Prior Repeated Stress Attenuates Cold-Induced Immunomodulation Associated with "Browning" in Mesenteric Fat of Rats.

Continuous exposure to cold leads to activation of adaptive thermogenesis in brown adipose tissue but also to induction of brown/beige cell phenotype in white adipose tissue. The aim of this work was to investigate whether prior exposure to immobilization (IMO) stress may affect immune response associated with adipocyte "browning" in mesenteric adipose tissue (mWAT). In the first experiment, Sprague-Dawley rats were exposed to acute (3 h) or prolonged (7 days) cold exposure (4 ± 1 °C). 7-day cold stimulated gene expression of uncoupling protein 1 and other "browning"-associated factors. In the second experiment, rats were immobilized for 7 days (2 h daily) followed by exposure to continuous cold for 1 or 7 days. Prior IMO exaggerated cold-induced sympathetic response manifested by elevated tyrosine hydroxylase (TH) protein and norepinephrine in mWAT. Induction of non-sympathetic catecholamine production demonstrated by elevated TH and PNMT (phenylethanolamine N-methyltransferase) mRNAs was observed after 7-day cold; however, prior IMO attenuated this response. 7-day cold-induced gene expression of anti-inflammatory mediators (IL-4, IL-13, IL-10, adiponectin), markers of M2 macrophages (Arg1, Retnlα), and eosinophil-associated molecules (eotaxin, IL-5), while inhibited expression of pro-inflammatory cytokines (IFNγ, IL-1b, IL-6, IL-17) and monocytes (MCP-1, Ly6C). This immune response was accompanied by elevated expression of uncoupling protein-1 and other thermogenic factors. Rats exposed to prior IMO exhibited inhibition of cold-induced immune and "browning"-related expression pattern. Overall, we demonstrated that 7-day cold-induced browning"-associated changes in rat mWAT, while prior history of repeated stress prevented this response.

Repeated Stress Exaggerates Lipopolysaccharide-Induced Inflammatory Response in the Rat Spleen.

Spleen is an immune organ innervated with sympathetic nerves which together with adrenomedullary system control splenic immune functions. However, the mechanism by which prior stress exposure modulates the immune response induced by immunogenic challenge is not sufficiently clarified. Thus, the aim of this study was to investigate the effect of a single (2 h) and repeated (2 h daily for 7 days) immobilization stress (IMO) on the innate immune response in the spleen induced by lipopolysaccharide (LPS, 100 µg/kg). LPS elevated splenic levels of norepinephrine and epinephrine, while prior IMO prevented this response. LPS did not alter de novo production of catecholamines, however, prior IMO attenuated phenylethanolamine N-methyltransferase gene expression. Particularly repeated IMO exacerbated LPS-induced down-regulation of α1B- and ß1-adrenergic receptors (ARs), while enhanced α2A- and ß2-AR mRNAs. Elevated expression of inflammatory mediators (iNOS2, IL-1ß, IL-6, TNF-α, IL-10) was observed following LPS and repeated IMO again potentiated this effect. These changes were associated with enhanced Ly6C gene expression, a monocyte marker, and elevated MCP-1, GM-CSF, and CXCL1 mRNAs suggesting an increased recruitment of monocytes and neutrophils into the spleen. Additionally, we observed increased Bax/Bcl-1 mRNA ratio together with reduced B cell numbers in rats exposed to repeated IMO and treated with LPS but not in acutely stressed rats. Altogether, these data indicate that repeated stress via changes in CA levels and specific α- and ß-AR subtypes exaggerates the inflammatory response likely by recruiting peripheral monocytes and neutrophils to the spleen, resulting in the induction of apoptosis within this tissue, particularly in B cells. These changes may alter the splenic immune functions with potentially pathological consequences.

Lipopolysaccharide induces catecholamine production in mesenteric adipose tissue of rats previously exposed to immobilization stress.

