Experimental Evidence that Phenylalanine Provokes Oxidative Stress in Hippocampus and Cerebral Cortex of Developing Rats

High levels of phenylalanine (Phe) are the biochemical hallmark of phenylketonuria (PKU), a neurometabolic disorder clinically characterized by severe mental retardation and other brain abnormalities, including cortical atrophy and microcephaly. Considering that the pathomechanisms leading to brain damage and particularly the marked cognitive impairment in this disease are poorly understood, in the present study we investigated the in vitro effect of Phe, at similar concentrations as to those found in brain of PKU patients, on important parameters of oxidative stress in the hippocampus and cerebral cortex of developing rats. We found that Phe induced in vitro lipid peroxidation (increase of TBA-RS values) and protein oxidative damage (sulfhydryl oxidation) in both cerebral structures. Furthermore, these effects were probably mediated by reactive oxygen species, since the lipid oxidative damage was totally prevented by the free radical scavengers alpha-tocopherol and melatonin, but not by L-NAME, a potent inhibitor of nitric oxide synthase. Accordingly, Phe did not induce nitric oxide synthesis, but significantly decreased the levels of reduced glutathione (GSH), the major brain antioxidant defense, in hippocampus and cerebral cortex supernatants. Phe also reduced the thiol groups of a commercial GSH solution in a cell-free medium. We also found that the major metabolites of Phe catabolism, phenylpyruvate, phenyllactate and phenylacetate also increased TBA-RS levels in cerebral cortex, but to a lesser degree. The data indicate that Phe elicits oxidative stress in the hippocampus, a structure mainly involved with learning/memory, and also in the cerebral cortex, which is severely damaged in PKU patients. It is therefore presumed that this pathomechanism may be involved at least in part in the severe cognitive deficit and in the characteristic cortical atrophy associated with dysmyelination and leukodystrophy observed in this disorder.