Urinary Norepinephrine Excretion Research Articles

Abstract P100: Associations Between Urinary Biomarkers at Baseline and Salt and Potassium Sensitivity: The GenSalt Study

Introduction: Salt sensitivity of blood pressure is associated with an increased risk of hypertension and cardiovascular disease. Measurement methods of salt sensitivity of BP are inconvenient thus identifying biomarkers for salt sensitivity of BP has important clinical significance. Hypothesis: The urinary excretion of dopamine, kallikrein, norepinephrine and uromodulin at baseline are associated with salt sensitivity of BP. Methods: The Genetic Epidemiology Network of Salt Sensitivity (GenSalt) (n=1887) and GenSalt Replication (n=690) studies dietary interventions included a 3-day baseline exam, 7-day low sodium feeding, a 7-day high sodium feeding, and a 7-day high sodium feeding with potassium supplementation. Baseline urinary dopamine, kallikrein, norepinephrine, and uromodulin were measured by ELISA methods. Salt and potassium sensitivity were defined as change in SBP and DBP during dietary intervention. Urinary biomarkers below limit of detection (LOD) were replaced using LOD/√2 and censored likelihood multiple imputation (CLMI). Linear models between standardized log transformed urinary biomarkers and change in SBP and DBP were assessed after adjustment for age, baseline BP, sex, education, physical activity, smoking status, alcohol intake, 24-hour sodium excretion and GenSalt cohort. Results: Norepinephrine was positively associated with SBP in CLMI models (0.10 mmHg, CI: 0.00, 0.19) and DBP in LOD/√2 models (0.31 mmHg, CI: 0.05, 0.57) and CLMI models (0.10 mmHg, CI: 0.02, 0.18) during high sodium diet. Uromodulin was associated with decreased SBP in LOD/√2 models (-0.36 mmHg, CI: -0.57, -0.14) and CLMI models (-0.19 mmHg, CI: -0.31, -0.08) during potassium supplementation. Similarly, dopamine was positively associated with DBP during high sodium diet and negatively associated with DBP during potassium supplementation. Conclusion: These data indicate urinary biomarkers dopamine, norepinephrine and uromodulin are associated with BP salt sensitivity and may be used in risk stratification.

Role of sympathetic pathway in light-phase time-restricted feeding-induced blood pressure circadian rhythm alteration.

Disruption of blood pressure (BP) circadian rhythm, independent of hypertension, is emerging as an index for future target organ damage and is associated with a higher risk of cardiovascular events. Previous studies showed that changing food availability time alters BP rhythm in several mammalian species. However, the underlying mechanisms remain largely unknown. To address this, the current study specifically investigates (1) the relationship between rhythms of food intake and BP in wild-type mice; (2) effects of light-phase time-restricted feeding (TRF, food only available during light-phase) on BP circadian rhythm in wild-type and diabetic db/db mice; (3) the roles of the autonomic system and clock gene in light-phase TRF induced changes in BP circadian rhythm. Food intake and BP of C57BL/6J and db/db mice were simultaneously and continuously recorded using BioDAQ and telemetry systems under ad libitum or light-phase TRF. Per2 protein daily oscillation was recorded in vivo by IVIS spectrum in mPer2Luc mice. Autonomic nerve activity was evaluated by heart rate variability, baroreflex, urinary norepinephrine (NE) and epinephrine (Epi) excretion, and mRNA expressions of catecholamines biosynthetic and catabolic enzymes, and alpha-adrenergic receptors in mesenteric resistance arteries. We found that in wild-type mice, the BP level was correlated with the food intake temporally across the 24 h. Reversing the feeding time by imposing light-phase TRF resulted in reverse or inverted BP dipping. Interestingly, the net changes in food intake were correlated with the net alteration in BP temporally under light-phase TRF. In db/db mice, light-phase TRF worsened the existing non-dipping BP. The food intake and BP circadian rhythm changes were associated with alterations in Per2 protein daily oscillation and the time-of-day variations in heart rate variability, baroreflex, and urinary excretion of NE and Epi, and increased mRNA expression of Slc6a2 (encoding NE transporter) and Adra1d (encoding alpha-adrenergic receptor 1d) in the mesenteric resistance arteries, indicating the sympathetic nervous system (SNS) was modulated after light-phase TRF. Collectively, our results demonstrated that light-phase TRF results in reverse dipping of BP in wild-type and diabetic db/db mice and revealed the potential role of the sympathetic pathway in light-phase TRF-induced BP circadian rhythm alteration.