Catecholamines (CAs) are mainly produced by sympathoadrenal system but their de novo production has been also observed in adipose tissue cells. The aim of this work was to investigate whether immune challenge induced by lipopolysaccharide (LPS) modulates biosynthesis of CAs in mesenteric adipose tissue (MWAT), as well as whether previous exposure to immobilization (IMO) stress could modulate this process. Sprague-Dawley rats were exposed to single (2 h) or repeated (2 h/7 days) IMO and afterwards injected with LPS (i.p., 100 µg/kg body weight) and sacrificed 3 h later. LPS did not alter CA biosynthesis in MWAT in control rats. Single and repeated IMO elevated CAs and expression of CA biosynthetic enzymes in MWAT, including adipocyte and stromal/vascular fractions (SVF). Repeated IMO followed by LPS treatment led to the up-regulation of CA-biosynthetic enzymes expression, elevation of CAs in SVF but depletion of norepinephrine and epinephrine in adipocyte fraction. Prior IMO caused a marked LPS-induced macrophage infiltration in MWAT as evaluated by F4/80 expression. A positive correlation between expression of tyrosine hydroxylase and F4/80 suggests macrophages as the main source of LPS-induced CA production in MWAT. Furthermore, prior exposure to the single or repeated IMO differently affected immune responses following LPS treatment by modulation of inflammatory cytokine expression. These data suggest that stress might be a significant modulator of immune response in MWAT via stimulation of the macrophage infiltration associated with cytokine response and de novo production of CAs.

Kinetics of oxytocin response to repeated restraint stress and/or chronic cold exposure.

Recently, several new atypical actions of circulating oxytocin are emerging, which may be of importance for the physiological effects of oxytocin released during stress. However, little information is available on oxytocin response to chronic stress stimuli. The aim of the present study is to deepen the knowledge on oxytocin secretion during chronic and repeated stress. The main hypothesis to be tested was that oxytocin release in response to single and to repeated or chronic stress is of different kinetics. Adult male Sprague-Dawley rats were exposed to 2 different stress stimuli or their combination. Restraint (immobilization) of different duration (10-120 min) and number of repetitions (1 or 7 times) as well as chronic exposure (28 days) to cold temperature were used. Concentrations of oxytocin in plasma and posterior pituitary were measured by a radioimmunoassay. Concentrations of oxytocin in plasma increased significantly in response to both single and repeated immobilization. Acute immobilization caused rapid increase already after 10 min of restraint, while the recovery occurred only after 24 h. Repeated restraint caused delayed onset of increased oxytocin release and a more rapid recovery to prestress levels after 3 h. In conclusion, the results of the present study show that though with a different kinetics, increased oxytocin release is preserved during repeated exposure to an intensive stressor, namely immobilization for 120 min. During repeated exposure to shorter stressors, an adaptation in oxytocin responses may occur. This should be taken into account with respect to cardiovascular and metabolic effects of stress-induced oxytocin.

Stress stimulates production of catecholamines in rat adipocytes.

The sympathoadrenal system is the main source of catecholamines (CAs) in adipose tissues and therefore plays the key role in the regulation of adipose tissue metabolism. We recently reported existence of an alternative CA-producing system directly in adipose tissue cells, and here we investigated effect of various stressors-physical (cold) and emotional stress (immobilization) on dynamics of this system. Acute or chronic cold exposure increased intracellular norepinephrine (NE) and epinephrine (EPI) concentration in isolated rat mesenteric adipocytes. Gene expression of CA biosynthetic enzymes did not change in adipocytes but was increased in stromal vascular fraction (SVF) after 28 day cold. Exposure of rats to a single IMO stress caused increases in NE and EPI levels, and also gene expression of CA biosynthetic enzymes in adipocytes. In SVF changes were similar but more pronounced. Animals adapted to a long-term cold exposure (28 days, 4°C) did not show those responses found after a single IMO stress either in adipocytes or SVF. Our data indicate that gene machinery accommodated in adipocytes, which is able to synthesize NE and EPI de novo, is significantly activated by stress. Cold-adapted animals keep their adaptation even after an exposure to a novel stressor. These findings suggest the functionality of CAs produced endogenously in adipocytes. Taken together, the newly discovered CA synthesizing system in adipocytes is activated in stress situations and might significantly contribute to regulation of lipolysis and other metabolic or thermogenetic processes.

Modulation of the sodium-calcium exchanger in the rat kidney by different sequential stressors.

