ENRICHMENT OF THE FRACTION OF NOCICEPTIVE NEURONS IN CULTURES OF PRIMARY SENSORY NEURONS

A method for enriching the fraction of nociceptive neurones in culture s of primary sensory neurones is described. Neurones from neonatal rat dorsal root ganglia were isolated, layered on 40% Ficoll and centrifu ged, separating the neurones into a low density fraction (LDF) and a h igh-density fraction (HDF). The LDF had a smaller mean diameter (19.7 mu m) than the HDF (27.3 mu m) and uncentrifuged cells (23.6 mu m). Th e proportion of cells immunoreactive for antibodies to substance P and calcitonin gene-related peptide, both of which are found in nocicepti ve neurones, was significantly greater in the LDF than in HDF. A subst antial enrichment of the proportion of neurones responding to the algo genic substances capsaicin and bradykinin with an increase in intracel lular calcium was observed in the LDF. The proportion of capsaicin and bradykinin-responsive neurones was also found to be increased by cult uring the neurones, with a particularly pronounced enhancement in the proportion of bradykinin-responsive cells. We conclude that separation on the basis of density followed by culture is a useful way of enrich ing the proportion of nociceptive neurones for the purpose of electrop hysiological, biochemical or other studies. (C) 1997 Elsevier Science B.V.