Peculiarities of hormonal regulation of the blood antioxidant system of adolescents with obesity

The study is designed to establish the relationship between the rate of the induced blood lipid peroxidation and production of some hormones in adolescents. The investigations have not shown any significant alteration in the blood insulin content as well as in the daily catecholamines and melatonin excretion in the urine of adolescents in early puberty with the I–II degree of neuroendocrine obesity and without insulin resistance. The rate of the induced blood lipid peroxidation in them remains to be at the initial level. In adolescents of the same age group hyperinsulinemia and an increased urinary excretion of catecholamines and melatonin occur against the background of the concomitant insulin resistance. The rate of the induced blood lipid peroxidation in such cases remains at the level of control values. Formation of hyperinsulinemia and an increased urinary excretion of norepinephrine and melatonin have been registered in late puberty in adolescents with neuroendocrine obesity of the I–II degree without insulin resistance. The emergence of these changes is accompanied by the maintenance of control values of the induced blood lipid peroxidation. The similar changes in the endocrine parameters with attendant increase in the rate of the induced blood lipid peroxidation are occurred in adolescents of the above-mentioned age group with neuroendocrine obesity against the background of insulin resistance.

LPIN1 is a new target gene for essential hypertension.

We previously showed Lipin1 (LPIN1) to be a candidate gene for essential hypertension by genome-wide association studies. LPIN1 encodes the Lipin 1 protein, which contributes to the maintenance of lipid metabolism and glucose homeostasis. However, little is known about the association between LPIN1 and blood pressure (BP). We evaluated the BP of LPIN1-deficient [fatty liver dystrophy (fld)] mice and explored related mechanisms. Fld mice have very low expression of LPIN1 and exhibit fatty liver, hypertriglyceridemia, insulin resistance and peripheral neuropathy. Fld mice had significantly elevated SBP and heart rate (HR) throughout the day as measured by a radiotelemetric method. Diurnal variation of SBP and HR was also absent in fld mice. Furthermore, urinary excretion of adrenaline and noradrenaline by fld mice was significantly higher compared with that of control mice. The BP response of fld mice to clonidine (a centrally acting α2-adrenergic receptor agonist) was greater than that of control mice. However, levels of Angiotensinogen and Renin 1 mRNA and urinary nitric oxide excretion were comparable between the two groups. The decrease in SBP at 8 weeks after fat grafting surgery was significantly greater in the transplant group compared with the sham operated group. The elevated BP in fld mice may result from activation of the sympathetic nervous system through decreased levels of adipose cytokines. These results indicate that LPIN1 plays a crucial role in blood pressure regulation and that LPIN1 is a new target gene for essential hypertension.

Overexpression of Growth Differentiation Factor 15 Reduces Blood Pressure and Produces Anti‐neuroinflammatory Effects in Mice with Diet‐induced Obesity

The growth differentiation factor 15 (GDF15) is a stress-responsive cytokine, which belongs to the super family of transforming growth factor beta (TGFb). In the past decade, GDF15 has been implicated in multiple biological processes and diseases, including anti-inflammation, anti-obesity and anti-diabetes effects. However, the role of GDF15 in sympathetic regulation of blood pressure has not been reported in the cardiovascular disease state. The purpose of the study was to investigate the effects of GDF15 on the blood pressure and central nervous system neuroinflammatory responses in diet-induced obese mice. NAG-Tg+ transgenic mice (TG) with overexpression of GDF15 and wild-type (WT) littermates were fed on normal chow or high-fat diet (HFD) for 16 weeks. During the experimental period, arterial pressure was recorded by radiotelemetry in conscious mice. WT mice with HFD had higher mean arterial pressure (MAP) compared with the normal diet fed WT mice (WT HFD 110±4 mmHg verse WT 93±4 mmHg). Compared with WT mice with HFD, GDF15 overexpression significantly reduced MAP (TG HFD: 99±3 mmHg verse WT HFD: 110±4 mmHg) accompanied by the decreased urinary norepinephrine excretion and baseline renal sympathetic nerve activity (TG HFD: 38.9±4.1% max verse WT HFD: 61.2±5.6% max). GDF15 TG mice with HFD had lower expressions of pro-inflammatory cytokines interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-a), interleukin 1β (IL-1β), C1q/TNF-related protein-1 (CTRP1) and reduced immune cell infiltration in the hypothalamus and brain stem tissue. A downregulation in GDF15 receptor (GFRAL) expression was detected in brain stem medullary nuclei in WT HFD mice, which was significantly upregulated in GDF15 TG mice with HFD. These data suggest that GDF15 overexpression exerts a potential protective effect on hypertension in obese mice through attenuating sympathetic outflow. The mechanism for this effect involves anti-inflammatory and neuronal protective properties of GDF15 in the central nervous system.

