Adina Michael-Titus

Neurological Benefits of Omega-3 Fatty Acids

The central nervous system is highly enriched in long-chain polyunsaturated fatty acid (PUFA) of the omega-6 and omega-3 series. The presence of these fatty acids as structural components of neuronal membranes influences cellular function both directly, through effects on membrane properties, and also by acting as a precursor pool for lipid-derived messengers. An adequate intake of omega-3 PUFA is essential for optimal visual function and neural development. Furthermore, there is increasing evidence that increased intake of the long-chain omega-3 PUFA, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), may confer benefits in a variety of psychiatric and neurological disorders, and in particular neurodegenerative conditions. However, the mechanisms underlying these beneficial effects are still poorly understood. Recent evidence also indicates that in addition to the positive effects seen in chronic neurodegenerative conditions, omega-3 PUFA may also have significant neuroprotective potential in acute neurological injury. Thus, these compounds offer an intriguing prospect as potentially new therapeutic approaches in both chronic and acute conditions. The purpose of this article is to review the current evidence of the neurological benefits of omega-3 PUFA, looking specifically at neurodegenerative conditions and acute neurological injury.

Omega-3 Fatty Acids Improve Recovery, whereas Omega-6 Fatty Acids Worsen Outcome, after Spinal Cord Injury in the Adult Rat

Spinal cord injury (SCI) is a cause of major neurological disability, and no satisfactory treatment is currently available. Evidence suggests that polyunsaturated fatty acids (PUFAs) could target some of the pathological mechanisms that underlie damage after SCI. We examined the effects of treatment with PUFAs after lateral spinal cord hemisection in the rat. The ω-3 PUFAs α-linolenic acid and docosahexaenoic acid (DHA) injected 30 min after injury induced significantly improved locomotor performance and neuroprotection, including decreased lesion size and apoptosis and increased neuronal and oligodendrocyte survival. Evidence showing a decrease in RNA/DNA oxidation suggests that the neuroprotective effect of ω-3 PUFAs involved a significant antioxidant function. In contrast, animals treated with arachidonic acid, an ω-6 PUFA, had a significantly worse outcome than controls. We confirmed the neuroprotective effect of ω-3 PUFAs by examining the effects of DHA treatment after spinal cord compression injury. Results indicated that DHA administered 30 min after spinal cord compression not only greatly increased survival of neurons but also resulted in significantly better locomotor performance for up to 6 weeks after injury. This report shows a striking difference in efficacy between the effects of treatment with ω-3 and ω-6 PUFAs on the outcome of SCI, with ω-3 PUFAs being neuroprotective and ω-6 PUFAs having a damaging effect. Given the proven clinical safety of ω-3 PUFAs, our observations show that these PUFAs have significant therapeutic potential in SCI. In contrast, the use of preparations enriched in ω-6 PUFAs after injury could worsen outcome after SCI.

Disruption of Neurogenesis on Gestational Day 17 in the Rat Causes Behavioral Changes Relevant to Positive and Negative Schizophrenia Symptoms and Alters Amphetamine-Induced Dopamine Release in Nucleus Accumbens

Gestational disruption of neurodevelopment has been proposed to lead to pathophysiological changes similar to those underlying schizophrenia. We induced such disruption by treating pregnant rat dams with methylazoxymethanol acetate (MAM) on gestational day 17 (GD17). Total brain size and that of the prefrontal cortex and hippocampus were reduced in adult rats exposed prenatally to MAM. When locomotor activity was assessed in an open field, MAM-exposed rats were hyper-responsive to a mild stress and to amphetamine (2 mg/kg, s.c.). They also engaged in less social interaction than controls. We studied, by microdialysis, the effect of amphetamine on extracellular dopamine in the nucleus accumbens and the medial prefrontal cortex of freely moving control and MAM-exposed rats. Amphetamine (2 mg/kg, s.c.) induced an increase in dopamine release that was larger in the nucleus accumbens of MAM-exposed rats than in controls, whereas no difference was seen in the medial prefrontal cortex. In controls, amphetamine infused into the medial prefrontal cortex (50 μM) led to a slight decrease in extracellular dopamine in the nucleus accumbens. This effect was absent in MAM-exposed rats, where a transient increase in nucleus accumbens dopamine levels was seen after amphetamine infusion. These results show that the late gestational disruption of neurogenesis in the rat leads to behavioral changes that mimic positive and negative schizophrenia symptoms, and also to a dysregulation of subcortical dopamine neurotransmission. This study contributes to the evaluation of the validity of the prenatal MAM GD17 treatment in rats as an animal model for schizophrenia.

Omega-3 fatty acids reverse age-related decreases in nuclear receptors and increase neurogenesis in old rats.

