The protective role of dl-α-lipoic acid in biogenic amines catabolism triggered by Aβ amyloid vaccination in mice
Introduction
Alzheimer's disease (AD) is the most common neurodegenarative disease that involves aggregation and deposition of misfolded proteins such as β-amyloid (Aβ) peptide [1], [39] leading to activation of glia (gliosis) prior to the deposition of amyloid plaques and formation of neurofibrillary tangles in degenerated neurons that ascend from the basal forebrain to cortical and hippocampal areas [19]. The search for AD pathogenic factors is hindered by the lack of diagnostic certainty in life and by the heterogeneity of the disease [40]. Recent reports suggest that perturbations in systems using the excitatory amino acid l-glutamate (l-Glu) may underlie such pathogenic mechanisms of AD [22], [23].
Along with l-Glu, biogenic amines viz., 5-hydroxytryptamine (5-HT), dopamine (DA), and norepinephrine (NE) have been identified as the principal transmitters mediating fast excitatory synaptic responses in the vertebrate central nervous system (CNS) and have widespread influence on the cortex, basal ganglia, limbic areas and spinal cord. They play an important role in regulation of brain functions: memory, mood, movement, behavior and autonomic functions such as blood pressure regulation. These functions are impaired in Alzheimer's disease (AD) due to loss of the catecholamines (CA) neurons, which regulate the brain functions [45]. The major and well-documented concept in neurodegenerative disorder is the deficiency of the cholinergic system due to the progressive loss of cholinergic presynaptic neurons situated in the basal fore brain that leads to structural and functional damage of monoaminergic systems [7], [8]. The neurodegenerative disease condition can be induced by different insults that have been related to systemic oxidative stress such as inflammatory stimulus [47].
Lemere et al. [26] demonstrated that intranasal immunization with freshly solubilized Aβ 1–40 reduced cerebral amyloid burden in PDAPP mouse (PDAPP mouse is a model of familial Alzheimer's disease in which mice express the human V717F mutation in the amyloid precursor protein (APP) gene under the control of the platelet derived (PD) growth factor-beta promoter). The Antibodies formed against Aβ might stimulate Aβ clearance by stimulating Fc-receptor-mediated phagocytosis of Aβ by microglia [4]. In addition antibodies against Aβ might act as an artificial chaperone for extra cellular amyloid-β and prevent aggregation of amyloid β by binding to epitopes [13]. However, phase II clinical trial with the immunization of AD patients with amyloid peptides has recently ended prematurely because of cerebral hemorrhage due to inflammation observed in several patients [11]. Neocortex and hippocampus are susceptible to neurodegeneration in response to an external insult such as infection and/or oxidative stress that leads to disruption of the neuronal circuitry of the hippocampus and neocortex cerebral areas [12], [20].
In the past decade an extraordinary progress has been made to understand the toxicity of Aβ peptide. Presently, compounds targeting Aβ vaccination hold much promise as therapy for AD. Therapies that decrease Aβ production, enhance its degradation or cause its clearance from CNS are the two major therapeutic ventures being pursued by various group of researchers. One of the most promising approaches towards the Aβ clearance is the use of immunotherapies, including vaccination with Aβ fragments [46]. However, inflammation is the limiting factor in therapy due to the sudden increase in the soluble Aβ peptide in the peripheral and brain tissues on vaccination.
dl-α-Lipoic acid (LA) and its reduced form, dihydrolipoic acid (DHLA) are natural potent antioxidants and cofactors of mitochondrial decarboxylation enzymatic reactions and essential for an adequate ATP production through citric acid cycle [36]. They are involved in different multi enzyme complexes such as pyruvate dehydrogenase, α-ketoglutarate dehydrogenase, branched chain α-ketoacid dehydrogenase and glycine decarboxylase complex [37]. LA, a low molecular substance which is absorbed and able to cross blood brain barrier [32] in animals, and preliminary human studies indicate that LA has neuroprotective effects on Aβ mediated cytotoxicity and may be effective in a number of neurodegenerative diseases [33].
