Glutamate transport and metabolism in dopaminergic neurons of substantia nigra: implications for the pathogenesis of Parkinson's disease

Parkinson's disease (PD) is associated with degeneration of the pigmented d opaminergic neurons located in the ventral mesencephalon. Although the mech anisms by which these neurons degenerate in PD are poorly understood, indir ect evidence suggests involvement of glutamatergic mechanisms in the pathog enesis of this disorder. Glutamate, the major excitatory transmitter in the mammalian central nervous system, is known to be neurotoxic when present i n excess at the synapses. Two major mechanisms protect neurons from glutama te-induced toxicity: (a) removal of synaptic glutamate via a high affinity uptake carried out by cytoplasmic membrane proteins known as excitatory ami no acid transporters (EAAT); and (b) metabolism and recycling of glutamate by synaptic astrocytes via glutamine synthetase, an ATP-requiring reaction. However, when extra-cellular glutamate levels are high (0.5-1.0 mM), gluta mate metabolism may be shifted toward the ATP-generating oxidative deaminat ion (glutamate dehydrogenase)-TCA cycle pathway. We have cloned and charact erized two human glutamate dehydrogenases (GDH), one of which is nerve tiss ue specific. This isoenzyme requires ADP for its activity and it may become functional when cellular energy charge is low. We have also cloned three h uman glutamate transporters. One of these (EAAT3) is neuron specific. In si tu hybridization studies using human brain revealed that the pigmented dopa minergic neurons, which degenerate in PD, express EAAT3 at high levels. Pri mary nerve tissue cultures derived from rat ventral mesencephalon were esta blished and studied for their ability to metabolize glutamate. Results show ed that mature cultures expressing high levels of CDH activity were capable of rapidly utilizing glutamate added to the medium at high concentrations (1-1.2 mM). This was associated with little release of aspartate and alanin e into the medium. In contrast, immature cultures expressing low GDH activi ty utilized glutamate at lower rates while releasing substantial amounts of aspartate and alanine into the medium. These data suggest that immature me sencephalic cells metabolize a substantial fraction of the glutamate they t ake up from the medium via the transamination pathway, compared to mature m esencephalic cultures. Immunocytochemical studies on these cultures reveale d that dopaminergic neurons (identified by their tyrosine hydroxylase conte nt) showed intense staining for GDH. Furthermore, inhibition of GDH express ion by antisense oligonucleotides was toxic to cultured mesencephalic neuro ns, with dopaminergic neurons being affected at the early stages of this in hibition. Hence, the dense expression by dopaminergic neurons of proteins i nvolved in the transport and metabolism of glutamate may serve particular b iological needs intrinsic to these cells. Further studies are required to t est whether these properties render these neurons vulnerable to excitotoxic mechanisms or to abnormalities of glutamate metabolism.