Arachidonic acid preserves hippocampal neuron membrane fluidity in senescent rats
Introduction
Polyunsaturated fatty acids (PUFA) such as arachidonic acid (AA: n-6 PUFA) and docosahexaenoic acid (DHA: n-3 PUFA) have an important role in the induction and maintenance of long-term potentiation (LTP) [6], [9]. Recent behavioral studies suggest a positive correlation between PUFA and cognition in mammals [1], [11], [15], [29], [34]. Previously, we reported that long-term dietary intake (>3 months) of AA in senescent rats older than 21 months improves Morris water–maze performance and the induction and maintenance of hippocampal LTP [19]. Neuronal membranes in the senescent rat hippocampal region have increased microviscosity together with the loss of some receptors, such as muscarinic 1 and NMDA receptors [30], [31], [36]. These age-dependent hippocampal deficits might be restored with long-term dietary intake of AA or l-α-glycerylphosphorylcholine, which protect against membrane oxidation and might facilitate the uptake of cholesterol into the membrane [24], [30].
Experimental studies of changes in hippocampal neural plasticity during the course of aging indicate that there is a shift in the mechanisms that regulate the thresholds for synaptic modification, including Ca2+ channel function and subsequent Ca2+-dependent processes, providing the basis for a model of age-related changes in hippocampal synaptic function [8]. Alterations in the neural membrane phospholipid components not only influence crucial intracellular and intercellular signaling, but also alter many physical properties of the membrane, such as fluidity, phase transition temperature, bilayer thickness and lateral domains [14]. A PUFA deficiency markedly affects neurotransmission, membrane-bound enzyme and ion channel activities and synaptic plasticity [45]. PUFA supplementation in the diet restores membrane fluidity and gene expression, decreases inflammation and modulates neurotransmission [2], [4], [20], [24], [25], [39], [41], [42]. For example, DHA supplementation increases the membrane fluidity of cardiomyocytes [20] and kidney cells [12] and ameliorates the age-related increases in lipid peroxidation and deterioration in fluidity in canalicular plasma membranes [13]. Although n-3 PUFA (e.g., eicosapentaenoic acid and DHA) and n-6 PUFA (e.g., linoleic acid and AA) have contrasting effects on inflammation [2], [4], gene expression [39], [41], [42], proliferation processes [3], platelet membrane fluidity [26], apoptosis [33] and cytoprotective and cytotherapeutic activity [35], PUFA supplementation improves signal transduction processes in association with improvements in learning and memory, peroxisomal disorders and psychotic changes in schizophrenia, depression, hyperactivity, stroke and Alzheimer disease [14], [40].
Several studies of membrane fluidity have assessed membrane microviscosity [17], [30], [46]. In these studies, a membrane suspension obtained from brain tissue homogenate is measured using fluorescence polarization. This method provides information only on the total viscosity of the membrane. The use of green fluorescent protein allows for studies of subcellular localization, mobility, transport routes and binding interactions of proteins in living cells. Live cell imaging, in combination with photobleaching, energy transfer or fluorescent correlation spectroscopy provides many insights into the movement of proteins and their interactions with cellular components. The mobility of a fluorescent protein can be assessed using a specific type of photobleaching technique called fluorescent recovery after photobleaching (FRAP). In this technique, fluorescent molecules in a small region of the cell are irreversibly photobleached using a high-power laser beam and subsequent movement of the surrounding non-bleached fluorescent molecules into the photobleached area is recorded with lower laser power. Two typical kinetic parameters of a protein can be discerned from quantitative studies using FRAP: the mobile fraction, Mf, which is the fraction of fluorescent proteins that can diffuse into the bleached region during the time course of the experiment and the diffusion constant, D, which is a measure of the rate of protein movement in the absence of low or active transport [10], [16], [22].
