Poly(ADP-ribose) polymerase: the nuclear target in signal transduction and its role in brain ischemia-reperfusion injury.

Poly(ADP-ribose) polymerase (PARP)-1 is a DNA nick sensor that transforms ADP-ribose from betaNAD+ in the form of polymer to over 40 nuclear proteins, particularly to histones, several transcription factors, and PARP itself, modulating their activities and functions. PARP-1 activated by DNA breaks facilitates transcription, replication, and DNA base excision repair. The last studies indicate that PARP-1 is the new nuclear target for fast signals evoked in cell membranes by depolarization and cholinergic and glutaminergic receptors stimulation. Excessive activation of PARP-1 by peroxynitrate-evoked DNA damage during oxidative stress can cause cell death by NAD+/ATP depletion after ischemia-reperfusion injury, inflammation, and diabetes mellitus. The PARP-1 through interaction with nuclear factor-kappaB, p53, and other transcription factors might significantly modulate cell survival and death and a type of death pathway. The pharmacological modulation of PARP-1 might offer a new effective approach for neuroprotection.

Age-related alteration of activity and gene expression of endothelial nitric oxide synthase in different parts of the brain in rats.

Nitric oxide (NO) plays important roles in aging and neurodegeneration. Our previous results indicated that aging differently affects NOS isoforms. Expression of nNOS mRNA was lower while iNOS was absent at any age. However, total NO synthesis increased in aged cerebral cortex and cerebellum as a consequence of changes of nNOS phosphorylation state. The question arise how aging influences activity and expression of eNOS in different parts of adult and aged brain. The levels of eNOS mRNA, protein and activity were measured using RT-PCR, immuno- and radiochemical methods, respectively. Our studies indicated that after inhibition of nNOS with 7-nitroindazole (7-NI) NO synthesis is lower in all parts of aged brain comparing to adults. However, eNOS activity significantly decreases only in cerebellum. The expression of eNOS determined on mRNA level was enhanced in all investigated aged brain parts to 140-190% of adult value and the data were statistically significant for cerebral cortex and cerebellum. The higher level of mRNA is probably the adaptive response to lower NOS activity. However, the Western-blot signal of eNOS protein was unchanged in aged brain parts comparing to adults suggesting age-related disturbances of protein synthesis and its function. It is also possible that a post-translational modification of the enzyme occurs in the aged rat brain. The lower eNOS activity in aged brain may significantly affects the signal transduction processes on the pathway NO/cGMP/PKG.

Transition metal ions significantly decrease phospholipase C activity degrading phosphatidylinositol-4,5-bisphosphate in the brain cortex.

Highly reactive transition metals, such as copper and iron play an obligatory role in generating of reactive oxygen species (ROS). Many neurodegenerative diseases including Alzheimer's disease (AD) and Parkinson's disease (PD) show increased accumulation of these metals. Phosphoinositide metabolism is altered in neurodegenerative diseases. In the present study, we examined the effect of CuSO(4) and FeCl(2) on phospholipase C (PLC) activity degrading phosphatidylinositol-4,5-bisphosphate (PIP(2)) and phosphatidylinositol (PI) in synaptic plasma membranes (SPM) from the rat brain cortex. We report that 25 microM CuSO(4) and FeCl(2) decreased PIP(2)-PLC activity by 60% and 75%, respectively. However, both compounds had no effect on PI-PLC activity. These data indicated that exclusively PIP(2)-PLC is sensitive to transition metal ions. We suggest that chelators of these metals may protect brain against alteration of phosphoinositide metabolism and might be beneficial in the treatment of neurodegenerative diseases.

Apolipoprotein E4 and A beta peptide 1-42 inhibit polyphosphoinositide biosynthesis in rat brain cortex.

Polyphosphoinositides synthesized by phosphatidylinositol 4-kinase, EC 2.7.1.67 (PI4K) and phosphatidylinositol 4-phosphate 5-kinase, EC 2.7.1.68 (PIP5K) are involved in cell signaling and cytoskeleton function. In this study, the effect of apolipoprotein E4 (apoE4) and amyloid beta peptide A beta 1-42 on PI4K and PIP5K activity in cortical synaptic plasma membranes from rat brain was investigated. The results indicated that 0.25 microM apoE4 inhibited PI4K and PIP5K by 40% and 30%, respectively, but 25 microM A beta 1-42 decreased exclusively PI4K activity by 20%. Although the mechanism of apoE4 and A beta action is unknown and needs future investigation, this study suggests that free radical-dependent protein oxidation may be involved in alteration of these enzymes.

Effects of aging and amyloid-beta peptides on choline acetyltransferase activity in rat brain.

Choline acetyltransferase (ChAT, acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6), involved in the learning and memory processes is responsible for the synthesis of acetylcholine. There are many discrepancies in literature concerning ChAT activity during brain aging and the role of amyloid beta peptides in modulation of this enzyme. The aim of the study was to investigate the mechanism of ChAT regulation and age-related alteration of ChAT activity in different parts of the brain. Moreover the effect of Abeta peptides on ChAT activity in adult and aged brain was investigated. The enzyme activity was determined in the brain cortex, hippocampus and striatum in adult (4-months-old), adult-aged (14-months-old) and aged (24-months-old) animals. The highest ChAT activity was observed in the striatum. We found that inhibitors of protein kinase C, A, G and phosphatase A2 have no effect on ChAT activity and that this enzyme is not dependent on calcium ions. About 70% of the total ChAT activity is present in the cytosol. Arachidonic acid significantly inhibited cytosolic form of this enzyme. In the brain cortex and striatum from aged brain ChAT activity is inhibited by 50% and 37%, respectively. The aggregated form of Abeta 25-35 decreased significantly ChAT activity only in the aged striatum and exerted inhibitory effect on this enzyme in adult, however, statistically insignificant. ChAT activity in the striatum was diminished after exposure to 1 mM H2O2. The results from our study indicate that aging processes play a major role in inhibition of ChAT activity and that this enzyme in striatum is selectively sensitive for amyloid beta peptides.