Neurobiology of tobacco smoking and other addictive disorders.

Advances in research of the neurobiology of addictive disorders have provided clinicians with an evolving perspective on addiction. All drugs of abuse seem to share a common neurobiologic substrate involving the mesocorticolimbic system. Considerable evidence shows that these dopaminergic projections are involved in the positive brain reward, which drives addictive disorders; however, recent studies also implicate the neurotransmitters glutamate and serotonin in learning and sensitization to drug use. A review of the neurobiology of tobacco smoking provides further examples of the mechanisms for reinforcing tobacco use, including the enhancement of memory and treatment of depression with nicotine and MAO-inhibiting chemicals in tobacco smoke respectively. The relevance of these advances may be realized through the destigmatization of addictive disorders and the development of new and improved treatment strategies.

Role of pyruvate carboxylase in facilitation of synthesis of glutamate and glutamine in cultured astrocytes.

CO2 fixation was measured in cultured astrocytes isolated from neonatal rat brain to test the hypothesis that the activity of pyruvate carboxylase influences the rate of de novo glutamate and glutamine synthesis in astrocytes. Astrocytes were incubated with 14CO2 and the incorporation of 14C into medium or cell extract products was determined. After chromatographic separation of 14C-labelled products, the fractions of 14C cycled back to pyruvate, incorporated into citric acid cycle intermediates, and converted to the amino acids glutamate and glutamine were determined as a function of increasing pyruvate carboxylase flux. The consequences of increasing pyruvate, bicarbonate, and ammonia were investigated. Increasing extracellular pyruvate from 0 to 5 mM increased pyruvate carboxylase flux as observed by increases in the 14C incorporated into pyruvate and citric acid cycle intermediates, but incorporation into glutamate and glutamine, although relatively high at low pyruvate levels, did not increase as pyruvate carboxylase flux increased. Increasing added bicarbonate from 15 to 25 mM almost doubled CO2 fixation. When 25 mM bicarbonate plus 0.5 mM pyruvate increased pyruvate carboxylase flux to approximately the same extent as 15 mM bicarbonate plus 5 mM pyruvate, the rate of appearance of [14C] glutamate and glutamine was higher with the lower level of pyruvate. The conclusion was drawn that, in addition to stimulating pyruvate carboxylase, added pyruvate (but not added bicarbonate) increases alanine aminotransferase flux in the direction of glutamate utilization, thereby decreasing glutamate as pyruvate + glutamate --> alpha-ketoglutarate + alanine. In contrast to previous in vivo studies, the addition of ammonia (0.1 and 5 mM) had no effect on net 14CO2 fixation, but did alter the distribution of 14C-labelled products by decreasing glutamate and increasing glutamine. Rather unexpectedly, ammonia did not increase the sum of glutamate plus glutamine (mass amounts or 14C incorporation). Low rates of conversion of alpha-[14C]ketoglutarate to [14C]glutamate, even in the presence of excess added ammonia, suggested that reductive amination of alpha-ketoglutarate is inactive under conditions studied in these cultured astrocytes. We conclude that pyruvate carboxylase is required for de novo synthesis of glutamate plus glutamine, but that conversion of alpha-ketoglutarate to glutamate may frequently be the rate-limiting step in this process of glutamate synthesis.

Glucose transporter isoform expression in Huntington's disease brain.

Several reports have suggested a characteristic decrease in glucose use in the striatum of patients with Huntington's disease (HD) may contribute to the cellular atrophy of the caudate and putamen. We examined the expression of the two major glucose transporter isoforms of brain, GLUT1 and GLUT3. GLUT1 is found largely in capillary endothelial cells and to a lesser extent in the brain parenchyma, whereas GLUT3 is localized primarily in neurons. Membranes prepared from postmortem samples of HD caudate and cortex and non-HD caudate and cortex were separated on 10% sodium dodecyl sulfate-polyacrylamide gels and probed with antisera to GLUT1 and GLUT3 by western blotting. Compared with controls, GLUT1 and GLUT3 transporter expression in caudate was decreased by three- and fourfold, respectively, in grade 3 of the disease. At earlier stages (grade 1), there was no significant difference in the expression of the two transporter isoforms compared with nondiseased controls. It is surprising that despite a substantial increase in glial fibrillary acidic protein immunoreactivity (an indicator of the extent of gliosis), glucose transporter expression was diminished significantly in HD caudate. The results suggest in the absence of a significant number of neurons, as in grade 3, glial cell GLUT1 and GLUT3 expression is down-regulated, perhaps reflecting the decreased metabolic demand of this brain region in HD.