Kessler and Yehuda reported a marked decrease in learning-induced cholesterol levels in the hippocampus and a learning-induced increase in membrane lipid fluidity. They postulated that upon learning, brain membranes undergo changes in the lipid membrane phase that facilitate the transduction of the learning process into a biochemical template [17]. The number of muscarinic receptors in the hippocampus of aged rats is significantly lower than that in young animals [30]. Treatment of aged rats with S-adenosyl-l-methionine, however, restores the number of muscarinic receptors [31]. Though the dissociation constants are not different among young and S-adenosyl-l-methionine-treated and-untreated aged rats, binding capacity varies. The reduction in the muscarinic receptor density might be related to the aging-induced decrease in neuronal membrane fluidity [31]. Scheuer et al. demonstrated that piracetam alters membrane fluidity in the hippocampus and that the effects of piracetam on NMDA density in the hippocampus might be involved in its positive effects on cognitive performance [36]. Chronic treatment of aged rats with l-α-glycerylphosphorylcholine restores the number of muscarinic 1 receptors and partially restores membrane fluidity [30].
Cotrina suggested that astrocytic gap junction coupling in mouse hippocampus tends to be reduced as animals age. Using the FRAP technique, fluorescence recovery after 2 min was 63 ± 6% in younger animals, 59 ± 5% in adult animals and 54 ± 4% in old brain. Their observations indicate that although astrocytic gap junction proteins are maintained at high levels throughout the lifespan of the animal, aging is associated with changes in the number and size of both Cx30 and Cx43 gap junction plaques [5].
In the present study, we used FRAP to evaluate and compare membrane fluidity of hippocampal pyramidal neurons among young, aged control and aged animals given long-term dietary supplementation with AA.
Section snippets
Animals
All experiments were performed with Fischer 344 male rats 2 months of age (young control, YC) or 18 months of age (aged animals) obtained from Clea Japan, Tokyo. Aged rats were subdivided into two groups; one group (old animals, OA) was fed experimental chow supplemented with AA containing triglyceride and the other (old control, OC) was fed control chow for 12 weeks. The AA chow contained 2 g AA [SUNTGA40-S (ARAVITA 40), SUNTORY, Osaka, Japan] per kilogram powder chow. The control diet
Results
In the present study, the number of animals used was 3 YC, 2 OC and 2 OA. Five or six hippocampal slices were obtained from each rat. The degree of staining with DiI C16(3) at the soma of the CA1 pyramidal neurons among the three groups was bright enough to detect the recovery of the fluorescent intensity based on microscopic observation. Because we did not measure the absolute intensity level of the photobleached area of each group, it was difficult to compare the staining level among YC, OC
Discussion
We used FRAP to demonstrate that there was a significant difference in membrane fluidity in hippocampal neurons between YC and OC; hippocampal pyramidal neurons from young animals were more fluid, i.e., had more lateral mobility, than those of senescent animals, as characterized by a larger diffusion constant and mobile fraction and a smaller time constant. Long-term (>3 months) AA supplementation kept the hippocampal neuronal membranes more fluid, but the amount of protein molecules available
Acknowledgements
We thank Mr. Shingo Maeda of Olympus Co. for technical assistance. This work was supported by grants from Tokai University for the Promotion of Scientific Research and Suntory Ltd. to M.S.
References (48)
- et al.
Different effects of n−6 and n−3 polyunsaturated fatty acids on the activation of rat smooth muscle cells by interleukin-1 beta
J Lipid Res
(2003) - et al.
Expression and function of astrocytic gap junctions in aging
Brain Res
(2001) Involvement of hippocampal synaptic plasticity in age-related memory decline
Brain Res Brain Res Rev
(1999)- et al.
Relationship between age-related increases in rat liver lipid peroxidation and bile canalicular plasma membrane fluidity
Exp Gerontol
(2001) - et al.
Docosahexaenoic acid in the diet: its importance in maintenance and restoration of neural membrane function
Prostaglandins Leukot Essent Fatty Acids
(2004) - et al.
