D-[3H]aspartate release from hippocampus slices studied in a multiwell system: controlling factors and postnatal development of release.

Medium components and various factors were tested to define optimal conditions for D-[3H]aspartate release. Isolation of the hippocampus and preparation of the slices in a medium without Ca2+ increased the release of D-[3H]aspartate in response to veratridine when subsequently tested in a regular Ca2+ containing medium. Apparently, the absence of Ca2+ during preparation of the slices reduced irreversible damage due to hypoxic conditions which prevail throughout the interval between killing the animal and immersion of the slices in a well oxygenated medium. Substitution of 10 mM Mg2+ for Ca2+ was an efficient procedure to test for Ca2+ dependence of D-[3H]aspartate release induced by veratridine. The inhibition was readily reversible when Ca2+ was readded. Veratridine (50 microM) was superior to high K+ (45 mM) in inducing D-[3H]aspartate release under all conditions tested in slices of mature animals. Furthermore, veratridine-induced release could be completely blocked by tetrodotoxin while K+-induced release was essentially unaffected by this toxin. Postnatal development of the D-[3H]aspartate release induced by veratridine was found to require 40-45 days, whereas release induced by K+ reached about 80% of maximum at postnatal day 22. K+-induced release appears to reach maturation when most hippocampal cells have been formed while veratridine-induced release probably requires completion of the neural circuit, involving also extensive sodium channel formation. These investigations were conveniently performed using a modified plastic culture box in which 24 slice systems can be studied simultaneously.

Early phenobarbital-induced alterations in hippocampal acetylcholinesterase activity and behavior.

Early exposure to phenobarbital (PhB) causes marked destruction of large neurons which are then forming both in the hippocampus and in the cerebellum. Such exposure to PhB also reduces the achievements of mice in hippocampus-related behaviors such as radial 8-arm maze performance. Experimental evidence suggests that these behaviors are partially mediated by cholinergic transmission. We studied the performance of mice, exposed to PhB prenatally or neonatally, in radial 8-arm maze. Both treatments caused significant impairments in the animals' performance in the maze. Acetylcholinesterase (AChE) and pseudocholinesterase (pChE) activities were studied in the hippocampus and cerebellum of mice who were exposed to PhB prenatally or neonatally. These enzymes are involved both in cholinergic transmission and in neuronal development. A significant decrease (13-16%, P less than 0.01) in hippocampal AChE specific activity was found between days 15 and 22 in animals exposed to PhB neonatally. The total hippocampal activity of AChE was also greatly reduced (25-39%, P less than 0.01) during that period as a result of both the reduction in specific activity and a reduction in hippocampal weight of the treated animals. These alterations were transient and were not detected in adulthood. No changes in hippocampal AChE or pChE activities were found in animals treated prenatally. Cerebellar AChE and pChE activities were not altered after prenatal nor after neonatal exposure to PhB. It is possible that the short-term effect of neonatal treatment on AChE specific activity might mediate the long-term impairments in hippocampus-related behaviors.