Levodopa, which is used in the treatment of Parkinson's disease, has known cytotoxic effects on dopaminergic neurons grown in culture. Calretinin (CR) is a cytosolic calcium-binding protein found in specific subpopulations of neurons as well as in some nonneuronal tissue. CR is expressed in 10% of rat embryo dopaminergic neurons grown in vitro. Since it has been postulated that CR provides neuroprotection due to its calcium-binding properties, we investigated whether CR-containing dopaminergic neurons were spared from levodopa toxicity. Incubation of mesencephalic cells with 10(-5) to 10(-7) M levodopa on Days 1-6 in vitro produced no significant effects on the number of dopaminergic neurons containing CR, but resulted in the loss of approximately 65% of the dopaminergic cells which did not contain CR. The remaining CR-negative dopaminergic neurons exhibited dose-dependent reductions in neurite length. The neuronal processes in CR-containing dopaminergic cells retained a smooth bipolar appearance. CR-immunoreactive cells which did not contain dopamine showed slight neurite length decreases at the highest drug concentrations but no changes in neuron number. These results indicate that CR may protect dopaminergic neurons from levodopa-induced toxicity.