Hypertension due to loss of water

In the current issue of Acta Physiologica, two papers are published investigating the development of arterial hypertension under conditions of pathological loss of free water in two disease models. Kovaric et al 1 studied rats with chronic renal failure, shedding excessive free water via their remnant kidney due to a hypostenuric concentrating defect. Wild et al. 2 investigated a transgenic mouse model for psoriasis, losing free water via the psoriatic skin lesions. In both cases the animals displayed hypertension, cutaneous vasoconstriction, and metabolic adaptations towards enhanced production of urea and organic osmolytes typical for a phylogenetically ancient water conservation response termed aestivation 3 . The international consortia of investigators spearheaded by Titze, Karbach, and Kitada therefore conclude that in both cases the hypertension was due to cutaneous vasoconstriction in an attempt to limit epidermal water loss. Because in either case the hypertension was associated with loss of water, and thus with hypo- rather than hypervolemia, the authors argue, that this concept of pathogenesis of hypertension is antipodal to the classic view that relies on hypervolemia due to sodium retention.

Metastatic pheochromocytoma and paraganglioma: signs and symptoms related to catecholamine secretion

BackgroundThe presence or future development of metastatic pheochromocytomas or paragangliomas (mPPGLs) can be difficult to diagnose or predict at initial presentation. Since production of catecholamines from mPPGLs is different from non-metastatic tumors (non-mPPGLs), this study aimed to clarify whether presenting catecholamine-related signs and symptoms (cSS) might also differ.MethodsThe study included 249 patients, 43 with mPPGL and 206 with non-mPPGL. Clinical data at the time of biochemical diagnosis (i.e. at entry into the study) were used to generate a cumulative score of cSS for each patient.ResultsPatients with mPPGL were significantly younger (43.3 ± 14 vs. 48.9 ± 16.1 years) and included a lower proportion of females (39.5% vs. 60.7%) than patients with non-mPPGLs. Frequencies of signs and symptoms did not differ between the two groups. Patients with mPPGLs had lower (P < 0.001) urinary excretion of epinephrine (3.5 (IQR, 1.9—6.5) µg/day) than those with non-mPPGLs (19.1 (IQR, 4.3—70.2) µg/day). There was no difference in urinary excretion of norepinephrine. In patients with mPPGLs a high cSS score was associated with high urinary excretion of norepinephrine and normetanephrine. In contrast, in patients with non-mPPGLs, a high cSS was associated with high urinary excretion of epinephrine and metanephrine.ConclusionAlthough presenting signs and symptoms were associated with production of norepinephrine in patients with mPPGLs and of epinephrine in patients with non-mPPGLs, there were no differences in signs and symptoms between the two groups. Therefore, consideration of signs and symptoms does not appear helpful for distinguishing patients with and without mPPGLs.

Pathogenetic treatment of patients with hypertension depending on the level of sympathоadrenal system activity

Purpose of the study. Recently, new opportunities to optimize clinical efficacy and tolerability of treatment of patients with essential hypertension appeared. Determination of the indicators of increased activity of the sympathetic nervous system (SNS) in practice could contribute to differentiated treatment of patients with hypertension. Comparative estimation of influence of complex treatment with bisoprolol or amlodipine on the SNS activity parameters in patients with arterial hypertension (AH) of II stage.&#x0D; Materials and methods. 121 patients (men and women) with documented II stage AH were examined clinically. Concentrations of adrenaline, noradrenaline, dopamine and DOPA in urine were evaluated by spectrochromatography, endotheliumdependent vasodilation was studied using complex rheographic ReoCom Professional (HAI-Medicom, Ukraine). Patients were divided into 2 groups: if concentration of noradrenalin in urine exceeds healthy subjects levels more than on 20% – patients were added to basic treatment (enalapril, atorvastatin, aspirin) bisoprolol, if concentration of noradrenalin in urine exceeds healthy subjects levels less than on 20% – we added amlodipine. Statistic analysis was done on Apache OpenOffice (version 4.1) and PSPP (version 0.7.9).&#x0D; Results. The study showed that after treatment there was significant decrease of adrenalin, noradrenalin in 1st group . Increase of DOPA levels was higher in 1st group (Δ% = +46,5 [30,0; 55,4]% vs Δ% = +8,8 [–2,1 ; 16,0]%, (p &lt; 0,05).). Trends in daily urinary excretion of adrenaline (–44,8%) and norepinephrine (–33,7%) showed the impact of effective Δ-blocker on the activity of pressor systems. DOPA excretion was increased significantly in both groups which can indirectly indicate hemodynamic load reduction in hypertensive patients. When choosing the individual antihypertensive treatment in patients with arterial hypertension, if excretion of noradrenalin is increased it is rationally to add adjuvant therapy with β-blockers.&#x0D; Conclusion. The results of our research can be applied into the practice of general practitioner as recommendations for individual selection of antihypertensive treatment depending on the activity of sympathetic nervous system.

