Journal of neurochemistry
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Journal of neurochemistry · Jul 2004
Comparative StudyDifferential contribution of plasmalemmal Na/Ca exchange isoforms to sodium-dependent calcium influx and NMDA excitotoxicity in depolarized neurons.
Inhibition of Na(+),K(+)-ATPase during NMDA applications greatly increased NMDA-induced excitotoxicity in primary cultures of forebrain neurons (FNs), but not in cerebellar granule cells (CGCs). Because Na(+),K(+)-ATPase inhibition promotes reversal of plasmalemmal Na(+)/Ca(2+) exchangers, we compared the activities of reversed K(+)-independent (NCX) and K(+)-dependent (NCKX) Na(+)/Ca(2+) exchangers in these cultures. To this end, we measured gramicidin-induced and Na(+)-dependent elevation in cytosolic [Ca(2+)] ([Ca(2+)](c)) that represents Ca(2+) influx via reversed NCX and NCKX; NCX activity was dissected out by removing external K(+). ⋯ We tested whether the NCX or NCKX family of exchangers contributes most to the toxic NMDA-induced Ca(2+) influx in depolarized neurons. We found that in FNs, inhibition of NCX alone was sufficient to significantly limit NMDA excitotoxicity, whereas in CGCs, inhibition of both NCX and NCKX was required. The data suggest that the high activity of NCX isoforms expressed in FNs, possibly NCX1, sensitizes these neurons to NMDA excitotoxicity.
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Journal of neurochemistry · Jul 2004
Long-term regulation of signalling components of adenylyl cyclase and mitogen-activated protein kinase in the pre-frontal cortex of human opiate addicts.
Opiate addiction involves the development of chronic adaptive changes in micro -opioid receptors and associated pathways (e.g. cAMP signalling) which lead to neuronal plasticity in the brain. This study assessed the status of cAMP and mitogen-activated protein kinase (MAPK) pathways in brains (pre-frontal cortex) of chronic opiate addicts. In these subjects (n = 24), the immunodensities of adenylyl cyclase-I, PKA Calpha, total and phosphorylated CREB were not different from those in sex-, age- and PMD-matched controls. ⋯ Chronic morphine treatment in rats (10-100 mg/kg for 5 days) was also associated with decreases of pERK1/2 (59-68%) in the cortex. In SH-SY5Y cells, morphine also stimulated the activity of pERK1/2 (2.5-fold) and the MEK inhibitor PD98059 blocked this effect (90%). The abnormalities of MAPK signalling might have important consequences in the long term development of various forms of neural plasticity associated with opiate addiction in humans.
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Journal of neurochemistry · Jul 2004
Shift in the localization of sites of hydrogen peroxide production in brain mitochondria by mitochondrial stress.
We have determined the underlying sites of H(2)O(2) generation by isolated rat brain mitochondria and how these can shift depending on the presence of respiratory substrates, electron transport chain modulators and exposure to stressors. H(2)O(2) production was determined using the fluorogenic Amplex red and peroxidase system. H(2)O(2) production was higher when succinate was used as a respiratory substrate than with another FAD-dependent substrate, alpha-glycerophosphate, or with the NAD-dependent substrates, glutamate/malate. ⋯ In addition, three forms of in vitro mitochondrial stress were studied: Ca(2+) overload, cold storage for more than 24 h and cytochrome c depletion. In each case we observed (i) a decrease in succinate-supported H(2)O(2) production by complex I and an increase in succinate-supported H(2)O(2) production by complex III, (ii) increased glutamate/malate-induced H(2)O(2) generation by complex I and (iii) increased alpha-glycerophosphate-supported H(2)O(2) generation by complex III. Our results suggest that all three forms of mitochondrial stress resulted in similar shifts in the localization of sites of H(2)O(2) generation and that, in both normal and stressed states, the level and location of H(2)O(2) production depend on the predominant energetic substrate.
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Journal of neurochemistry · Jul 2004
Regulation of RANTES/CCL5 expression in human astrocytes by interleukin-1 and interferon-beta.
In the CNS, astrocytes are significant sources of RANTES/CCL5 (regulated upon activation, normal T cell expressed and secreted), a CC-chemokine with important biological function. Astrocyte RANTES/CCL5 has been shown to be induced by interleukin-1 (IL-1), with interferon-gamma (IFNgamma) as a primer, but whether type I interferons play any role in the expression of RANTES/CCL5 is not known. In this report, we studied the detailed mechanism of RANTES/CCL5 induction in primary human astrocytes activated with IL-1 and IFNbeta. ⋯ IFNbeta synergized with IL-1 to induce nuclear accumulation of C/EBPbeta protein. They also synergized to form nuclear ISRE complexes with Stat1, Stat2 and interferon regulatory factor-1 (IRF-1) proteins. Together, our results demonstrate that IFNbeta plays a positive regulatory role in the expression of RANTES/CCL5 in human astrocytes through several distinct mechanisms.