Selective neurotoxicity induced by lasalocid in dissociated cerebral cultures

Dispersed cerebral cells prepared from 15-16-day mouse foetuses were cultured for 7 days and exposed to lasalocid for periods of 4, 18 or 48 hr. Cultures were examined by phase contrast microscopy and processed for scanning electron microscopy. Lasalocid (1 and 2 mum) induced neurotoxic effects that were evident already at 4 hr, including swelling of perikarya, followed by cytolysis of most neurons present in the cultures. During the following 18 hr virtually the entire neuronal population degenerated completely. Cultures exposed to a lower concentration of lasalocid (0.5 mum) for 48 hr showed only mild damage to some neurons, and at 0.2 mum no damage could be observed, even after several days of continuous exposure. Notably, glial and other non-neuronal cells were not damaged by exposure to 2 mum-lasalocid, and cell division resumed on returning the cultures to regular growth media. Similarly, lasalocid (2 mum) was not toxic to cultured rat astrocytes. These morphological observations were followed by measurements of (45)Ca(2+) influx in the dissociated cerebral cultures. Lasalocid (1 mum) induced (45)Ca(2+) influx of 40% above dimethyl sulphoxide control values. The voltage-sensitive calcium channel antagonists nimodipine (10 mum) and D-600 (50 mum) did not inhibit lasalocid-mediated (45)Ca(2+) influx, whereas MK-801 [10 mum; a non-competitive N-methyl-d-aspartate (NMDA) receptor/channel antagonist] exclusively blocked this influx and prevented cytotoxic damage. Conversely, NMDA and glutamate, which by themselves mediated (45)Ca(2+) influx in these cultures, both potentiated lasalocid-induced (45)Ca(2+) influx and cellular damage. These observations clearly demonstrate the selective neurotoxicity of lasalocid to cultured cerebral neurons, and may imply involvement of the NMDA receptor/channel in lasalocid-mediated neurotoxicity.