A food ingredient containing phytoecdysteroids and polyphenols from quinoa grain: technology and physiological and biochemical evaluation in vivo

An effective technological approach to produce adaptogenic microingredients is to concentrate plant biologically active substances via sorption on various matrices. The aim was to develop and evaluate in vivo the phytoecdysteroids and flavonoids concentrate extracted from quinoa grain and sorbed on the coagulated chicken egg protein. The consecutive extraction of phytoecdysteroids and flavonoids was conducted followed by sorption on the protein. The in vivo experiment was performed during 37 days using Wistar male rats. The developed concentrate was added into the diet of experimental animals in the dose 0.59±0.02 g/100 g of diet. As a stress model we used daily 40-min immobilization and exhaustive 3-hour immobilization at the end of the experiment. Urinary catecholamines content was determined, blood corticosterone, malon dialdehyde, glutathione reductase and superoxide dismutase content was determined. Phytoecdysteroids and flavonoids were concentrated 20 and 80 times respectively compared to the initial raw materials. The concentrate consumption neutralized negative effect of immobilization stress on anxiety level of Wistar rats. The significant decrease in urinary excretion of adrenaline and noradrenaline shows the pronounced adaptogenic effect of the concentrate, whereas phytoecdysteroids and flavonoids act as prostressors activating body antioxidant protection system, what is substantiated by significantly increased blood superoxide dismutase level of experimental animals.

Pathophysiological significance of Stim1 mutation in sympathetic response to stress and cardiovascular phenotypes in SHRSP/Izm: In vivo evaluation by creation of a novel gene knock-in rat using CRISPR/Cas9

ABSTRACT Genetic approach using rat congenic lines between SHRSP/Izm and WKY/Izm identified stromal interaction molecule 1 (Stim1), an essential component of store-operated Ca2+ entry (SOCE), as a promising candidate gene responsible for the exaggerated sympathetic response to stress in SHRSP. Since SHRSP has a nonsense mutation in Stim1 resulting in the expression of a truncated form of STIM1 that caused reduction of SOCE activity in primary cultured cerebral astrocytes, we created SHRSP/Izm knocked-in with the wild-type Stim1 (KI SHRSP) by the CRISPR/Cas9 method to investigate whether the functional recovery of STIM1 would mitigate sympatho-excitation to stress in vivo in SHRSP. No potential off-target nucleotide substitutions/deletions/insertions were found in KI SHRSP. Western blotting and fluorescent Ca2+ imaging of astrocytes confirmed wild-type STIM1 expression and restored SOCE activity in astrocytes from KI SHRSP, respectively. Blood pressure (BP) measured by the tail-cuff method at 12, 16, and 20 weeks of age did not significantly differ between SHRSP and KI SHRSP, while the heart rate of KI SHRSP at 16 and 20 weeks of age was significantly lower than that of age-matched SHRSP. Unexpectedly, the sympathetic response to stress (evaluated with urinary excretion of norepinephrine under cold stress and BP elevation under cold/restraint stress) did not significantly differ between SHRSP and KI SHRSP. The present results indicated that the functional deficit of STIM1 was not a genetic determinant of the exaggerated sympathetic response to stress in SHRSP and that it would be necessary to explore other candidates within the congenic fragment on chromosome 1.

Carbidopa for Afferent Baroreflex Failure in Familial Dysautonomia: A Double-Blind Randomized Crossover Clinical Trial.

Afferent lesions of the arterial baroreflex occur in familial dysautonomia. This leads to excessive blood pressure variability with falls and frequent surges that damage the organs. These hypertensive surges are the result of excess peripheral catecholamine release and have no adequate treatment. Carbidopa is a selective DOPA-decarboxylase inhibitor that suppresses catecholamines production outside the brain. To learn whether carbidopa can inhibit catecholamine-induced hypertensive surges in patients with severe afferent baroreflex failure, we conducted a double-blind randomized crossover trial in which patients with familial dysautonomia received high dose carbidopa (600 mg/day), low-dose carbidopa (300 mg/day), or matching placebo in 3 4-week treatment periods. Among the 22 patients enrolled (13 females/8 males), the median age was 26 (range, 12-59 years). At enrollment, patients had hypertensive peaks to 164/116 (range, 144/92 to 213/150 mm Hg). Twenty-four hour urinary norepinephrine excretion, a marker of peripheral catecholamine release, was significantly suppressed on both high dose and low dose carbidopa, compared with placebo (P=0.0075). The 2 co-primary end points of the trial were met. The SD of systolic BP variability was reduced at both carbidopa doses (low dose: 17±4; high dose: 18±5 mm Hg) compared with placebo (23±7 mm Hg; P=0.0013), and there was a significant reduction in the systolic BP peaks on active treatment (P=0.0015). High- and low-dose carbidopa were similarly effective and well tolerated. This study provides class Ib evidence that carbidopa can reduce blood pressure variability in patients with congenital afferent baroreflex failure. Similar beneficial effects are observed in patients with acquired baroreflex lesions.

