Pb 2+-induced toxicity is associated with p53-independent apoptosis and enhanced by glutamate in GT1–7 neurons

Abstract

Recent studies indicate that the glutamatergic neurotransmitter system is involved in neurotoxicity caused by inorganic lead (Pb 2+). We studied the role of apoptosis in the effects induced by Pb 2+ (0.01–100 μM) and glutamate (0.1 and 1 mM) in mouse hypothalamic GT1–7 neurons. Although glutamate alone had no effect on cell viability, it enhanced neuronal cell death induced by Pb 2+ (1–100 μM) within 72 h. Glutamate alone neither induced caspase-3-like protease activity nor promoted internucleosomal DNA fragmentation, both biochemical hallmarks of apoptosis. However, concurrent exposure to Pb 2+ (10 or 100 μM) and glutamate (1 mM) resulted in more prominent cleavage of the fluorogenic caspase-3 substrate (Ac-DEVD-AMC) than caused by the same Pb 2+ concentrations alone at 24–72 h. The highest caspase-3-like protease activities were measured at 48 h. Internucleosomal DNA fragmentation caused by Pb 2+ (10 or 100 μM) alone or together with glutamate (1 mM) was evident at 96 h, less clear at 72 h and absent at 48 h. Immunoblotting did not reveal any changes in p53 protein levels in cells exposed to Pb 2+, glutamate or their combination at any studied time point (3–72 h). Our results suggest that Pb 2+-induced neurotoxicity may partially be mediated through p53-independent apoptosis and enhanced by glutamate.