Dendrotoxin (DTX), a well characterized IA-type potassium channel blocker, directly added to the culture medium had no effect on survival of cultured cortical neurons at 6 or 14 days in vitro. On the contrary, neurons exposed to DTX remained in better condition than untreated ones. In an attempt to demonstrate the mechanisms by which DTX may affect neuronal survival we studied its effect in co-cultures of cortical neurons and astrocytes submitted to successive medium changes. After the second change of medium, at 9 days in vitro, the neuronal number in controls decreased by 43%, while in cultures receiving astrocyte-conditioned medium the cell loss was significantly reduced (15%, p < 0.01) with respect to control conditions. When DTX was added to the culture medium neuronal loss was also significantly prevented (25% for 1 microM DTX, p < 0.01) with respect to control conditions. 4-Aminopyridine (4-AP) and 21 mM K+ also preserved neurons. The L-type calcium channel antagonist nifedipine (5 microM) abolished the protective effect of DTX and 4-AP. These results show that K+ channel blockade induces protection against damage produced by repetitive medium change and that this effect is mediated by L-type Ca2+ channels.