Neuronal Nitric Oxide Synthase Associated Protein: Nos1ap mediates Sympathoexcitation through Paraventricular Nucleus of the Hypothalamus

Exaggerated sympathoexcitation is associated with multiple disorders, including hypertension, heart failure, diabetes leading to various cardiovascular complications. Within the paraventricular nucleus of the hypothalamus (PVN), nitric oxide (NO•) is synthesized by neuronal nitric oxide synthase (NOS1) and acts as a non‐conventional neurotransmitter that inhibits neuronal firing and consequent sympathetic outflow. NOS1 is tethered to N‐methyl D‐aspartate receptor (NMDAR) channels harboring NMDAR subunit 2B by the scaffolding protein postsynaptic density 95 (PSD95). Activation of the NMDAR at the synaptic membrane initiates a Ca2+ influx that stimulates the production of NO•. NOS1 adaptor protein (Nos1ap) competes with PSD95 for NOS1 binding and thus influences the production of NO•. We investigated the effect of over‐expressing Nos1ap in the PVN on sympathetic outflow using Cre recombinase‐conditional Nos1ap‐transgenic (Tg) mice. The Nos1ap Tg mice were microinjected with lentiviral expressing Cre‐GFP or GFP bilaterally into the PVN following our standardized method, referred to as CAPON‐OE and Control, respectively. Two weeks of post‐viral administration, the sympathetic outflow, was evaluated. CAPON‐OE mice displayed a significantly elevated urinary excretion of norepinephrine, an index of overall sympathetic outflow compared to Control (0.64±0.11* vs. 0.17±0.03 ug/day). Basal renal sympathetic nerve activity, measured as a percent of maximal activity was elevated 1.5‐fold in CAPON‐OE (46.09 ± 3.56* vs.31.04 ± 3.24) mice. Furthermore, FosB (a member of the AP‐1 family of transcription regulatory proteins) immunocytochemistry revealed increased expression in the PVN of CAPON‐OE mice when compared to control mice suggesting a chronic activation of PVN neurons after Nos1ap over‐expression. Interestingly, a robust over‐expression of Nos1ap in the PVN of CAPON‐OE (0.49±0.04* vs. 0.21±0.03) compared to control mice does not lead to any significant change in NOS1 expression. However, DAF‐FM staining of PVN slices showed decreased NO• levels in CAPON‐OE mice when compared to control mice suggesting that the decrease in NMDAR‐mediated NOS1 activity due to sequestering of NOS1 by Nos1ap leads to less NO• production. Taken together, our data indicate that specific over‐expression of Nos1ap within the PVN reduces the levels of NO• generated by NOS1, leading to increased sympathetic outflow.Support or Funding InformationSupported by AHA ‐ 14SDG19980007 and NIH grant HL62222.

Urinary Norepinephrine Is a Metabolic Determinant of 24-Hour Energy Expenditure and Sleeping Metabolic Rate in Adult Humans.

Interindividual variability in 24-hour energy expenditure (24EE) during energy-balance conditions is mainly determined by differences in body composition and demographic factors. Previous studies suggested that 24EE might also be influenced by sympathetic nervous system activity via catecholamine (norepinephrine, epinephrine) secretion. Therefore, we analyzed the association between catecholamines and energy expenditure in 202 individuals from a heterogeneous population of mixed ethnicities. Participants (n = 202, 33% female, 14% black, 32% white, 41% Native American, 11% Hispanic, age: 36.9 ± 10.3 y [mean ± SD], percentage body fat: 30.3 ± 9.4) resided in a whole-room calorimeter over 24 hours during carefully controlled energy-balance conditions to measure 24EE and its components: sleeping metabolic rate (SMR), awake-fed thermogenesis (AFT), and spontaneous physical activity (SPA). Urine samples were collected, and 24-h urinary epinephrine and norepinephrine excretion rates were assessed by high-performance liquid chromatography. Both catecholamines were associated with 24EE and SMR (norepinephrine: +27 and +19 kcal/d per 10 μg/24h; epinephrine: +18 and +10 kcal/d per 1 μg/24h) in separate analyses after adjustment for age, sex, ethnicity, fat mass, fat-free mass, calorimeter room, temperature, and physical activity. In a multivariate model including both norepinephrine and epinephrine, only norepinephrine was independently associated with both 24EE and SMR (both P < .008), whereas epinephrine became insignificant. Neither epinephrine nor norepinephrine were associated with adjusted AFT (both P = .37) but epinephrine was associated with adjusted SPA (+0.5% per 1 μg/24h). Our data provide compelling evidence that sympathetic nervous system activity, mediated via norepinephrine, is a determinant of human energy expenditure during nonstressed, eucaloric conditions.

