Mechanism of oxidative stress‐induced intracellular acidosis in rat cerebellar astrocytes and C₆ glioma cells

Abstract

Following ischaemic reperfusion, large amounts of superoxide anion (.O2−), hydroxyl radical (.OH) and H2O2 are produced, resulting in brain oedema and changes in cerebral vascular permeability. We have found that H2O2 (100 μm) induces a significant intracellular acidosis in both cultured rat cerebellar astrocytes (0.37 ± 0.04 pH units) and C6 glioma cells (0.33 ± 0.07 pH units). Two membrane‐crossing ferrous iron chelators, phenanthroline and deferoxamine, almost completely inhibited H2O2‐induced intracellular acidosis, while the non‐membrane‐crossing iron chelator apo‐transferrin had no effect. Furthermore, the acidosis was completely inhibited by two potent membrane‐crossing .OH scavengers, N‐(2‐mercaptopropionyl)‐grycine (N‐MPG) and dimethyl thiourea (DMTU). Since .OH can be produced during iron‐catalysed H2O2 breakdown (Fenton reaction), we have shown that a large reduction in pH1 in glial cells can result from the production of intracellular .OH via H2O2 oxidation. We have ruled out the possible involvement of: (i) an increase in intracellular Ca2+ levels; and (ii) inhibition of oxidative phosphorylation. Our results suggest that .OH inhibits glycolysis, leading to ATP hydrolysis and intracellular acidosis. This conclusion is based on the following observations: (i) in glucose‐free medium, or in the presence of iodoacetate or 2‐deoxy‐D‐glucose, H2O2‐induced acidosis is completely suppressed; (ii) H2O2 and iodoacetate both produce an increase in levels of intracellular free Mg2+, an indicator of ATP breakdown; and (iii) direct measurement of intracellular ATP levels and lactate production show 50 and 55% reductions in ATP content and lactate production, respectively, following treatment with 100 μm H2O2. Inhibition of the pH1 regulators (i.e. the Na+–H+ exchange and possibly the Na+–HCO3−–dependent pH1 transporters) resulting from H2O2‐induced intracellular ATP reduction may also be involved in the H2O2‐evoked intracellular acidosis in glial cells.

Mycothiol protects <i>Corynebacterium glutamicum</i> against acid stress via maintaining intracellular pH homeostasis, scavenging ROS, and <i>S</i>-mycothiolating MetE

“…5). Thus, MSH functions to assist in the scavenging of deleterious ROS, which is known to damage a wide range of biological molecules including those involved in pH i maintenance (Mulkey et al, 2004;Tsai et al, 1997). Consistently, it has been reported previously that GSH, the main low-molecularweight thiol in eukaryotes and Gram-negative bacteria, has the capability of maintaining a significantly higher pH i value under acid stress in Lactococcus lactis (Zhang et al, 2007).…”

“…In addition, data is accumulating to demonstrate that the exposure of microorganisms to various stresses, such as heavy metals, antibiotics, xenobiotics, heat and salt stress, can also increase the production of ROS and induce secondary oxidative stress (Kohanski et al, 2007;Mols and Abee, 2011a). Interestingly, previous studies have indicated that ROS can regulate pH i , at least in eukaryotic cells, via inhibiting proteins and biochemical pathways that affect pH i (Mulkey et al, 2004;Tsai et al, 1997). Our results demonstrate that acid stress does induce the generation of ROS in vivo in C. glutamicum, and that MSH-deficient mutants show a significantly higher ROS level than that of the wild-type due to losing the ability to synthesis MSH (Fig.…”

Mitochondrial translocation of α-synuclein is promoted by intracellular acidification

“…Mechanisms proposed for cytosolic acidification include: dysregulation of ion transport [42]; proton leakage from acidic organelles [43]; and hydrolysis of high energy nucleotides [44]. Notably, the PD toxin 1-methyl-4-phenylpyridinium (MPP + ) impairs cellular energy metabolism and leads to a decrease in pHi [45].…”

Hyperbaric oxygen and chemical oxidants stimulate CO2/H+-sensitive neurons in rat brain stem slices

“…Conversely, hyperoxia may increase neuronal responsiveness to hypercapnia. For example, previous studies found that hyperoxia caused intracellular acidosis in astrocytes, C6 glioma cells (67), and cardiac myoblasts (69) by reducing glycolysis and increasing ATP hydrolysis. Moreover, previous work in glomus cells found that hyperoxia can affect sensitivity to subsequent exposure to low O 2 (hypoxia) and hypercapnia (33).…”

“…38) and/or that O2 free radicals produced during HBO2 cause an intracellular acidification (b; Refs. 67,69). The result of either interaction would conceivably increase the neuronal response to the stimulus.…”