Maria Noêmia Martins de Lima

Haloperidol- and clozapine-induced oxidative stress in the rat brain.

Haloperidol (HAL) is a typical neuroleptic that acts primarily as a D2 dopamine receptor antagonist. It has been proposed that reactive oxygen species play a causative role in neurotoxic effects induced by HAL. Adult male Wistar rats received daily injections of HAL (1.5 mg/kg) or clozapine (CLO, 25 mg/kg), an atypical neuroleptic, for 28 days. Control animals were given saline (SAL; NaCl 0.9%). Oxidative parameters in the brain were measured in the striatum (ST), hippocampus (HP) and cortex (CX). Thiobarbituric acid (TBA) reactive substances (TBAR) levels were increased by HAL treatment in the ST and decreased in CX of both of the HAL- and CLO-treated rats. Protein carbonyls were significantly increased by both HAL and CLO in the HP. The nonenzymatic antioxidant potential was decreased in the HP, and superoxide production was significantly increased in the ST following treatment with HAL. CLO induced an increase in superoxide production in the HP. Neither HAL nor CLO affected catalase (CAT) and superoxide dismutase (SOD) activities. The findings suggest that HAL and CLO can induce oxidative damage to the ST and HP in rats.

Recognition memory impairment and brain oxidative stress induced by postnatal iron administration.

Iron accumulation in the brain has been implicated in the pathogenesis of neurodegenerative disorders. It is known that iron catalyses the formation of highly reactive hydroxyl radicals. Recent studies have implicated oxidative damage in memory deficits in rats and humans. The purpose of the present study was to investigate the long‐term effects of iron treatment in four different phases of the neonatal period on recognition memory in rats. Additionally, parameters of oxidative stress in cerebral regions related to memory formation were evaluated. Male Wistar rats received vehicle or 10.0 mg/kg of Fe2+ orally at postnatal days 5–7, 12–14, 19–21 or 30–32. Animals given iron at any phase of the neonatal period showed impairments in long‐term retention of object recognition memory, although only the group given iron from postnatal days 12–14 showed a complete memory blockade. Iron treatment induced oxidative damage in the brain as assessed by the thiobarbituric acid reactive species assay. Moreover, iron administration increased superoxide production in submitochondrial particles, suggesting impaired mitochondrial function; and there was an increase in superoxide dismutase activity in brain regions susceptible to iron administration. The results show that iron load in the early stages of life induces cognitive impairment possibly by inducing oxidative damage in the brain. These findings are consistent with the view that oxidative stress may be related to the cognitive decline observed in normal ageing.

Selegiline protects against recognition memory impairment induced by neonatal iron treatment.

Excess of iron in the brain has been implicated in the pathogenesis of several human neurodegenerative diseases, for example Alzheimers disease and Parkinsons disease. It has been shown that the neonatal period is critical for the establishment of normal iron content in the adult brain. Moreover, it is known that aging alters the cerebral distribution of this metal. We have recently described that neonatal administration of iron severely impaired novel object recognition memory in rats. The aim of the present study was to determine whether selegiline, a monoamine oxidase (MAO) inhibitor known for its neuroprotective properties, could protect rats against cognitive impairment induced by neonatal administration of iron. In the first experiment, male Wistar rats received vehicle (5% sorbitol in water) or iron (10.0 mg/kg) orally from postnatal days 12 to 14 and saline (0.9% NaCl) or selegiline (1.0 or 10.0 mg/kg) intraperitoneally for 21 days, starting 24 h before the first iron dosing. In the second experiment, rats were given either vehicle or iron (10.0 mg/kg) orally from postnatal days 12 to 14 followed by saline or selegiline (1.0 or 10.0 mg/kg) intraperitoneally for 21 days, starting when rats reached adulthood (50th day after birth). Iron-treated rats given selegiline in both doses showed no deficits in recognition memory. Rats receiving iron but no selegiline presented memory deficits. This is the first study reporting the reversion of iron-induced memory impairment, supporting the view that our model can be considered as a useful tool in the search for new drugs with neuroprotective and/or memory enhancing properties.

Iron Leads to Memory Impairment that is Associated with a Decrease in Acetylcholinesterase Pathways

Increasing evidence indicates that excessive iron in selective regions of the brain may be involved in the etiology of neurodegenerative disorders. Accordingly, increased levels of iron have been described in brain regions of patients in Parkinsons and Alzheimers diseases. We have characterized neonatal iron loading in rodents as a novel experimental model that mimics the brain iron accumulation observed in patients with neurodegenerative diseases and produces severe cognitive impairment in the adulthood. In the present study we have investigated the involvement of the cholinergic system on iron-induced memory impairment. The effects of a single administration of the acetylcholinesterase (AChE) inhibitor galantamine or the muscarinic receptor agonist oxotremorine on iron-induced memory deficits in rats were examined. Male Wistar rats received vehicle or iron (10.0 mg/kg) orally at postnatal days 12 to 14. At the age of 2-3 months, animals were trained in a novel object recognition task. Iron-treated rats showed long-term impairments in recognition memory. The impairing effect was reversed by systemic administration of galantamine (1 mg/kg) immediately after training. In addition, iron-treated rats that received oxotremorine (0.5 mg/kg) showed enhanced memory retention. Rats given iron showed a decreased AChE activity in the striatum when compared to controls. The results suggest that, at least in part, iron-induced cognitive deficits are related to a dysfunction of cholinergic neural transmission in the brain. These findings might have implications for the development of novel therapeutic strategies aimed at ameliorating cognitive decline associated with neurodegenerative disorders.