Exercise training upregulates Nrf2 protein in the rostral ventrolateral medulla of mice with heart failure.

Chronic heart failure (CHF) is associated with global oxidative stress, which contributes to sympathoexcitation. Increased reactive oxygen species in the brain accumulate within neurons and lead to enhanced neuronal excitability. Exercise training (ExT) is associated with a reduction of oxidative stress by upregulation of antioxidant enzymes. The link between ExT and antioxidant enzyme expression in the brain of animals with CHF is not clear. We hypothesized that ExT enhances transcription and translation of the nuclear factor erythroid 2-related factor 2 (Nrf2) gene, a master transcription factor that modulates antioxidant enzyme gene expression, in the rostral ventrolateral medulla (RVLM) of mice with CHF. Mice were divided into the following groups: Sham sedentary (Sham-Sed), Sham-ExT, CHF-Sed, and CHF-ExT. After 8 wk of ExT, we measured Nrf2 and NAD(P)H dehydrogenase [quinone] 1 (NQO-1) message and protein expression along with maximal exercise tolerance and urinary norepinephrine (NE) excretion. We found that Nrf2 and NQO-1 mRNA and protein expression in the RVLM were lower in CHF-Sed mice compared with Sham-Sed. ExT attenuated the CHF-induced reduction of Nrf2 and NQO-1 mRNA and protein expression in the RVLM. NE excretion was higher in CHF-Sed mice compared with Sham-Sed (666.8 ± 79.3 ng/24 h, n = 6 vs. 397.8 ± 43.7 ng/24 h, P = 0.04). CHF-ExT mice exhibited reduced urinary NE excretion compared with CHF-Sed (360.7 ± 41.7 ng, n = 4 vs. 666.8 ± 79.3 ng, n = 6; P = 0.03). We conclude that ExT-induced upregulation of Nrf2 in the RVLM contributes to the beneficial effects of ExT on sympathetic function in the heart failure state.NEW & NOTEWORTHY This study provide evidence for an important role for exercise training in the modulation of antioxidant enzyme production in the rostral ventrolateral medulla (RVLM) in the heart failure state. We show here a correlation between exercise training and the expression of the antioxidant transcription factor Nrf2 in the RVLM. Exercise training reduced sympathetic function (norepinephrine excretion) and upregulated both Nrf2 and the antioxidant enzyme NQO-1. We conclude that Nrf2 in the RVLM may be an important target for controlling sympathetic outflow in heart failure.

Central Glucagon-like Peptide-1 Receptor Signaling via Brainstem Catecholamine Neurons Counteracts Hypertension in Spontaneously Hypertensive Rats

Glucagon-like peptide-1 receptor (GLP-1R) agonists, widely used to treat type 2 diabetes, reduce blood pressure (BP) in hypertensive patients. Whether this action involves central mechanisms is unknown. We here report that repeated lateral ventricular (LV) injection of GLP-1R agonist, liraglutide, once daily for 15 days counteracted the development of hypertension in spontaneously hypertensive rats (SHR). In parallel, it suppressed urinary norepinephrine excretion, and induced c-Fos expressions in the area postrema (AP) and nucleus tractus solitarius (NTS) of brainstem including the NTS neurons immunoreactive to dopamine beta-hydroxylase (DBH). Acute administration of liraglutide into fourth ventricle, the area with easy access to the AP and NTS, transiently decreased BP in SHR and this effect was attenuated after lesion of NTS DBH neurons with anti-DBH conjugated to saporin (anti-DBH-SAP). In anti-DBH-SAP injected SHR, the antihypertensive effect of repeated LV injection of liraglutide for 14 days was also attenuated. These findings demonstrate that the central GLP-1R signaling via NTS DBH neurons counteracts the development of hypertension in SHR, accompanied by attenuated sympathetic nerve activity.

