Chlorella vulgaris reduces the impact of stress on hypothalamic–pituitary–adrenal axis and brain c-fos expression
Introduction
Stressors are processed within the brain and this involves activation of several important groups of neurons. Immediate early genes, such as c-fos, are extensively used to map brain areas involved in stress responses (Armario, 2006b). Studies indicate that inputs from different brain regions converge upon the neurons of the hypothalamic paraventricular nucleus (PVN), particularly the medial dorsal parvocellular division (mpdPVN). The neurons from the mpdPVN, which synthesize and release corticotropin-releasing factor (CRF) and other secretagogues, represent the origin of the hypothalamic–pituitary–adrenal (HPA) axis activation, culminating in the release of adrenocorticotropic hormone (ACTH) and the subsequent ACTH-induced release of glucocorticoids from the adrenal gland (Armario, 2006a). Increase in levels of mRNA for CRF occurs in the mpdPVN 2–4 h after initial exposure to stress. However intronic probes (which detect nuclear primary transcripts, hnRNA) are needed to analyze the CRF response to brief exposures or low intensity stressors (i.e., Rivier and Lee, 1996).
Chlorella vulgaris (CV) is a microscopic single-celled freshwater algae that is reported to be a rich source of antioxidants, such as lutein, α- and β-carotene, ascorbic acid and tocopherol, also supplying large quantities of vitamins, essential fatty acids, minerals and dietary fiber (Rodriguez-Garcia and Guil-Guerrero, 2008). Treatment with CV demonstrated protective activities against infections, tumors, stress and high-fat-diet-induced insulin resistance (i.e., Dantas and Queiroz, 1999, Ramos et al., 2010, Souza Queiroz et al., 2013, Vecina et al., 2014). Distinct Chlorella-based molecules, as glycoproteins and polysaccharides, were isolated and they seem to justify some of its immunostimulatory effects (i.e., Noda et al., 1996, Pugh et al., 2001). Of interest, we demonstrated that CV treatment increases the levels of INF-γ and reduces the levels of IL-10 from animals exposed to stress. These effects were related with increased resistance to Listeria monocytogenes infection and reduced suppression of myelopoiesis (Souza Queiroz et al., 2008). Likewise, by using the method of long term bone marrow culture, we found CV treatment to reduce the negative impact that both single and repeated stressors had over the production of myeloid and lymphoid cells and on the production of IL-1α and IL-6 (Souza Queiroz et al., 2013).
Considering the aforementioned results obtained with treatment with CV in stressful situations, in the present study we aim to further investigate the action of this algae. Therefore, we evaluate the effects of CV on the activation of the HPA axis, hyperglycemia and c-fos expression in key brain areas in response to forced swimming. An additional experiment was performed to test whether two of the most abundant fatty acids found in CV composition, α-linonelic (ALA) and linoleic (LA) acids, were responsible for the effect of CV on ACTH response to stress.
Section snippets
Animals
Male 2–3 months old Sprague–Dawley rats bred at the Animal Facility Service of the Universitat Autònoma de Barcelona were used. The animals were housed in pairs in polypropylene opaque wide-topped, solid-bottom cages (21.5 × 46.5 × 14.5 cm; “Type 1000 cm2”, Panlab S.L.U., Barcelona, Spain) containing wood shaving bedding (Lignocel ¾, Harlan Interfauna Ibérica, Barcelona, Spain), kept at standard conditions of temperature (21 ± 1 °C) and a 12:12 h light/dark schedule (lights on at 8:00 h). Food (SAFE-diet
Experiment 1
Fig. 1 shows the effects of treatment with CV (50 mg/kg or 200 mg/kg) on levels of corticosterone and glucose at 1 h, 2 h and 4 h after treatment (Figs. 1A and B) and on the plasma levels of ACTH (Fig. 1C), corticosterone (Fig. 1D) and glucose (Fig. 1E) before and after forced swimming. GEE analysis of corticosterone levels (Fig. 1A) revealed marginally significant effect of CV (Wald X2(2) = 5.8, P = 0.054), significant effect of sampling time (Wald X2(2) = 25.0, P < 0.001), and no effect of the interaction
Discussion
The results from these two experiments demonstrate that a single oral dose of CV given the day before stress application has a modest effect per se, but markedly reduced both peripheral and central responses to forced swimming stress, suggesting an important anti-stress brain locus of action. To the best of our knowledge, this is the first study to demonstrate the efficacy of an acute dose of CV. Our results were observed despite no influence on behavior during swimming, indicating that the
Conclusions
The data presented in this study reinforces the usefulness of CV to reduce the impact of stressors. Its effect was observed both peripherally and centrally, with the reduction of the hnCRF response in the mpdPVN and the marked and widespread inhibition of c-fos expression in most brain regions. Although the ultimate mechanisms of this action need additional studies, these findings can provide a venue for promoting the translation of this understanding into effective prevention strategies to
Acknowledgements
This project received financial support from CNPq (300764/2010-3; 200033/2011-5 and 142853/2008-9), FAPESP (09/51866-3) and from Ministerio de Economía y Competitividad (SAF2014-53876-R), Instituto de Salud Carlos III (RD12/0001/0015, Redes Temáticas de Investigación Cooperativa en Salud, Ministerio de Sanidad y Consumo) and Generalitat de Catalunya (SGR2014-1020).
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2019, Brain Research BulletinCitation Excerpt :Several health-protecting effects are attributed to CV, including anti-atherogenic, antitumoral, hepatoprotective, antioxidant, anti-inflammatory, and anti-hypertensive activities. Due to those supposed beneficial effects, thousands of tons of CV are produced in several countries each year and are sold widely as a nutraceutical (Panahi et al., 2015a, 2016; Souza Queiroz et al., 2016). So far, only one study (with an open-label, randomized design) has been conducted to assess the efficacy of adjunctive CV to conventional AD treatment (Panahi et al., 2015a).