Stress, if exaggerated, modulates a variety of metabolic pathways and results in development of serious health consequences. The cell membrane sodium-calcium exchanger (NCX) is a major calcium extrusion system and is also modulated by stress. It has been shown previously that mRNA, protein levels and activity of the type 1 NCX (NCX1) in the left ventricle of the rat heart are increased by stressors, such as immobilization or hypoxia. In this study we investigated whether exposure to a subsequent different stressor can affect gene expression, protein level and activity of the NCX1 in rat kidney compared to exposure to only one type of stressor. In these experiments, we used immobilization and cold as the model stressors.We found that cold exposure at 4 degrees C for 24 h, when applied after immobilization repeated seven times, completely abolished the immobilization-induced increase in NCX mRNA level and after 7 days cold exposure the increases in NCX1 protein and activity in rat kidney were also abolished. Permanently increased NCX1 expression can result in imbalance of cellular calcium homeostasis and thus contribute to kidney dysfunction. Based on our results, we conclude that exposure to a cold stressor can have a protective effect on the kidney in rats exposed previously to repeated immobilization stress. This might be explained by differential stimulation of sympathetic neural and adrenal medullary responses by these different stressors.

Neuroendocrine and cardiovascular parameters during simulation of stress-induced rise in circulating oxytocin in the rat.

Physiological functions of oxytocin released during stress are not well understood. We have (1) investigated the release of oxytocin during chronic stress using two long-term stress models and (2) simulated stress-induced oxytocin secretion by chronic treatment with oxytocin via osmotic minipumps. Plasma oxytocin levels were significantly elevated in rats subjected to acute immobilization stress for 120 min, to repeated immobilization for 7 days and to combined chronic cold stress exposure for 28 days with 7 days immobilization. To simulate elevation of oxytocin during chronic stress, rats were implanted with osmotic minipumps subcutaneously and treated with oxytocin (3.6 microg/100 g body weight/day) or vehicle for 2 weeks. Chronic subcutaneous oxytocin infusion led to an increase in plasma oxytocin, adrenocorticotropic hormone, corticosterone, adrenal weights and heart/body weight ratio. Oxytocin treatment had no effect on the incorporation of 5-bromo-2-deoxyuridine into DNA in the heart ventricle. Mean arterial pressure response to intravenous phenylephrine was reduced in oxytocin-treated animals. Decrease in adrenal tyrosin hydroxylase mRNA following oxytocin treatment was not statistically significant. Oxytocin treatment failed to modify food intake and slightly increased water consumption. These data provide evidence on increased concentrations of oxytocin during chronic stress. It is possible that the role of oxytocin released during stress is in modulating hypothalamic-pituitary-adrenocortical axis and selected sympathetic functions.

Decreased stress responsivity of central and peripheral catecholaminergic systems in aged 344/N Fischer rats.

We investigated the effects of stress on central and peripheral sympatho-adrenal and sympatho-neural functions in healthy, intact young (3-4 mo) and aged (24 mo) male Fischer 344/N rats. Extracellular fluid (ECF) levels of the catecholamines norepinephrine (NE), dihydroxyphenylglycol (DHPG), methoxyhydroxyphenylglycol (MHPG), and dihydroxyphenylacetic acid (DOPAC) were obtained by microdialysis in the paraventricular nucleus (PVN) of the hypothalamus at baseline and during immobilization (IMMO). The baseline levels of these substances were similar in both age groups, and their concentrations increased significantly in response to IMMO. The IMMO-induced increases of NE and MHPG, however, were significantly smaller in old than in young rats. Plasma levels of the catecholamines NE, DHPG, MHPG, DOPAC, dihydroxyphenylalanine (DOPA), epinephrine (EPI), dopamine (DA), and HVA were also determined in young and old rats during IMMO. Basal levels of these substances were significantly higher in old than in young rats. The magnitude of the IMMO-induced increases in the majority of these compounds however, was significantly smaller in old than in young rats. We conclude that, at the basal state, aging in the Fischer rat is associated with normal PVN ECF, but high plasma catecholamine levels; at stress state, however, old rats have substantially lesser activation of their central and peripheral catecholaminergic systems than young rats.

