Alterations in Norepinephrine and Dopamine Content in Selected Brain Areas of Guinea Pigs Adapted to Simulated High Altitude

Abstract: The alterations in brain content of norepinephrine (NE) and dopamine (DA) were studied in guinea pigs adapted to simulated high altitude (hypobaric hypoxia) equivalent to 5500 meters. The animals were adapted for 46 days over a period of 82 days to a pressure of 375 mm Hg. The animals were then killed and the following brain parts dissected: cerebellum, neocortex, caudate head/basal forebrain, diencephalons/rhinencephalon, and brain stem. NE and DA content were analyzed by high pressure liquid chromatography with electrochemical detection by a technique described. Results showed a significant increase of NE and DA in neocortex; a significant increase of DA but not NE in caudate/basal forebrain, and a significant decrease of NE and not DA in diencephalons/rhinencephalon.     

AMPA-Induced Excitotoxicity Increases Nuclear Levels of CAD, Endonuclease G, and Acinus and Induces Chromatin Condensation in Rat Hippocampal Pyramidal Neurons

Programmed cell death has been linked to AMPA-receptor-mediated excitotoxicity in pyramidal neurons of the hippocampus. The intent of this study was to investigate the roles of caspase-dependent and independent nuclear death-related factors in mediating AMPA-induced nuclear changes in PyNs by use of immunohistochemistry and transmission electron microscopy (TEM). Data indicate increases in the nuclear levels of caspase-activated acinus and DNase and Endonuclease G (a caspase-independent endonuclease) in CA1 and CA3 PyN nuclei with different temporal patterns following an AMPA-insult. Hoechst staining and TEM confirm AMPA-induced chromatin condensation. The presence of active acinus in nuclei suggests it mediates chromatin condensation. Interestingly, a DNA fragmentation labeling protocol showed that there was no chromatin cleavage up to 90 min after AMPA-insult. Overall, we conclude that: 1) AMPA-induced excitotoxicity increases nuclear immunoreactivity of pro-death enzymes from multiple programmed cell death pathways, 2) differential chromatin condensation patterns occur between CA1 and CA3, and 3) there is no chromatin cleavage within our experimental timeframe. Abbreviations: AIF, apoptosis inducing factor; AMPA, α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid; CAD, caspase-activated DNase; CIP, calf intestinal alkaline phosphatase; EndoG, endonuclease G; ICAD, inhibitor of CAD; NMDA, N-methyl D-aspartate; TdT, terminal deoxynucleotidyl transferase; TEM, transmission electron microscopy; TUNEL, terminal deoxynucleotidyl transferase biotin-UTP nick end labeling     

AMPA Receptor-Mediated Alterations of Intracellular Calcium Homeostasis in Rat Cerebellar Purkinje Cells In Vitro: Correlates to Dark Cell Degeneration

In the rat cerebellar slice preparation in vitro, excessive DL-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA)-receptor activation elicits a characteristic type of excitotoxicity of Purkinje cells (PCs) known as dark cell degeneration (DCD). DCD models neurotoxicity of PCs and hippocampal pyramidal neurons in vivo following hyperexcitable states. The intent of this study was to: a) determine whether AMPA-induced neurotoxicity of PCs is correlated with temporally and spatially restricted rises in intracellular Ca²⁺ and b) whether GYKI 52466 and nominal external Ca²⁺, conditions that reduced expression of AMPA-elicited DCD, altered the induced Ca²⁺ patterns. Employing the Ca²⁺-sensitive dye Fluo-3 and a confocal laser scanning microscope, we evaluated changes in intracellular Ca²⁺ within PCs in a cerebellar slice preparation. AMPA application alone (30 M for 30 min) caused a significant initial rise in perinuclear and cytoplasmic Ca²⁺ that returned to control levels during the latter part of the AMPA exposure period. Following removal of AMPA (expression period), perinuclear and cytoplasmic Ca²⁺ displayed a significant delayed rise peaking transiently 60 min after AMPA removal. The efficacy of GYKI 52466 and nominal external Ca²⁺ conditions to attenuate AMPA-induced DCD was correlated to reductions in AMPA-induced transient elevations in perinuclear and cytoplasmic Ca²⁺ levels during the expression phase and to a lesser extent during the exposure period. The present data suggest that during the expression phase, the delayed perinuclear and cytoplasmic Ca²⁺ transient may be the harbinger of impending loss of Ca²⁺ homeostasis and cell damage.