Environmental and health risk assessments of genetically modified eucalypts in Brazil

Motivated by the Brazilian Ministry of Agriculture, Farming and Supply (MAPA), we created in 2009 the “Collaborating Center in Agriculture Defense Relative to the Biosafety of Genetically Modified Eucalypts” (Project “CDA Eucalyptus”) in order to collect information and conduct research to assess the biosafety of GM eucalypts in the Brazilian context. The Normative Resolution Nr. 5 of the National Biosafety Technical Commission (CTNBio) is the official document presenting all information needed to propose the commercial release of GMOs in Brazil. Based on this document and along with the personnel of the Suzano Paper & Cellulose Co., we conducted a series of experiments with GM and non-GM eucalypts planted in a test field in the state of Sao Paulo to start collecting the necessary information. Two independent groups of transgenic plants, harboring two different transgene constructions along with non-GM control plants are being assayed. The genetic traits, the identity or names of the transgenes as well as the identity of each tree individual will not be revealed due to intellectual property request still pending. Each group of plants was represented by four independent events in triplicates (2 groups x 4 events x 3 clonal trees + 3 non-GM clonal trees), therefore totaling 27 individuals under analysis. Samples were identified by random numbers and all assays were conducted in a simple-blind or a double-blind fashion. Tests concluded until now included (i) the detection of transgene regulatory sequences in purified DNA samples by conventional PCR and RT-qPCR, confirming the expected sampling conducted; (ii) extraction, chemical characterization and analysis of the antifungal effects of essential (volatile) oils extracted from leaves; (iii) pollen germination in vitro; (iv) flower morphology; (v) seed production; (vi) initial seedling development; (vii) leaf allelopathy; (viii) measurements of total phenolic compounds in leaves and roots; and (ix) effects of leaf extracts on the viability of human colon cells. All results obtained from experiments (ii) to (ix) revealed no statistical differences between GM- and non-GM-derived samples. A second round of experiments will be conducted to confirm these results. Proteomic and transcriptomic profiling of GM and non-GM trees are under analysis, as well as a series of experiments that include the chemical, nutritional and biological analysis of honey samples derived from bee hives located in fields of GM versus non-GM plants; and bee (Apis mellifera) population dynamics.

Neuropathological Changes in Adult and Old Rats Following Early Post-Natal Iron Administration

acetylcholine esterase (AChE) activity. Evidence for cholinergic dysfunction in early stages of Alzheimer’s disease (AD) or mild cognitive impairment (MCI) is inconclusive, and human studies of the relationship of cholinergic function with neuropsychological performance in vivo are rare. Methods: We included 21 healthy controls, 20 MCI and 15 mild AD patients. A comprehensive neuropsychological battery was administered to indicate the general cognitive function and to define performance of verbal and non-verbal memory, language, executive function and visuoconstructive abilities. [11C]-MP4A PET was applied in all subjects, parametric images of tracer hydrolysis rate constant k3 were generated and analysed using standard atlas regions. After principal component analysis (PCA) of k3 values correlational analysis with neuropsychological test results was performed. Results: The mean global k3 values of MCI (0.076 6 0.011 min-1) and AD patients (0.066 6 0.009 min-1) were significantly lower than that of controls (0.084 6 0.006 min-1) (p < 0.05 and p < 0.001, respectively) with the temporal regions showing the strongest decline in both patient groups. PCA revealed three main components representing frontal and parietal regions (PC1), temporal regions (PC2) and hippocampus and amygdala (PC3). PC 2 could separate all three diagnostic groups. Significant correlations in the whole group between PC2 and neuropsychological performances were found, most strongly with impaired verbal and visual memory function as well as impaired working memory, but also with deficient visuo-spatial and executive functions. The association of verbal and non-verbal memory with the ‘‘temporal’’ PC 2 was confirmed in MCI patients. This is in line with neuropathological studies showing the greatest and most consistent loss of cholinergic neurons in AD in the posterior part of the nucleus basalis Meynert (nbM-Ch4p), which primarily sends cholinergic projections to the temporal cortex (Geula and Mesulam, 1999). Conclusions: Cholinergic dysfunction in the temporal lobe is an early hallmark in MCI and mild AD and it is associated with impaired neuropsychological function.