A Novel Method to Isolate Odontoblasts from Rat Incisor

Abstract

Historically, odontoblasts have been isolated from rat incisor using a surgical curette to separate these cells from the dentin. Isolation of odontoblasts using this approach typically resulted in cells with membrane properties that made the application of patch-clamp electrophysiological techniques prohibitive. The studies here describe a new procedure for isolating mature odontoblasts from adult rat incisor to obtain enriched populations of intact, viable odontoblasts that can be readily studied using patch-clamp methodologies. Identification of isolated cells as odontoblasts was confirmed using in situ mRNA hybridization for expression of dentin sialoprotein, osteocalcin, bone sialoprotein, and type I collagen, and calcium flux was monitored in these cells by means of fura-2 microfluorometry. We suggest that either single odontoblasts or clusters of these cells isolated by this new method would be an ideal preparation for the study of odontoblast properties using electrophysiological techniques, in situ hybridization and/or microfluorometry.