Nitric oxide consumption through lipid peroxidation in brain cell suspensions and homogenates

Abstract

Mechanisms which inactivate NO (nitric oxide) are probably important in governing the physiological and pathological effects of this ubiquitous signalling molecule. Cells isolated from the cerebellum, a brain region rich in the NO signalling pathway, consume NO avidly. This property was preserved in brain homogenates and required both particulate and supernatant fractions. A purified fraction of the particulate component was rich in phospholipids, and NO consumption was inhibited by procedures that inhibited lipid peroxidation, namely a transition metal chelator, the vitamin E analogue Trolox and ascorbate oxidase. The requirement for the supernatant was accounted for by its content of ascorbate which catalyses metal-dependent lipid peroxidation. The NO-degrading activity of the homogenate was mimicked by a representative mixture of brain lipids together with ascorbate and, under these conditions, the lipids underwent peroxidation. In a suspension of cerebellar cells, there was a continuous low level of lipid peroxidation, and consumption of NO by the cells was decreased by approx. 50% by lipid-peroxidation inhibitors. Lipid peroxidation was also abolished when NO was supplied at a continuously low rate (approximately 100 nM/min), which explains why NO consumption by this process is saturable. Part of the activity remaining after the inhibition of lipid peroxidation was accounted for by contaminating red blood cells, but there was also another component whose activity was greatly enhanced when the cells were maintained under air-equilibrated conditions. A similar NO-consuming process was present in cerebellar glial cells grown in tissue culture but not in blood platelets or leucocytes, suggesting a specialized mechanism.

Altered Flow-Mediated Vasodilatation, Low Paraoxonase-1 Activity, and Abnormal High-Density Lipoprotein Subclass Distribution in Takayasu's Arteritis

“…19,38 Our data demonstrated significant low plasma levels of PON1 activity in TA patients and low PON1 activity has also been observed in other physiological situations characterized by ED, such as HT, 39 chronic renal failure, 11,40 and peripheral artery disease, 37 as well as diabetes mellitus. 6,10,41 Considering all this evidence, we expected that PON1 would correlate with FMD, but it did not nor did it predict the FMD in the multiple regression analysis.…”

Inactivation of nitric oxide by rat cerebellar slices

“…In cerebellar cell suspensions, NO consumption is predominantly the result of ascorbate and iron together initiating the formation of lipid peroxyl radicals, which avidly react with NO (O'Donnell et al 1997;Keynes et al 2005a). To determine whether this process accounts for NO consumption in cerebellar slices, we tested the transition metal chelator, DTPA and the vitamin E analogue, Trolox (Britt et al 1992), both of which inhibit lipid peroxidation-dependent NO consumption in the cell suspensions (Keynes et al 2005a).…”

“…To determine whether this process accounts for NO consumption in cerebellar slices, we tested the transition metal chelator, DTPA and the vitamin E analogue, Trolox (Britt et al 1992), both of which inhibit lipid peroxidation-dependent NO consumption in the cell suspensions (Keynes et al 2005a). Slices were stimulated with both a sub-EC 50 concentration of NO (240 ± 30 nm NO, n = 10; generated from 10 μm Sper/NO), and a maximal concentration (100 μm DEA/NO).…”

“…The reaction of NO with superoxide (O 2 − ) is almost diffusion limited (Kissner et al 1997) but is limited physiologically by high concentrations of superoxide dismutase (SOD) which scavenges O 2 − (Beckman & Koppenol, 1996;Wink & Mitchell, 1998), such that this reaction is likely to be of greater significance in pathophysiological situations when O 2 − production is enhanced, for example during leakage of electrons from the respiratory chain during reperfusion following ischaemia, or from NADPH oxidase in activated microglia (Sankarapandi et al 1998). NO also reacts at an 550 C. N. Hall and J. Garthwaite J Physiol 577.2 almost diffusion-limited rate with lipid peroxyl radicals (O'Donnell et al 1997), which are generated at increased rates during pathological conditions (Moosmann & Behl, 2002) and which account for a component of NO consumption by acutely prepared brain cell suspensions and brain homogenates (Keynes et al 2005a). Enzymatic NO consumption by lipoxygenases and prostaglandin H synthase (PGHS) in reticulocytes and platelets (O'Donnell et al 1999(O'Donnell et al , 2000 and unidentified flavohaemoprotein(s) in mammalian cell lines (Gardner et al 2001;Hallstrom et al 2004), endothelial cells (Schmidt & Mayer, 2004) and cultured cerebellar glia (Keynes et al 2005b) have also been reported.…”

Original article Increased nitric oxide levels in cerebellum of cachectic rats with Walker 256 solid tumor

“…According to Keynes et al [13], the concentration of NO in the cerebellum is dependent on its generation and degradation rate. The consumption of NO by these cells decreases by 50% when lipid peroxidation inhibitors are used.…”