Peripheral p70S6k levels and emotional memory in patients with Alzheimer's disease.

Alzheimer's disease (AD) is clinically marked at the onset, by memory disturbances affecting explicit memory. Emotional explicit memory is enhanced in normal subjects and remained less affected at the beginning of AD. The kinase p70S6k participates in the control of protein translation and seems also implicated in the process of synaptic plasticity and the formation of memory at the molecular level. In a previous study, we have shown that peripheral p70S6k level is correlated with the decline of cognitive and memory functions in patients with AD. The goal of the present study was to analyse emotional and neutral explicit memory in AD patients and to evaluate the levels of active p70S6k in lymphocytes by western blots. The results reveal that the difference between emotional and neutral memories are correlated with the levels of peripheral p70S6k in patients with AD, as well as with the global cognitive scores assessed by the Mini Mental Status Examination. The decline of emotional memory in AD patients is reflected by the decrease of p70S6k levels.

Activated mTOR and PKR kinases in lymphocytes correlate with memory and cognitive decline in Alzheimer's disease.

BACKGROUND: The control of translation, involving the kinases mTOR (mammalian target of rapamycin) and PKR (double-stranded RNA-dependent protein kinase), modulates cell survival and death and is altered in the brains of patients with Alzheimer's disease (AD). In AD increased susceptibility of lymphocytes to apoptosis has been reported. METHODS: We investigated the level of the kinases mTOR and PKR and the eukaryotic initiation factor 2alpha (eIF2alpha) in lymphocytes of patients with AD in comparison with controls. In AD patients we also looked for a correlation between activated proteins and cognitive and memory tests. RESULTS: We report significant alterations of the levels of these kinases and eIF2alpha in lymphocytes of AD patients that were also significantly correlated with cognitive and memory test scores. CONCLUSION: These results suggest that the levels of mTOR, PKR and eIF2alpha in lymphocytes could follow the cognitive decline in AD.

Effect of ischemia on TBARS and lactate production in several cerebral regions of anaesthetised and awake rats.

The premise of neuroprotective therapy for acute ischemic stroke is based upon the possibility to interfere with the cellular ischemic cascade, so the understanding of the mechanisms and consequences of cerebral ischemia is necessary. The relationship between lipid peroxidation and acidosis was investigated in several regions of rat brain following ischemia without reperfusion. Male Wistar rats (280-300 g) were anaesthetised (Ketalar 33 mg/kg and Rompun 6.66 mg/kg) or not and underwent a four-vessel occlusion for 5 minutes. Then, thiobarbituric acid-reactive substances (TBARS) and lactate levels were measured in different brain regions (cerebellum, bulb, striatum, hippocampus, cortex). Induction of ischemia by ligation of two common carotid arteries and two vertebral arteries resulted in a production of TBARS (40-120%, p < 0.05) and lactate (20-60%, p < 0.05) in all cerebral regions of awake rats, especially in striatum, suggesting a potential link between lipid peroxidation and acidosis. When ischemia was realised on anaesthetised animals, an increase of lactate levels (30-50%, p < 0.05) was shown in all brain regions but TBARS were produced only in striatum (82%, p < 0.05). These data showed the particular vulnerability of striatum to ischemia and the possible opposite effects of an anaesthesia.

Evidence that acidosis alters the high-affinity dopamine uptake in rat striatal slices and synaptosomes by different mechanisms partially related to oxidative damage.

Several experimental studies have shown that acidosis impairs neurotransmitter uptake processes. The purpose of this study was to determine the mechanism underlying acidosis-induced alterations of the high-affinity dopamine (DA) uptake in rat striatal synaptosomes and slices. Acidosis (pH 5.5) performed either by lactic acid or phosphoric acid induced a decrease in the high-affinity DA uptake in the two striatal models, slices being lesser affected than synaptosomes. Addition of the acid prior to uptake measurement led to a strong reduction of the DA uptake velocity. This early inhibitory effect was completely reversed when acid was removed from the medium by washings. Conversely, when slices and synaptosomes were pre-incubated for different times with each acid, DA uptake remained inhibited in spite of washings. This later inhibition was accompanied by the production of thiobarbituric acid reactive substances, a marker of lipid peroxidation, and was partially prevented by the antioxidant Trolox. Taken together, these results suggest that acidosis, in a degree encountered during ischemia, alters the high-affinity DA uptake by at least two ways: an early and direct effect of H(+) ions on the DA transporters, and subsequently an inhibition partially mediated by free radical damage.

