Neuronal Cell Death Induced by Mechanical Percussion Trauma in Cultured Neurons is not Preceded by Alterations in Glucose, Lactate and Glutamine Metabolism.

Traumatic brain injury (TBI) is a devastating neurological disorder that usually presents in acute and chronic forms. Brain edema and associated increased intracranial pressure in the early phase following TBI are major consequences of acute trauma. On the other hand, neuronal injury, leading to neurobehavioral and cognitive impairments, that usually develop months to years after single or repetitive episodes of head trauma, are major consequences of chronic TBI. The molecular mechanisms responsible for TBI-induced injury, however, are unclear. Recent studies have suggested that early mitochondrial dysfunction and subsequent energy failure play a role in the pathogenesis of TBI. We therefore examined whether oxidative metabolism of (13)C-labeled glucose, lactate or glutamine is altered early following in vitro mechanical percussion-induced trauma (5 atm) to neurons (4-24 h), and whether such events contribute to the development of neuronal injury. Cell viability was assayed using the release of the cytoplasmic enzyme lactate dehydrogenase (LDH), together with fluorescence-based cell staining (calcein and ethidium homodimer-1 for live and dead cells, respectively). Trauma had no effect on the LDH release in neurons from 1 to 18 h. However, a significant increase in LDH release was detected at 24 h after trauma. Similar findings were identified when traumatized neurons were stained with fluorescent markers. Additionally (13)C-labeling of glutamate showed a small, but statistically significant decrease at 14 h after trauma. However, trauma had no effect on the cycling ratio of the TCA cycle at any time-period examined. These findings indicate that trauma does not cause a disturbance in oxidative metabolism of any of the substrates used for neurons. Accordingly, such metabolic disturbance does not appear to contribute to the neuronal death in the early stages following trauma.

Neuron-glia interactions in glutamatergic neurotransmission: roles of oxidative and glycolytic adenosine triphosphate as energy source.

Glutamatergic neurotransmission accounts for a considerable part of energy consumption related to signaling in the brain. Chemical energy is provided by adenosine triphosphate (ATP) formed in glycolysis and tricarboxylic acid (TCA) cycle combined with oxidative phosphorylation. It is not clear whether ATP generated in these pathways is equivalent in relation to fueling of the energy-requiring processes, i.e., vesicle filling, transport, and enzymatic processing in the glutamatergic tripartite synapse (the astrocyte and pre- and postsynapse). The role of astrocytic glycogenolysis in maintaining theses processes also has not been fully elucidated. Cultured astrocytes and neurons were utilized to monitor these processes related to glutamatergic neurotransmission. Inhibitors of glycolysis and TCA cycle in combination with pathway-selective substrates were used to study glutamate uptake and release monitored with D-aspartate. Western blotting of glyceraldehyde-3-P dehydrogenase (GAPDH) and phosphoglycerate kinase (PGK) was performed to determine whether these enzymes are associated with the cell membrane. We show that ATP formed in glycolysis is superior to that generated by oxidative phosphorylation in providing energy for glutamate uptake both in astrocytes and in neurons. The neuronal vesicular glutamate release was less dependent on glycolytic ATP. Dependence of glutamate uptake on glycolytic ATP may be at least partially explained by a close association in the membrane of GAPDH and PGK and the glutamate transporters. It may be suggested that these enzymes form a complex with the transporters and the Na(+) /K(+) -ATPase, the latter providing the sodium gradient required for the transport process.

Role of astrocytic transport processes in glutamatergic and GABAergic neurotransmission.

The fine tuning of both glutamatergic and GABAergic neurotransmission is to a large extent dependent upon optimal function of astrocytic transport processes. Thus, glutamate transport in astrocytes is mandatory to maintain extrasynaptic glutamate levels sufficiently low to prevent excitotoxic neuronal damage. In GABA synapses hyperactivity of astroglial GABA uptake may lead to diminished GABAergic inhibitory activity resulting in seizures. As a consequence of this the expression and functional activity of astrocytic glutamate and GABA transport is regulated in a number of ways at transcriptional, translational and post-translational levels. This opens for a number of therapeutic strategies by which the efficacy of excitatory and inhibitory neurotransmission may be manipulated.