
authors.initials: F
authors.lastname: Zhou
authors.forename: Jiayou
authors.initials: J
authors.lastname: Wang
authors.forename: Hongxing
authors.initials: H
authors.lastname: Zhang
authors.forename: He
authors.initials: H
authors.lastname: Liu
authors.forename: Guangping
authors.initials: G
authors.lastname: Zhao
authors.forename: Cuihua
authors.initials: C
authors.lastname: Zu
authors.forename: Xiaoxing
authors.initials: X
authors.lastname: Lu
authors.forename: Licai
authors.initials: L
authors.lastname: Zhang
mesh_terms: D000818:Animals; D002555:Cerebrospinal Fluid; D002772:Cholera Toxin; D005455:Fluorescent Antibody Technique / Q000379:methods*; D005456:Fluorescent Dyes; D006735:Horseradish Peroxidase; D007091:Image Processing, Computer-Assisted; D008297:Male; D009474:Neurons / Q000166:cytology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D005456:Fluorescent Dyes; C052635:cholera toxin, B subunit-horseradish peroxidase; D002772:Cholera Toxin; D006735:Horseradish Peroxidase