The peripheral noradrenergic terminal as possible site of action of salsolinol as prolactoliberin.

Salsolinol, an endogenous isoquinoline, induces selective prolactin release in rats [Tóth, B.E., Homicskó, K., Radnai, B., Maruyama, W., DeMaria, J.E., Vecsernyés, M., Fekete, M.I.K., Fülöp, F., Naoi, M., Freeman, M.E., Nagy, G.M., 2001. Salsolinol is a putative neurointermediate lobe prolactin releasing factor. J. Neuroendocrinol. 13, 1042-1050]. The possible role of dopaminergic and adrenergic signal transduction was investigated to learn the mechanism of this action. The effect of salsolinol (10mg/kg i.v.) was inhibited by reserpine treatment (2.5mg/kg i.p.) and reinstated by pretreatment with monoamine oxidase inhibitor (pargyline 75 mg/kg i.p.). Salsolinol did not affect the in vitro release of dopamine (DA) in the median eminence, and did not inhibit the L-DOPA induced increase of DA level in the median eminence. 1-Methyl dihydroisoquinoline (1MeDIQ) is an antagonist of salsolinol induced prolactin release and causes increase in plasma NE level [Mravec, B., Bodnár, I., Fekete, M.I.K., Nagy, G.M., Kvetnansky, R., 2004. An antagonist of prolactoliberine induces an increase in plasma catecholamine levels in the rat. Autonom. Neurosci. 115, 35-40]. Using tissue catecholamine contents as indicators of the interaction between salsolinol and 1MeDIQ we found no interaction between these two agents to explain the changes in prolactin release in the median eminence, lobes of the pituitary, superior cervical and stellate ganglion. Increasing doses of salsolinol caused a dose dependent decrease of tissue dopamine concentration and increase of NE/DA ratio in the salivary gland, atrium and spleen. These changes of DA level and NE/DA ratio run parallel in time with the increase of prolactin release. 1MeDIQ antagonized the increase of prolactin release and decrease of tissue DA content caused by salsolinol. Neither this increase of prolactin secretion nor the decrease of DA level in spleen could be demonstrated in NE transporter (NET) knock out mice. The results presented argue for the possible role of peripheral norepinephrine release as a target for salsolinol in its action releasing prolactin. The dominant role of norepinephrine transporter may be suggested.

Cardiovascular diseases and molecular variants of the renin-angiotensin system components in Slovak population.

Cardiovascular diseases associated with molecular variants of individual components of renin-angiotensin system are reported to constitute inherited predisposition in humans. Molecular variant frequencies are race- and population-dependent. We examined frequencies of the M235T variant of angiotensinogen gene and I/D polymorphism of gene for angiotensin-converting enzyme in Slovak population: in hypertensive patients, coronary heart disease (CHD), dilated cardiomyopathy (DCM) and myocardial infarction (MI) patients compared to healthy subjects. Frequency of M235T was significantly increased in hypertensive, CHD and DCM patients compared to controls (0.48 and 0.50 vs. 0.40, p < 0.001). Significant increase in D allele frequency compared to controls was observed in the group of patients after MI (0.58 vs. 0.50, p < 0.001), CHD (0.59 vs. 0.50, p < 0.001) and DCM (0.60 vs. 0.50, p < 0.001). These results correlate with other Caucasian populations. In Slovak population, M235T is associated with increased blood pressure and D allele of ACE gene is associated with MI, chronic CHD and DCM, rather than with hypertension. Our results suggest that in Slovak population, D alelle and M235T variant represent a risk factor for several cardiovascular diseases and these polymorphisms might have a cumulative effect on development of cardiovascular diseases.

Modulation of expression of the sigma receptors in the heart of rat and mouse in normal and pathological conditions.

