The acetylcholine precursor choline is transported into cholinergic neurons by a high-affinity, sodium-dependent mechanism that is selectively localized to the cholinergic nerve terminal. In addition, a low-affinity, sodium-independent choline uptake system is present in cholinergic and non-cholinergic cells which deliver choline for cell membrane anabolism. Here, we show that uptake of [3H]choline into cultured fibroblast cell lines exhibits high affinity (Km < or = 10 microM), is sodium-dependent, and is blocked by hemicholinium, a classical inhibitor of neuronal high-affinity choline uptake. Our data indicate that sodium-dependent high-affinity choline transport systems are also present in non-cholinergic cells.