Sulfatide and GM1 ganglioside modulate the high-affinity dopamine uptake in rat striatal synaptosomes: evidence for the involvement of their ionic charges.

The present study was undertaken to examine the effects of the anionic glycolipids GM1 ganglioside and sulfatide on the high-affinity dopamine (DA) uptake in rat striatal synaptosomes. After 1h of incubation, GM1 stably bound to synaptosomes and modified the activity of the neuronal dopamine transporter (DAT). With 1.2 and 12 microM GM1, V(max) decreased by 13 and 23%, respectively, reflecting a slight reduction of the number of functional uptake sites and K(m) was lowered by 21 and 33%, thus showing an increase of the affinity. Treatment of synaptosomes with 1.2 microM of sulfatide, which possesses an anionic sulfated group, led to a similar decrease of V(max) (19%) than GM1, but to a significantly higher reduction of K(m) (35%). In fact, sulfatide associated to synaptosomes in a 3.5-fold higher extent than GM1. Conversely, when GM1 and sulfatide were replaced by GM1 alcohol and galactosylceramide, respectively, no modification of the DA uptake occurred, although these neutral glycolipids incorporated into the synaptosomes to the same extent as the related anionic compounds.Altogether, these results demonstrate the key role of negative charges linked to the oligosaccharide chains of glycolipids in the modulation of DA transport across the synaptosomal membrane.

Progressive alteration of neuronal dopamine transporter activity in a rat injured by an intranigral injection of MPP+.

MPTP or its metabolite MPP+ are used to produce a Parkinsonism syndrome in a variety of animal species. The present study describes the effects of intranigral MPP+ administration either at 10 or 40 microg on the neuronal dopamine transporter (DAT) activity measured in rat striatal synaptosomes at different times after lesion. The 40 microg MPP+ injection induced a maximal toxic effect on day 7. However, 10 microg MPP+ progressively inhibited DA uptake on the injured side. V(max) decreased in a time-dependent manner and the lowest value was observed on day 21 after lesion. At this time, the K(m) value began to increase and was continuously accentuated until day 45 as compared to the contralateral side. Treatments either with the antioxidant alpha-tocopherol acetate or the MAO inhibitor pargyline, given daily for 7 days after lesion, partially prevented the 40 microg MPP(+)-induced inhibition of DA uptake. Conversely, both treatments given daily for 21 days after lesion completely prevented the alteration of DAT activity in the ipsilateral striatum induced by 10 microg MPP+. The absence of protection when both treatments were stopped 2 weeks before DA uptake measurements indicated that free radicals and DA oxidized products were continuously accumulated and gradually affected the functionality of the DAT. These results demonstrate that a rat intranigral lesion with 10 microg MPP+ led to a progressive impairment of DAT activity.

Relevance of different striatal markers in assessment of the MPP+-induced dopaminergic nigrostriatal injury in rat.

Many striatal dopaminergic markers are available for estimating the degree of the nigrostriatal lesion by MPTP/MPP+, but the changes of these markers are not perfectly matched. In this study we investigated different striatal markers and determined which ones closely reflected the nigrostriatal alteration. The in vivo binding of (E)-N-(3-iodoprop-2-enyl)-2-beta-carbomethoxy-3beta-(4'-methylphenyl)nortropane (PE2I), a selective and potent inhibitor of the neuronal dopamine transporter (DAT) was considered as the reference index of injury of striatal dopaminergic nerve-endings. Rats received a 10-microg MPP+ injection in the right substantia nigra and were killed at 7 days after lesion. The results were as follows: (i) a decrease (66%) of the biodistribution of [125I]PE2I; (ii) a great reduction of the DAT expression measured by the binding of [125I]PE2I in striatal membranes (Bmax decreased by 54%) and in cerebral slices (88%); (iii) an 80% inhibition of the vesicular monoamine transporter expression revealed by the binding of [3H]dihydrotetrabenazine in cerebral slices; (iv) a robust decrease in the quantity of DA and its metabolites (about 50-60%); (v) a slight modification of the DAT activity with a decreased number of functional sites (Vmax decreased by 12%, p < 0.05) without change of the affinity in striatal synaptosomes. Among these markers the binding of [125I]PE2I in membrane homogenates and the content of DA, and its metabolites, in striatum could be the most relevant in vitro indexes of the degenerative state of the nigrostriatal pathway after MPP+ lesion.