The antioxidant therapy along with Aβ vaccination reduces the inflammation associated with Aβ vaccination. Our preliminary biochemical and histological investigation shows that LA therapy reduces the oxidative vulnerability of Aβ vaccination and that it is a good candidate in inflammatory AD models where active immunization is administered [35]. This study was focused on the quantitative analysis of central nervous system neurotransmitters (DA, NE and 5-HT) and its metabolites (HVA and 5-HIAA) in neocortex and hippocampus by the aid of isocratic RP–HPLC system with electrochemical detection. We have used one of the important antigenic fragment of Aβ25–35 as peptide vaccination candidate which exhibit all the biological activity of the full length Aβ [16], [43] and is used by other researchers to study the neuronal toxicity and oxidative vulnerability extensively [18].
Section snippets
Materials and methods
Six- to eight-week-old Swiss strain male mice weighing 25–30 g, were obtained from Tamilnadu Veterinary and Animal Sciences University, Madhavaram, Chennai, Tamilnadu, India. The animals were maintained in the Poly acrylic cages under hygienic conditions at ambient room temperature (28–30 °C) on a 12 h light/dark cycle. The animals had free access to water. They were fed with commercial pellet diet (Hindustan Lever Ltd., Bangalore, India) ad libitum. All experimental conditions were in accordance
Statistical analysis
Data are expressed as mean ± S.E.M. for six animals in each group. Statistical analysis of variance (ANOVA) followed by the Tukey's test was applied to determine the significant differences among the groups. p values less than 0.05 were considered significant.
Results
HPLC–EC technique was employed to measure biogenic amines and their metabolites of control and experimental mice. The endogenous content of monoamines (5-HT) and catecholamines (DA, NE) was found to be heterogeneously distributed in the neocortex and hippocampus of saline treated control animals (Table 1, Table 2, Table 3). Among the biogenic amines examined in the neocortex area of control mice, the level of DA was found to be at highest concentration followed by 5-HT and NE. Although similar
The efficacy of LA on the biogenic amines in systemic inflammation induced mice
Recently, we demonstrated that SI induced by silver nitrate leads to accumulation of reactive oxygen species (ROS) and lipid peroxidation (LPO) in the lymphocytes, hepatocytes, astrocytes of mice [17]. In the current study, the efficacy of LA therapy in SI induced mice was assessed by the level of biogenic amines. The neurochemical responses in the neocortex and hippocampus during SI are depicted in Table 1. Result shows a significant (p < 0.05) depletion in the level of 5-HT, DA, and NE in the
The efficacy of LA on biogenic amines in Aβ immunized mice
Fu et al. reported that catecholamines exacerbate oxidative stress induced by Aβ with mitochondrial dysfunction and perturbed calcium homeostasis [14]. In the present study, we monitored the protective effects of LA on biogenic amines catabolism induced by Aβ. Result shows that Aβ significantly (p < 0.05) impair the levels of DA, NE and 5-HT with an increase in their metabolites in the neocortex and hippocampus on 1st day as shown in Table 2. However, LA therapy restored Aβ depleted biogenic
The efficacy of LA on biogenic amines in the mice after systemic inflammation and Aβ immunization
In an earlier work, we reported that Aβ immunization with inflammation induces accumulation of ROS and LPO in lymphocytes, hepatocytes, astrocytes of mice [35]. In the present study, Aβ immunization in inflammation induced mice was used to evaluate the role of LA through the changes in aminergic system. The level of biogenic amines was significantly (p < 0.05) low in the neocortex and hippocampus of IV and IVL groups mice when compared with control on 1st day (Table 3). Regarding the level of
Discussion
Systemic inflammation activates inflammatory cells to produce a variety of proinflammatory and cytotoxic factors, including the cytokines, tumor necrosis factor-α, free radicals like nitric oxide (NO) and ROS. These proinflammatory cytokines are transported to various tissues through the blood stream [6], including the brain parenchyma and circumventricular sites, which lack a functional blood brain barrier [21]. These factors lead to loss of the dopaminergic neuronal function [28]. This may be
Acknowledgements
We are grateful to Dr. T. Ramasami, Director, Central Leather Research Institute, Chennai, for his kind permission to publish this work. The authors J.G.M., and E.P.J. thank the Council of Scientific and Industrial Research (CSIR, New Delhi), India for awarding fellowship. We thank Dr. S. Kathiroli, Director, National Institute of Ocean Technology, Chennai, for extending the HPLC–EC facility required for this work. Finally, we thank the reviewers for their valuable comments and suggestion.
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