This study showed a reduction of GLUT4 protein content of insulin-sensitive tissues in an animal model of metabolic syndrome, a fundamental mechanism to impair glucose uptake and glucose homeostasis. This regulation, not yet seen, began at 3 months of age, and was not reverted as time passed. Furthermore, this glucose transporter modulation was accompanied by inflammation, insulin resistance and hypertension, also here described in the same animal model.
The use of MSG in genetically hypertensive rats led these animals to progressively increase body adiposity and hypertriglyceridemia; besides, developing and maintaining insulin resistance, low HDL-cholesterol, high blood pressure levels, and inflammation throughout the period studied. Other animal models used showed that high-fat-fed SHRs did not show changes in plasma concentrations of total cholesterol and triglycerides, although plasma concentrations of free fatty acids were higher as compared to normal diet-fed SHRs . Mice fed on high-carbohydrate and high-fat diet (cafeteria diet) had increased body weight, abdominal fat deposition, hyperinsulinaemia, hyperglycaemia and hyperleptinaemia, but no hypertension [36, 37]. Although these are examples of models of metabolic syndrome , our study clearly shows the possibility of reproducing the whole metabolic syndrome in laboratory animals in a low-cost and easily-obtained model.
The increased Lee index and epididymal fat mass observed in MetS animals confirmed their obese condition, as it has been described in MSG-treated normotensive rats and mice [32, 39]. MSG-treated animals, compared to their controls, may have a lower absolute weight, as we also found in MetS, which has been proposed to be a result of decreasing growth hormone (GH) secretion . Accordingly, lean mass may be decreased in MSG animals , a feature that we also observed in SHR treated with MSG, which presented lower heart and skeletal muscle mass. In addition to low GH levels, high corticosterone levels  were also described in MSG animals, and both hormonal modulations contribute to decrease synthesis and increase catabolism of proteins, thus diminishing muscle mass . Furthermore, together with decreasing sympathetic activity  this hormonal imbalance contributes to conserve energy, accumulating fat. It is important to highlight that MSG should be administered in the neonatal period to determine all the derangements related to obesity described, as use in later periods of life  would not result in the characteristic hypothalamic lesions [20, 21], because of the protective effects of the blood–brain barrier .
In metabolic syndrome, however, beyond obesity, hypertension is a key feature, which is not present when MSG is used in mice  and Wistar rats , but was effectively shown in the present study treating SHR. Other authors described attenuation of high BP levels in MSG-treated SHR [32, 39], a finding that we did not observe, probably because the cardiovascular evaluation method we used is more appropriate (direct BP measurement). The studies mentioned found controversial results, possibly due to the fact that all measured mean BP using tail plethysmography. Furthermore, variable protocols of neonatal MSG treatment have been used, probably altering severity and/or time of induction of the alterations. It is well-known that, in SHR, BP rises progressively over lifetime, while in our study, the MSG-treated SHR had higher BP than the normotensive rats at all ages studied, but remained stable, not increasing over time. This fact might be related to low sympathetic activity described in MSG-treated animals .
Fulfilling the picture of metabolic syndrome, MSG-treated SHR presented obesity and high BP levels associated with insulin resistance, hypertriglyceridemia and reduced HDL cholesterol levels with normal total cholesterol levels. Spontaneously hypertensive rats are typically insulin resistant , which we also observed in the MSG-treated SHR. However, SHR are not obese, and their visceral fat content is similar to that of normotensive Wistar rats , which was strongly changed by using MSG. Insulin resistance in SHR is partially ascribed to the characteristic sympathetic hyperactivity they present, which promotes reduced activity of the insulin signaling cascade and, consequently, can reduce GLUT4 translocation and/or expression . Furthermore, norepinephrine inhibits insulin-mediated glucose uptake in muscle  and blocks insulin inhibitory action on liver glucose production , all these effects contributing to impair glycemic homeostasis. The characteristic lipid profile of insulin-resistant states was observed from 6 months of age and beyond for the MSG-treated SHR, as observed for non-MSG treated SHR , but not for Wistar-Kyoto rats. Based on these facts, we can understand that increased sympathetic activity may induce or worsen installed insulin resistance, closing the circle which perpetuates the existence of both insulin resistance and high BP levels.
