The levels of the N-methyl-D-aspartate subclass of glutamate receptor were determined in organotypic neocortical explants chronically exposed to a growth medium containing 25 mM potassium (K25). Explants exposed to 25 mM potassium for 2-3 weeks evinced significantly less binding of the non-competitive N-methyl-D-aspartate receptor-associated channel antagonist 125I-MK801 than did age-matched controls. Surprisingly, cultures that were returned to control growth medium for a further 2 or 7 days showed even less binding of the ligand. The Kd values of binding were not affected and were similar to those of fresh postnatal cortex. The maximum number of binding sites did not vary between postnatal day 6 and 14 days in vitro control cultures, but were significantly less than those measured at postnatal day 20 (comparable age: 14 days in vitro). Several conclusions can be drawn from these findings: (i) the density of the N-methyl-D-aspartate subclass glutamate receptor does not attain in vivo levels under the present culturing conditions, but remains at those levels associated with the stage of development at which the tissue was brought into culture, (ii) chronic depolarization results in a drastic reduction in N-methyl-D-aspartate receptor density, which is not compensated for after the return to normal growth conditions, (iii) depolarization selectively inhibits cellular maturation of the neocortex (but not survival, as shown previously), including neurotransmitter receptor production and/or insertion into membranes or assembly.