The aim of the present work was to study the effect of various stressors (hypoxia, cold, immobilization) on the gene expression of sigma receptors in the left ventricles of rat heart. We have clearly shown that gene expression of sigma receptors is upregulated by strong stress stimuli, such as immobilization and/or hypoxia. Nevertheless, cold as a milder stressor has no effect on sigma receptor's mRNA levels. Signalling cascade of sigma receptors is dependent on IP(3) receptors, since silencing of both, type 1 and 2 IP(3) receptors resulted in decreased mRNA levels of sigma receptors. Physiological relevance of sigma receptors in the heart is not clear yet. Nevertheless, based on the already published data we can assume that sigma receptors might participate in contractile responses in cardiomyocytes.

Stress-triggered activation of gene expression in catecholaminergic systems: dynamics of transcriptional events.

Stress triggers important adaptive responses that enable an organism to cope with a changing environment. However, when prolonged or repeated, stress can be extremely harmful. The release of catecholamines is a key initial event in responses to stressors and is followed by an increase in the expression of genes that encode catecholamine-synthesizing enzymes. This process is mediated by transcriptional mechanisms in the adrenal medulla and the locus coeruleus. The persistence of transcriptional activation depends on the duration and repetition of the stress. Recent work has begun to identify the various transcription factors that are associated with brief or intermediate duration of a single or repeated stress. These studies suggest that dynamic interplay is involved in converting the transient increases in the rate of transcription into prolonged (potentially adaptive or maladaptive) changes in gene expression.

Insertion/deletion polymorphism on ACE gene is associated with endothelial dysfunction in young patients with hypertension.

Endothelial dysfunction, insulin resistance (IR) and genetic predispositions are important risk factors of hypertension. Aim of our study was to test the hypothesis, whether insertion/deletion (I/D) polymorphism on the angiotensin converting enzyme (ACE) gene and M235T polymorphism on angiotesinogen gene (AGT) correlates with parameters of insulin sensitivity and plasminogen activator inhibitor (PAI-1) levels in newly diagnosed hypertensive patients as compared with normotensive controls. Blood pressure (BP), fasting plasma glucose, insulin, epinephrine, norepinephrine and PAI-1 concentrations were determined in 30 male patients with hypertension grade 1 (HT) and in 31 matched healthy subjects (NT). Insulin resistance was estimated using IR HOMA formula. Patients with HT had increased levels of PAI-1, norepinephrine, fasting plasma insulin levels, IR HOMA (p<0.001) compared to controls. Subjects (HT and NT) with DD and ID genotype had a significantly higher systolic BP (p<0.05) and PAI-1 compared to those with II genotype. Homozygous subjects 235T had a higher systolic BP and higher levels of epinephrine and norepinephrine than heterozygous or homozygous M235 (p<0.05). In conclusion, no association was found between M235T polymorphism and insulin resistance or PAI-1 levels, but results indicate relationship between I/D polymorphism of the ACE gene and plasma PAI-1 levels in the early stage of hypertension.

Decreased pituitary response to insulin-induced hypoglycaemia in young lean male patients with essential hypertension.

Essential hypertension is associated with changes in central catecholaminergic pathways which might also be reflected in the pituitary response to stress stimuli. The aim of this study was to determine whether the response of pituitary hormones, cortisol, plasma renin activity, aldosterone and catecholamines to insulin-induced hypoglycaemia is changed in hypertension. We studied 22 young lean male patients with newly diagnosed untreated essential hypertension and 19 healthy normotensive, age- and body mass index (BMI)-matched controls. All subjects underwent an insulin tolerance test (0.1 IU insulin/kg body weight intravenously) with blood sampling before and 15, 30, 45, 60 and 90 min after insulin administration. Increased baseline levels of norepinephrine (P<0.05), increased response of norepinephrine (P<0.001) and decreased response of growth hormone (P<0.001), prolactin (P<0.001), adrenocorticotropic hormone (P<0.05) and cortisol (P<0.001) were found in hypertensive patients when compared to normotensive controls. Increased norepinephrine levels and a decreased pituitary response to metabolic stress stimuli may represent another manifestation of chronically increased sympathetic tone in early hypertension.

IP3 type 1 receptors in the heart: their predominance in atrial walls with ganglion cells.