Retinoic acid receptors (RARs), retinoid X receptors (RXRs), and peroxisome proliferator‐activated receptors (PPARs) are transcription factors involved in many cellular processes, such as learning and memory. RAR and RXR mRNA levels decrease with ageing, and the decreases can be reversed by retinoic acid treatment, which also alleviates age‐related memory deficits. The omega‐3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have neuroprotective effects in the aged brain and are endogenous ligands of RXR and PPAR. We investigated whether dietary EPA and DHA supplementation reverses age‐related declines in protein levels of these receptors in rat forebrain. Two studies were conducted comparing adult and old rats. In the first, old rats were fed standard or EPA/DHA‐enriched (270 mg/kg/day, EPA to DHA ratio 1.5:1) diets for 12 weeks. Analysis by Western blot revealed significant decreases in RARα, RXRα, RXRβ, and PPARγ in the forebrain with ageing, which were reversed by supplementation. Immunohistochemical analysis of the hippocampus showed significant age‐related decreases in RARα and RXRβ expression in CA1 and the dentate gyrus, which were restored by supplementation. Decreases in hippocampal doublecortin expression were also partially alleviated, suggesting a positive effect on neurogenesis. We also investigated the effects of DHA supplementation (300 mg/kg/day for 12 weeks) on RARα, RXRα, and RXRβ expression in the prefrontal cortex, striatum, and hippocampus. Overall, DHA supplementation appeared to increase receptor expression compared with the untreated old group. These observations illustrate additional mechanisms that might underlie the neuroprotective effects of omega‐3 fatty acids in ageing.

Decrease in parvalbumin-expressing neurons in the hippocampus and increased phencyclidine-induced locomotor activity in the rat methylazoxymethanol (MAM) model of schizophrenia

Treatment of rats with methylazoxymethanol (MAM) on gestational day (GD)17 disrupts corticolimbic development in the offspring (MAM‐GD17 rats) and leads to abnormalities in adult MAM‐GD17 rats resembling those described in schizophrenic patients. The underlying changes in specific cortical and limbic cell populations remain to be characterised. In schizophrenia, decreases in inhibitory γ‐aminobutyric acid (GABA)‐containing interneurons that express the calcium‐binding protein parvalbumin have been reported in the prefrontal cortex and hippocampus. In this study we analysed the expression of parvalbumin (PV), calretinin (CR) and calbindin (CB) in the prefrontal cortex and hippocampus of MAM‐GD17 rats. Exposure in utero to MAM led to a significant decrease in the number of neurons expressing PV in the hippocampus, but not the prefrontal cortex. Neurons expressing CR or CB were not affected in either structure. The neurochemical changes in MAM‐GD17 rats were accompagnied by increased hyperlocomotion after administration of phencyclidine (PCP), analogous to the hypersensitivity of schizophrenic patients to PCP. Therefore, the developmental MAM‐GD17 model reproduces key neurochemical and behavioural features that reflect cortical and subcortical dysfunction in schizophrenia, and could be a useful tool in the development of new antipsychotic drugs.

Imipramine and phenelzine decrease glutamate overflow in the prefrontal cortex—a possible mechanism of neuroprotection in major depression?

Antidepressant drugs have been used for decades, but the neurobiological substrate of their efficacy is not completely understood. Although these drugs have well-established effects on monoamines, evidence is emerging that they may also affect other neurotransmitter systems. It has been shown that treatment with a wide range of antidepressants changes the binding characteristics of the N-methyl-D-aspartate type of glutamate receptor. This change is delayed and occurs only in the cortex. The mechanism that triggers it is unknown. We hypothesized that N-methyl-D-aspartate receptor alterations may be due to changes in the dynamics of cortical excitatory amino acid release. Such changes are of particular interest in areas such as the prefrontal cortex, a region involved in stress responses and affected in major depression. We investigated the effects of two antidepressants with different modes of action, imipramine and phenelzine, on glutamate and aspartate outflow in rat prefrontal cortex and striatum. We showed that antidepressants significantly decreased stimulated glutamate outflow. The effect had a rapid onset, was sustained during chronic administration and was only seen in the prefrontal cortex. This change may initiate receptor alterations. Furthermore, if antidepressants can dampen states of hyperglutamatergic activity and the subsequent excitotoxicity, their chronic use may have a considerable neuroprotective potential in major depression.

