dinitrobenzenes and Brain-Diseases

Exposure to 1,3-dinitrobenzene (DNB) is associated with neuropathologic changes in specific brainstem nuclei, mediated by oxidative stress and mitochondrial dysfunction. The expression of Bcl-2-family proteins as a function of sensitivity to 1, 3-dinitrobenzene (DNB)-induced mitochondrial permeability transition (MPT) was examined in C6 glioma and SY5Y neuroblastoma cells. Neuroblastoma cells were 10-fold more sensitive than glioma cells to DNB-induced decreases in mitochondrial reducing potential, measured by reduction of the tetrazolium compound, 3-[4, 5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT). The IC(50) values for DNB-related inhibition of MTT reduction were 107+/-25 microM in SY5Y cells and 1047+/-101 microM in C6 cells. Levels of reactive oxygen species (ROS) were increased in both SY5Y and C6 cells following DNB exposure by 4.6- and 6.0-fold above control, respectively. DNB caused abrupt depolarization of mitochondria in both neuroblastoma and glioma cells that was inhibited by trifluoperazine. The first order rate constants for mitochondrial depolarization were: C6, k=0.31+/-0.02 min(-1); SY5Y, k=0.14+/-0.01 min(-1). Onset of MPT occurred at 10-fold lower concentration of DNB in SY5Y cells than in C6 cells. The antioxidants, deferoxamine and alpha-tocopherol, effectively prevented DNB-induced MPT in C6 and SY5Y cells, suggesting involvement of ROS in the initiation of MPT. Exposure to DNB resulted in decreased cellular ATP content in SY5Y cells and efflux of mitochondrial calcium in both SY5Y and C6 cells, concurrent with onset of MPT. The expression of Bcl-2, Bcl-X(L), and Bax was evaluated in both cell types by Western blot analysis. C6 glioma cells strongly expressed Bcl-X(L) and only weakly expressed Bcl-2 and Bax, whereas SY5Y neuroblastoma cells expressed lower levels of Bcl-X(L) and higher levels of both Bcl-2 and Bax. Collectively, these results suggest that higher constitutive expression of Bcl-X(L), rather than Bcl-2, correlates with resistance to DNB-induced MPT in SY5Y and C6 cells and that differential regulation of the permeability transition pore may underlie the cell-specific neurotoxicity of DNB.

Topics: Adenosine Triphosphate; Antioxidants; bcl-2-Associated X Protein; bcl-X Protein; Brain Diseases; Calcium; Deferoxamine; Dinitrobenzenes; Electrophysiology; Glioma; Humans; Mitochondria; Neuroblastoma; Oxidation-Reduction; Permeability; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species; Tetrazolium Salts; Thiazoles; Trifluoperazine; Tumor Cells, Cultured; Vitamin E

m-Dinitrobenzene (m-DNB) is an industrial chemical causing gliovascular lesions in the brain stem similar to those produced by nitroimidazoles and by thiamine deficiency. To identify early preneuropathic indices of toxicity we examined the action of m-DNB on glycolysis and on measures of oxidative stress in the brain both in vivo and in vitro. Significant increases in local cerebral glucose utilization were seen in 14 of 30 brain regions prior to development of lesions. Rat brain astrocyte cultures also showed increases in both glucose consumption and lactic acid formation in the first 24 h following exposure to 0.5 mM m-DNB and prior to the development of cytotoxicity. The concentration of reduced glutathione in these cultures was decreased to about half of control values over a 2-h incubation period, indicating an early disturbance of redox balance. The rate of reduction of nitroblue tetrazolium increased eightfold during a 1-h incubation period, suggesting a free radical-mediated process. Superoxide dismutase partially prevented this increase, although other protective agents failed to do so possibly due to lack of cellular penetration. These observations show that m-DNB neurotoxicity involves early metabolic stimulation and redox disruption that may be causally associated with the production of free radicals.

Topics: Animals; Animals, Newborn; Astrocytes; Blood-Brain Barrier; Brain; Brain Diseases; Cell Death; Cells, Cultured; Dinitrobenzenes; Glutathione; Glycolysis; Kinetics; Lactates; Lactic Acid; Nitroblue Tetrazolium; Oxidative Stress; Rats; Rats, Inbred F344; Time Factors

Using a 3 x 10 mg/kg dose schedule of 1,3-dinitrobenzene (DNB) over two days in Fischer rats, we have found the following changes in vascular function and structure during the early phase of the symmetrical brain stem lesions. 1. Marked increase in cerebral blood flow generally but especially in the inferior colliculi, from 6 h after the final dose of DNB. 2. Increasing incidence of petechial haemorrhages in inferior colliculi, cerebellar roof, vestibular and superior olivary nuclei from 12 h. 3. Focal leakage of horseradish peroxidase and many sleeve-like arteriolar haemorrhages seen in vibratome sections and by scanning electron microscopy (SEM) in these regions from 12 h. 4. Periarteriolar oedema and protein leakage present in step-serial sections in these regions from 12 h, with astrocyte swelling and occasional small infarcts. These changes suggest that the vascular bed may play an important role in the pathogenesis of these lesions, perhaps in parallel with early astroglial damage. They are discussed in relation to (i) the known presence of xanthine oxidase in the vascular bed of the brain and the likelihood of "useless redox cycling' with free radical generation from this enzyme's interaction with nitroheterocyclic compounds, and (ii) the possible role of free radical damage to endothelial cells in this intoxication and in the analogous lesions of natural and experimental Wernicke's encephalopathy.

Topics: Animals; Brain Diseases; Brain Stem; Cerebral Hemorrhage; Cerebrovascular Circulation; Dinitrobenzenes; Horseradish Peroxidase; Male; Rats; Rats, Inbred F344