Previously we have shown that inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) are abundantly expressed in the atria of rat hearts. Since arrangement of atria is very heterogeneous, in this work we focused on the precise localization of IP3 receptors in the left atrium, where the gene expression of the type 1 IP3R was the highest. The mRNA levels of the IP3 type 1 receptors in the left atrium, left ventricle and myocytes were determined using real-time polymerase chain reaction and Taqman probe. For precise localization, immunohistochemistry with the antibody against type 1 IP3Rs was performed. The mRNA of type 1 IP3 receptor was more than three times higher in the left atrium than in the left ventricle, as determined by real-time PCR. Expression of the type 1 IP3 receptor mRNA was higher in the atria, especially in parts containing cardiac ganglion cells. The atrial auricles, which are particularly free of ganglion cells, and the ventricles (wall of the right and left ventricle and ventricular septum) contained four to five times less IP3 receptors than atrial samples with ganglia. IP3R type 1 immunoreactivity detected by a confocal microscope attributed the most condensed signal on ganglionic cells, although light immunoreactivity was also seen in cardiomyocytes. These results show that type 1IP3 receptors predominate in intrinsic neuronal ganglia of cardiac atria.

Gene expression of the phenylethanolamine N-methyltransferase is differently modulated in cardiac atria and ventricles.

Phenylethanolamine N-methyltransferase (PNMT) is a final enzyme in catecholamine synthesizing cascade that converts noradrenaline to adrenaline. Although most profuse in adrenal medulla, PNMT is expressed also in the heart, particularly in cardiac atria and ventricles. In atria, the PNMT mRNA is much more abundant compared to ventricles. In present study we aimed to find out whether there is a difference in modulation of the PNMT gene expression in cardiac atria and ventricles. We used three methodological approaches: cold as a model of mild stress, hypoxia as a model of cardiac ischemic injury, and transgenic rats (TGR) with incorporated mouse renin gene (mREN-2)27, to determine involvement of renin-angiotensin pathway in the PNMT gene expression. We have found that PNMT gene expression was modulated differently in cardiac atria and ventricles. In atria, PNMT mRNA levels were increased by hypoxia, while cold stress decreased PNMT mRNA levels. In ventricles, no significant changes were observed by cold or hypoxia. On the other hand, angiotensin II elevated PNMT gene expression in ventricles, but not in atria. These results suggest that PNMT gene expression is modulated differently in cardiac atria and ventricles and might result in different physiological consequences.

The effects of stress on muscarinic receptors. Heterologous receptor regulation: yes or no?

1 Stress is usually comprehended as an event affecting mainly the catecholaminergic system, the hypothalamo-pituitary-adrenocortical (HPA) axis and the receptor systems connected to these neurotransmitters/hormones. Other neurotransmitter/hormone systems can be affected too. Here we review the available data on the effects of different stressful stimuli (physical, chemical, psychological/social, cardiovascular, affecting multiple system) on muscarinic receptors (MR). 2 The data suppose the existence of specific mechanisms that regulate the signalization through MR during different type of stress. 3 Physical stressors (cold vs. heat) reveal opposite type of changes on peripheral-tissue MRs. Chemical stressors (oxidative stress) are tightly connected with MR and it is especially interesting that the sensitivity of MR to oxidative stress is subtype-specific. It is also suggested that heterologous regulation can occur with psychological/social stressors on the organism. Cardiovascular system-disturbing stressors cause imbalance between autonomic receptors or down-regulate MR in the peripheral tissue. Immobilization caused opposite effects on MR in the central nervous system and periphery, where the changes are supposed to be due to heterologous regulation between receptor systems. 4 In conclusion, some data indicate that in specific conditions MR are regulated as a consequence of other changes rather than as a primary effect of stress. On the contrary, in some situations, MR are the first targets to respond to the stress. 5 These findings on stress-induced activity of the cholinergic system and changes in muscarinic receptors support the view that stress is a specific response of the organism.

Developmental changes of some G-protein coupled receptors affected by c-fos knock-out.