Beyond its classic metabolic actions, insulin is also anti-inflammatory, decreasing activity of pro-inflammatory cytokines, such as TNF-α and IL-6, as well as repressing the transcription factor nuclear factor кB (NFкB) . In MSG-treated mice, Furuya and collaborators  demonstrated that hypertrophyc adipocytes triggered local inflammatory activity with increased macrophage infiltration and TNF-α and IL-6 expression, depicting high plasma concentration of the cytokines. This was accompanied by decreased GLUT4 content in white adipose tissue and reversed by atorvastatin treatment . We showed that inflammation is exacerbated in MSG-treated SHR from 3 months of age on, along with GLUT4 reduction in all insulin-sensitive tissues. TNF-α rise, especially, can reduce the expression of IRS-1 and GLUT4, as well as of the hormone-sensitive lipase, adiponectin and PPARγ . All these processes are known to contribute to lipolysis and insulin resistance, and were also observed in these animals since the age of 3 months. Adiponectin reduction in MSG-treated SHR at the age of 9 months is probably due to the exacerbation of hypertriglyceridemia at this age, which acts as an independent causal factor for hypoadiponectinemia . Besides, hypoadiponectinemia probably occurs only at the age of 9 months because of its known relationship with the severity of obesity .
Insulin resistance of MSG-treated SHR is in accordance with the reduction of GLUT4 content in all insulin-sensitive tissues analyzed, corroborating previous data in MSG-treated mice in all cell fractions of adipose tissue with no change in the relative GLUT4 translocation to the plasma membrane  and also in the same tissue in humans . Moreover, low GLUT4 protein content in skeletal muscle and heart was also shown in MSG-treated mice . In MSG-treated SHR, GLUT4 on insulin-sensitive tissues had not been reported yet, especially its time-course changes through aging, as here reported.
Skeletal and heart GLUT4 protein were reduced from the age of 3 months and did not worsen over time, following the stable profile of insulin resistance in MSG-treated SHR. It is known that in the SHR, plasma membrane myocyte GLUT4 increases with age (12 weeks), but at 20 weeks, GLUT4 contents tends to reach the initial levels (8 weeks) . Moreover, reduced insulin-induced GLUT4 translocation  and/or total content was also showed before . Pharmacological treatment of arterial hypertension with captopril can virtually re-establish the glucose transporter contents . On the other hand, investigations have shown that GLUT4 translocation in skeletal muscle and heart is stimulated by bradykinin , which enhances insulin-induced phosphorylation of insulin receptors and insulin-stimulated association of IRS-1 and phosphatidylinositol-3-kinase in skeletal muscle of aged rats , all of which are essential for insulin-mediated GLUT4 translocation and glucose transport. Both cases demonstrate that the amount of GLUT4 is related to blood pressure levels. Anyhow, it is known that the behavior of this transporter in the skeletal muscle does not always reflect what happens in the white adipose tissue , and, indeed, in this tissue, we find a progressive reduction of its content in MSG-treated SHR, especially at the age of 9 months.
In the heart, in turn, it is known that insulin resistance together with the excess of free fatty acids – as we find in MSG-treated SHR – are responsible for contractility dysfunction [61, 62]. Since the muscle contraction also induces GLUT4 translocation to the plasma membrane, contractility dysfunction is directly related to reduced plasma membrane GLUT4 content . Furthermore, the excess of free fatty acids alone can interfere in glucose transport, since it reduces GLUT4 expression in the heart, but not that of the free fatty acid transporter . In rats with insulin resistance induced by a fructose-rich diet, cardiomyocytes present low glucose input in response to ischemia, a consequence of GLUT4 translocation reduction . We found a reduction of GLUT4 content in heart of MSG-treated SHR at the age of 3, 6 and 9 months, which can be explained by the hypothesis that the reduction of this transporter in the heart occurs when obesity and insulin resistance are established . The normotensive and hypertensive control groups did not present differences in the GLUT4 content in the heart, at the age of 6 months . In the MSG-treated SHR, time-course of changes of insulin-sensitive GLUT4 protein content had not yet been described in the literature.