Abstract 088: Overexpression of Neuronal Nitric Oxide Synthase Associated Protein in the Paraventricular Nucleus Contributes to Increased Sympathoexcitation in Male Mice by Modulating Subcellular Localization of Neuronal Nitric Oxide Synthase

Nitric oxide (NO•) bioavailability within the paraventricular nucleus (PVN) plays a pivotal role in the pathogenesis of many cardiovascular diseases including hypertension. Aligning neuronal nitric oxide synthase (NOS1) with N -methyl-D-aspartate (NMDA) receptor by tethering to PSD95 (postsynaptic density 95 protein) allows the production of NO• initiated by an influx of Ca 2+ in neurons within the PVN. NOS1 associated protein (Nos1ap) competes with PSD95 for NOS1 binding and thus influences the production of NO•. We investigated the effect of specifically overexpressing Nos1ap in the PVN on sympathetic outflow using cre recombinase-conditional Nos1ap over-expression transgenic(Tg) mice injected with lentivirus expressing Cre-GFP or GFP into the PVN. Two weeks of post-viral administration, the sympathetic outflow was evaluated in both male and female mice. Male-Cre-Nos1ap mice displayed a significantly elevated urinary excretion of norepinephrine, an index of overall sympathetic outflow compared to GFP-Nos1ap (0.64±0.11* vs. 0.17±0.03 ug/day) while female littermates showed no difference (0.44±0.06 vs. 0.36±0.03 ug/day). Basal renal sympathetic nerve activity assessed as a percent of maximal activity was elevated 1.5-fold in Cre-Nos1ap male mice (46.1 ± 3.6* vs.31.0 ± 3.2) while there was no significant difference in Cre-Nos1ap female mice(22.6± 1.2 vs. 24.8 ± 2.6) compared to respective GFP controls. Interestingly, female mice had higher levels of basal NOS1 in the PVN compared to male littermates (0.79 ±0.01* vs. 0.37±0.05). Further, a robust overexpression of Nos1ap in the PVN of Cre-Nos1ap (0.49±0.04* vs. 0.21±0.03 male; 0.53±0.08* vs. 0.28±0.04 female) compared to GFP-Nos1ap mice does not lead to any change in NOS1, NMDA receptor(NR)2A, NR2B, while NR1 expression tended to increase in both sexes. Overexpression of Nos1ap in NG108 neurons, in vitro, led to relocalization of 46% of membrane-associated NOS1 to the cytosolic compartment, suggesting that the decrease in NMDA receptor-mediated NOS1 activity leads to reduced NO• production. Taken together, our data indicate that overexpression of Nos1ap within the PVN reduces the levels of NO• generated by NOS1 leading to increased sympathetic outflow in male mice but not in female mice.

Association Between Urinary Catecholamine Excretion and Urine Volume.

Several confounders must be considered in the evaluation of urinary catecholamine excretion. However, literature is contradictory about potential confounders. The aim of the present study was to assess correlations between catecholamine excretion and anthropometric or clinical parameters with special attention to urine volume. A total of 967 24-h urinary catecholamine measurements were performed in 593 patients for diagnostic purposes. The indication for urine examination was suspicion of secondary hypertension, phaeochromocytoma, or paraganglioma. From the patients examined, 57% were females and 43% were males. The patients' age ranged between 15 and 87 years with a median [Q1; Q3] of 51 [39; 62] years. Seventy-eight percent of the patients suffered from hypertension. Seventy percent of patients took one or more antihypertensive drugs. The most commonly used drugs were ACE inhibitors (43%), while α-blockers (15%) were the least used drugs. Urinary excretion was between 500 and 11 950 ml/24 h with a median of 2200 [1600; 2685] ml/24 h. The median body mass index (BMI) was 26.7 [24.0; 30.4] kg/m2. The excretion of all catecholamines was greater in men than in women (all p<0.0001). Epinephrine (p=0.0026), dopamine (p<0.0001), and metanephrine (p=0.0106) excretion decreased with age. BMI was associated with urinary excretion of dopamine (p<0.0001), norepinephrine (p=0.0026), normetanephrine (p<0.0001), and homovanillylmandelic acid (HVMA; p=0.0251). Urine volume correlated with urinary dopamine (p=0.0127), metanephrine (p<0.0001), normetanephrine (p=0.0070), and HVMA (p<0.0028) excretion. In addition to the established associations between urinary catecholamine excretion and age, gender, and BMI in the present study, urinary catecholamine excretion correlated also with urine volume.