Dietary enrichment with omega-3 polyunsaturated fatty acids reverses age-related decreases in the GluR2 and NR2B glutamate receptor subunits in rat forebrain

Ageing is associated with a decrease in the brain content of omega-3 polyunsaturated fatty acids (PUFA), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), and with decreased neuroplasticity. The glutamate receptor subunits GluR2 and NR2B play a significant role in forebrain synaptic plasticity. We investigated GluR2 and NR2B in the aged prefrontal cortex, hippocampus and striatum, and tested if treatment with a preparation containing EPA and DHA can reverse age-related changes. The study compared adult and old (3-4 and 24-26 month) rats, and the latter were fed a standard diet or a diet supplemented for 12 weeks with omega-3 PUFA at 270mg/kg/day (ratio EPA to DHA 1.5:1). Ageing was associated with decreases in the GluR2 and NR2B subunits in all structures. These decreases were fully reversed by omega-3 PUFA supplementation. Age-related changes in the phospholipid PUFA content were also seen. Decreases in DHA were mostly corrected by supplementation. This study supports the neuroprotective effect of omega-3 fatty acids in brain ageing, and illustrates specific mechanisms underlying this effect.

Omega-3 polyunsaturated fatty acids increase the neurite outgrowth of rat sensory neurones throughout development and in aged animals

Polyunsaturated fatty acids (PUFA) of the omega-3 series and omega-6 series modulate neurite outgrowth in immature neurones. However, it has not been determined if their neurotrophic effects persist in adult and aged tissue. We prepared cultures of primary sensory neurones from male and female rat dorsal root ganglia (DRG), isolated at different ages: post-natal day 3 (P3) and day 9 (P9), adult (2-4 months) and aged (18-20 months). Cultures were incubated with the omega-6 PUFA arachidonic acid (AA) and the omega-3 PUFA eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), at 0.8, 4, 8 and 40muM. PUFA increased neurite outgrowth throughout the developmental stages studied. The effects of omega-3 PUFA, in particular DHA, were still prominent in aged tissue. The amplitude of the effects was comparable to that of nerve growth factor (NGF; 50ng/ml) and all-trans-retinoic acid (ATRA; 0.1muM). The effects of PUFA were similar in cells positive or negative for the N52 neurofilament marker. Our results show that omega-3 PUFA have a marked neurite-promoting potential in neurones from adult and aged animals.

Erythropoietin and carbamylated erythropoietin are neuroprotective following spinal cord hemisection in the rat

The cytokine erythropoietin (EPO) has been shown to be neuroprotective in a variety of models of central and peripheral nervous system injury. Derivatives of EPO that lack its erythropoietic effects have recently been developed, and the initial reports suggest that they have a neuroprotective potential comparable to that of EPO. One such derivative is carbamylated EPO (CEPO). In the current study we compared the effects of treatment with EPO and CEPO on some of the early neurodegenerative events that occur following spinal cord injury (SCI) induced by hemisection. Adult male Wistar rats received a unilateral hemisection of the spinal cord. Thirty minutes and 24 h following injury, animals received an intraperitoneal injection of saline, EPO (40 µg/kg) or CEPO (40 µg/kg). Results indicated that 3 days post‐injury, both CEPO and EPO decreased to a similar extent the size of the lesion compared with control animals. Both compounds also decreased the number of terminal transferase‐mediated dUTP nick‐end labelling (TUNEL)‐labelled apopotic nuclei around the lesion site, as well as the number of axons expressing the injury marker β‐amyloid precursor protein. EPO and CEPO also increased Schwann cell infiltration into the lesion site, although neither compound had any effect on macrophage infiltration either within the lesion site itself or in the surrounding intact tissue. In addition, immunohistochemistry showed an increased expression of both the EPO receptor and the β common receptor subunit, the components of the receptor complex proposed to mediate the neuroprotective effects of EPO and CEPO in neurons near the site of the injury. The results show that not only does CEPO have an efficacy comparable to that of EPO in its neuroprotective potential following injury, but also that changes in the receptors for these compounds following SCI may underlie their neuroprotective efficacy.

DISTRIBUTION OF TEMOPORFIN, A NEW PHOTOSENSITIZER FOR THE PHOTODYNAMIC THERAPY OF CANCER, IN A MURINE TUMOR MODEL

Abstract— The biodistribution of temoporfin (tetra[m‐hydroxyphenyl]chlorin, m‐THPC), a recently developed photosensitizer, was investigated in BALB/c mice. The drug was administered intravenously (0.35‐0.75 μmol/kg) to tumor‐free mice or to mice implanted with the Colo 26 colorectal carcinoma. Blood and tissue samples were collected for up to 96 h post‐dose. Drug concentrations were determined by HPLC coupled to photometric detection at 423 nm. Concentrations in blood and liver fell relatively rapidly such that blood concentrations at later time points were below the limit of detection. Tumor concentrations rose at first and then remained constant from 24 h. Temoporfin concentrations in some tissues, notably heart and skeletal muscle, declined only slowly when compared to blood. The tumor: tissue ratios for those organs that showed a more rapid decline in temoporfin concentrations were higher at later times, whereas in tissues such as muscle the ratio remained relatively constant. The organs with the highest tumor: tissue ratios were small intestine (8.6), liver (6.9) and skeletal muscle (5.0).