The role of c-fos gene disruption on binding characteristics of selected G protein-coupled receptors has been investigated. The following receptors were studied: muscarinic receptors (MR), alpha1-adrenoceptors (AAR), beta-adrenoceptors (BAR), D1-like dopamine receptors (D1R), D2-like dopamine receptors (D2R). We have tested the hypothesis that c-fos gene disruption would not influence the receptor density in central nervous system (CNS: brain cortex and cerebellum) and in the periphery (lung, heart). We found that both in the periphery and in the CNS there were important changes in receptor density. Both MR and BAR were increased in the lung and heart. The effects of c-fos gene disruption in CNS were more selective. In general, the receptors that activate Gq-phospholipase C-proteinkinase C pathway (AAR, MR) were affected, while the others (that activate/or inhibit adenylyl cyclase: BAR, D1R, D2R) were not. These results show that disruption of c-fos gene could significantly change the expression of G protein-coupled receptors. Moreover, these changes could be comprehended as one of the adaptive mechanisms that help the organism to cope with c-fos gene disruption.

Continuous cold exposure induces an anti-inflammatory response in mesenteric adipose tissue associated with catecholamine production and thermogenin expression in rats.

OBJECTIVE: Continuous exposure to cold leads to an activation of adaptive thermogenesis in the brown adipose tissue and induction of brown/beige cell phenotype in the white adipose tissue. Thermogenic response is associated with alternatively activated macrophages producing catecholamines, which subsequently activate the uncoupling protein 1 (UCP-1). The aim of this work was to elucidate the effect of cold exposure on catecholamine and immune responses associated with adipocyte browning in the mesenteric adipose tissue (mWAT) of rat. METHODS: The rats were exposed to continuous cold (4 °C) for 1 or 7 days. Catecholamines production and gene expressions of inflammatory and other factors, related to adipocyte "browning", were analyzed in the homogenized mWAT samples using 2-CAT ELISA kits. RESULTS: Cold exposure induced a sympathetic response in the mWAT, evidenced by the tyrosine hydroxylase (TH) protein level rise. Induction of non-sympathetical catecholamine production was observed 7 days after cold exposure by elevated TH and phenylethanolamine-N-methyltransferase (PNMT) expression, leading to an increased epinephrine levels. Cold exposure for 7 days stimulated the infiltration of macrophages, evaluated by F4/80 and CD68 expressions, and expression of anti-inflammatory mediators, while pro-inflammatory cytokines were inhibited. Anti- inflammatory response, accompanied by de novo catecholamine production and up-regulation of ß3-adrenergic receptors, led to the stimulation of UCP-1 and PGC1α expression, suggesting a cold-induced "browning" of the mWAT, mediated by alternatively activated macrophages. CONCLUSIONS: The present data indicate that prolonged cold exposure may induce anti-inflammatory response in mWAT associated with induction of UCP-1 expression. Although functional thermogenesis in the mWAT is most likely redundant, a highly efficient dissipation of energy by UCP1 may affect the energy homeostasis in this visceral fat.

Mapping of genetic determinants of the sympathoneural response to stress.

Activation of the sympathoadrenal system (SAS, comprising the sympathetic nervous system and the adrenal medulla) in response to stressful stimuli is an important defense mechanism as well as a contributor to several cardiovascular diseases. There is variability in the SAS response to stress, although the extent to which this is genetically regulated is unclear. Some rodent models, including the hereditary hypertriglyceridemic (hHTg) rat, are hyperresponsive to stress. We investigated whether quantitative trait loci (QTLs) that affect sympathoadrenal response to stress could be identified. Second filial generation rats (n = 189) derived from a cross of the hHTg rat and the Brown Norway rat had plasma norepinephrine (NE) and epinephrine (Epi) levels, indices of activation of the sympathoneural and adrenal medulla components, respectively, measured in the resting state and in response to an immobilization stress. Responses were assessed early (20 min) and late (120 min) after the application of the stress. A genome scan was conducted using 153 microsatellite markers. Two QTLs (maximum peak LOD scores of 4.17 and 3.52, respectively) influencing both the early and late plasma NE response to stress were found on chromosome 10. Together, the QTLs accounted for approximately 20% of the total variation in both the early and late NE responses in the F(2) rats. Interestingly, the QTLs had no effect on plasma Epi response to stress. These findings provide evidence for a genetic determination of the response of a specific component of the SAS response to stress. Genetically determined variation in sympathetic nervous system response to stress may contribute to cardiovascular diseases.