Modulation of Sympathetic Activity and Innervation With Chronic Ivabradine and β-Blocker Therapies: Analysis of Hypertensive Rats With Heart Failure

Whether the reduction of heart rate with ivabradine (IVA) could affect sympathetic activation and cardiac innervation in heart failure (HF) remains unknown. The present study assessed the chronic effects of IVA and β-blocker on the systemic and local sympathetic nervous systems of hypertensive animals with HF. The Dahl salt-sensitive rats received chronic IVA, bisoprolol (BIS), or placebo (CTL) therapy. The survival of the animal models with IVA and BIS significantly improved (median; 19.7 in IVA and 19.7 in BIS vs 17.0 weeks in CTL, P < .001). A similar decrease in 24-hour heart rate (mean; 305 in IVA and 329 in BIS vs 388 beats/min in CTL, P < .001) without effect on blood pressure, and an improvement in the left ventricular dysfunction (mean fractional shortening; 56.7% in IVA and 47.8% in BIS vs 39.0% in CTL, P < .001) were observed in the animals with IVA and BIS. However, a negative inotropic effect was only observed in the animals with BIS. Excessive urinary noradrenaline excretion in animals with CTL was only suppressed with the use of IVA (mean; 1.35 μg/d in IVA and 1.95 μg/d in BIS vs 2.27 μg/d in CTL, P = .002). In contrast, atrial noradrenaline and acetylcholine depletion in the animals with CTL improved and the tyrosine hydroxylase expression in the both atria were restored with the use of both IVA and BIS. IVA therapy improved the survival of hypertensive animals with HF. Furthermore, it was associated with the amelioration of systemic sympathetic activation and cardiac sympathetic and parasympathetic nerve innervations. Chronic β-blocker therapy with negative inotropic effects had beneficial effects only on cardiac innervations.

A long-term perspective on cardiovascular job stress research.

This review provides perspectives on cardiovascular occupational stress research since the 1960s until now. The author argues for closer links between basic science and clinical follow‐up examinations of patients. In an excellent way urinary excretion of adrenaline and noradrenaline during wake hours mirrors day to day or week to week variations in sympathomedullary activity which could be related to variations in the patient's and cardiovascular and psychosocial situation. Modern methods for following variations over time in heart contractility should also be related to the patients' psychosocial situation. In addition the author argues for more extensive use of the increasing knowledge regarding regeneration and vagal activity in relation to variations in job conditions and development or prevention of cardiovascular disease.

Abstract 083: Role of Alpha2a-Adrenoceptors in the Development of Hypertensive Nephropathy

Objective and Methods: Hypertensive nephropathy is associated by increased renal sympathetic nerve activity. Alpha2A-adrenoceptors (alpha2A-AR) regulate sympathetic tone by controlling norepinephrine (NE) release by a negative feedback mechanism. Recently, we have shown that deletion of alpha2A-adrenoceptors increases renal sympathetic neurotransmission. To investigate whether deletion of alpha2A-AR play a role in the development of hypertensive nephropathy, we induced hypertension by infusing angiotensin (Ang)II (1000ng/kg/min) for 4 weeks in uninephrectomized wild-type (WT) and alpha2A-AR knockout (alpha2A-AR-/-) mice on an FVB background. Blood pressure was assessed by radiotelemetry. Results: NE release stimulated by renal nerve stimulation 5Hz was significantly higher in untreated kidneys of alpha2A-AR-/- compared controls. During AngII infusion (1000ng/kg/min) urinary NE excretion was significantly higher in alpha2A-AR-/- than control mice (4.3±0.6 vs. 2.1±0.4mg/dl creatinine, P &lt;0.05). In line with an increased sympathetic NE release, the severity of hypertension caused by AngII infusion was significantly higher in alpha2A-AR-/- mice compared to control mice (176±4 vs. 163±3mmHg, P &lt;0.01). Moreover, albuminuria (140±24 vs. 66±19mg/mg creatinine, P &lt;0.05) as well as kidney injury assessed by glomerular injury score and renal fibrosis were accelerated and kidney function measured by cystatin C (1082±66 vs. 906±52ng/ml, P&lt;0.05) was significantly decreased in alpha2A-AR-/- compared to WT mice. To test whether reduced renal sympathetic activity reversed this effect, renal nerve denervation (RDN) was performed in alph2A-AR-/- mice. RDN reduced renal NE content by 70%. In alpha2A-AR-/- mice, RDN reduced the severity of AngII induced hypertension 138±9 vs. 169± 11mmHg, P&lt;0.05) and albuminuria compared to non-denervated alpha2A-AR-/- mice. Conclusion: Deletion of alpha2A-ARs exaggerates hypertensive nephropathy by increasing sympathetic NE activity. Reduction of NE release by RDN reduced blood pressure and albuminuria in alpha2A-AR-/- mice suggesting an important role of renal sympathetic tone in the pathogenesis of hypertensive nephropathy.