Health benefits of docosahexaenoic acid (DHA)
Pharmacol Res
(1999) - et al.
Synaptic plasticity preserved with arachidonic acid diet in aged rats
Neurosci Res
(2003) - et al.
Membrane fluidity changes are associated with the antiarrhythmic effects of docosahexaenoic acid in adult rat cardiomyocytes
J Nutr Biochem
(2000) - et al.
Activation of synaptic NMDA receptors induces membrane insertion of new AMPA receptors and LTP in cultured hippocampal neurons
Neuron
(2001) Age-related impairment in long-term potentiation in hippocampus: a role for the cytokine, interleukin-1 beta?
Prog Neurobiol
(1998)
Effects of dietary fish oil supplementation on platelet aggregability and platelet membrane fluidity in normolipemic subjects with and without high plasma Lp(a) concentrations
Atherosclerosis
Long-term potentiation in aged rats is restored when the age-related decrease in polyunsaturated fatty acid concentration is reversed
Prostaglandins Leukot Essent Fatty Acids
Effect of l-alpha glycerylphosphorylcholine on muscarinic receptors and membrane microviscosity of aged rat brain
Prog Neuropsychopharmacol Biol Psychiatry
Effect of S-adenosyl-l-methionine on brain muscarinic receptors of aged rats
Eur J Pharmacol
Dietary supplementation with vitamin E reverses the age-related deficit in long term potentiation in dentate gyrus
J Biol Chem
Arachidonic acid improves aged rats’ spatial cognition
Physiol Behav
The role of omega-3 long-chain polyunsaturated fatty acids in health and disease of the retina
Prog Retin Eye Res
Regulation of selenoprotein GPx4 expression and activity in human endothelial cells by fatty acids, cytokines and antioxidants
Atherosclerosis
Docosahexaenoic acid: membrane properties of a unique fatty acid
Chem Phys Lipids
Polyunsaturated fatty acids (PUFA) and eicosanoids in human health and pathologies
Biomed Pharmacother
The role of polyunsaturated fatty acids in restoring the aging neuronal membrane
Neurobiol Aging
Modification by docosahexaenoic acid of age-induced alterations in gene expression and molecular composition of rat brain phospholipids
Proc Natl Acad Sci USA
Modulation of cyclin D1 and early growth response factor-1 gene expression in interleukin-1beta-treated rat smooth muscle cells by n−6 and n−3 polyunsaturated fatty acids
Eur J Biochem
Dietary modification of inflammation with lipids
Proc Nutr Soc
Cited by (94)
The association between long-chain polyunsaturated fatty acid intake and changes in brain volumes among older community-dwelling Japanese people
2022, Neurobiology of AgingCitation Excerpt :It was considered that the dendric complexity contributed to the brain volume (Chen et al., 2006; Fjell and Walhovd, 2010; Sugiyama et al., 2021). Appropriate neural membrane fluidity is important for maintaining the dendrite complex, and an age-related decrease in membrane fluidity was improved by LCPUFA intake (Fukaya et al., 2007; Suzuki et al., 1998). Another potential mechanism is that the increase in brain-derived neurotrophic factor (BDNF) by LCPUFA (Jiang et al., 2009) leads to larger dendritic branching (Chen et al., 2006; Horch and Katz, 2002).
Lymnaea stagnalis as model for translational neuroscience research: From pond to bench
2020, Neuroscience and Biobehavioral ReviewsAging and FADS1 polymorphisms decrease the biosynthetic capacity of long-chain PUFAs: A human trial using [U-<sup>13</sup>C]linoleic acid
2019, Prostaglandins Leukotrienes and Essential Fatty AcidsComprehensive analysis of the metabolomic characteristics on the health lesions induced by chronic arsenic exposure: A metabolomics study
2019, International Journal of Hygiene and Environmental HealthRole of the cell membrane interface in modulating production and uptake of Alzheimer's beta amyloid protein
2018, Biochimica et Biophysica Acta - Biomembranes