Establishment and properties of separate cultures of enteric neurons and enteric glia

Abstract In this paper methods are described for the preparation of two types of culture derived from myenteric explants: (a) highly enriched neuronal cell cultures, and (b) purified glial cells (>98%). Both procedures combine the technique of antibody complement-mediated cytolysis with the use of an antimitotic agent. Immunohistochemical methods were used to compare the purified cells to their counterparts in mixed cultures (see accompanying paper). Antibodies to the glycoprotein Thy-1 and the monoclonal antibody A2B5 which recognizes gangliosides, labelled the cell surface of all enteric neurons in enriched cultures while subpopulations of the neurons expressed the Leu 7 carbohydrate epitope, the neurotransmitter 5-hydroxytryptamine and the neuropeptides substance P, methionine-enkephalin and vasoactive intestinal polypeptide. Autoradiographic experiments show that a subpopulation of enriched neurons exhibit high-affinity uptake sites for γ-[ 3 H]aminobutyric acid (GABA). All purified enteric glia continue to express the calcium binding protein S100, the basement membrane glycoprotein laminin and the antigens recognized by the A2B5 antibody, and subpopulations of glia are labelled by the monoclonal antibodies LB1 which binds to GD3 gangliosides, and Leu 7. Thus enteric neurons and glia can survive independently of each other and express molecular properties which are present in cultures normally containing both cell types.