A polymorphism in the norepinephrine transporter gene is associated with affective and cardiovascular disease through a microRNA mechanism.

Norepinephrine released from sympathetic nerves is removed from the neuroeffector junction via the action of the norepinephrine transporter (NET). NET impairment is evident in several clinically important conditions including major depressive disorder (MDD), panic disorder (PD), essential hypertension and the postural orthostatic tachycardia syndrome (POTS). We aimed to determine whether a single nucleotide polymorphism (SNP) in the 3' untranslated region (UTR) of the NET gene is associated with NET impairment and to elucidate the mechanisms involved. The analyses were carried out in two cohorts of European ancestry, which included healthy controls and MDD, PD, hypertensive and POTS patients. Compared with controls, cases had significantly higher prevalence of the T allele of rs7194256 (C/T), arterial norepinephrine, depression and anxiety scores, larger left ventricular mass index, higher systolic and diastolic blood pressures, and heart rate. Bioinformatic analysis identified that the microRNA miR-19a-3p could bind preferentially to the sequence created by the presence of the T allele. This was supported by results of luciferase assays. Compared with controls, cases had significantly lower circulating miR-19a-3p, which was associated with pathways related to blood pressure and regulation of neurotransmission. In vitro norepinephrine downregulated miR-19a-3p. In conclusion, the T allele of the rs7194256 SNP in the 3'UTR of the NET gene is more prevalent in diseases where NET impairment is evident. This might be explained by the creation of a binding site for the microRNA miR-19a-3p. A defect in NET function may potentiate the sympathetic neurochemical signal, predisposing individuals with affective diseases to increased risk of cardiovascular disease development.

Soluble vascular endothelial growth factor receptor-1 is reduced in patients with resistant hypertension after renal denervation.

Renal denervation (RDN) has been shown in several studies to reduce blood pressure (BP) in patients with resistant hypertension (RH). Data on potential biomarkers associated with BP changes remain scarce. We evaluated whether soluble vascular endothelial growth factor receptor (sVEGFR-1) is affected by the procedure. A total of 57 patients with RH participated in this study. BP and heart rate were recorded at baseline and at 3 months follow-up, at which time blood samples were collected to determine the levels of sVEGFR-1, VEGF-A, VEGF-C, nitric oxide (NO), soluble vascular adhesion molecule 1 and soluble intracellular adhesion molecule 1. None of the biomarkers had a predictive value that could identify responders vs non-responders to RDN. However, sVEGFR-1 concentration was dramatically reduced after RDN (5913±385 vs 280±57 pg ml-1, P<0.001). At the same time VEGF-A levels were significantly increased (10.0±3.0 vs 55.5±7.9 pg ml-1, P<0.001), without significant changes in VEGF-C. NO levels were significantly increased after RDN in the whole group (82.6±6.2 vs 106.9±7.8 µM, P=0.021). Interestingly, the elevation in NO levels at 3 months was only seen in patients who demonstrated a reduction in systolic BP of ⩾10 mm Hg (78.9±8.3 vs 111.6±11.7 µM, P=0.018). We report a significant reduction in sVEGFR-1 levels after RDN procedure, which was accompanied by a significant increase in VEGF-A concentration as well as NO. Changes in plasma cytokines were not quantitatively linked to magnitude of BP reduction. An RDN-induced reduction in sVEGFR-1 plasma levels and increase in VEGF-A would raise the VEGF-A/sVEGFR-1 ratio, thereby increasing VEGF-A bioavailability to act on its full-length receptor and may contribute to the BP-lowering effect potentially via NO-mediated pathways.

Norepinephrine transporter expression is inversely associated with glycaemic indices: a pilot study in metabolically diverse persons with overweight and obesity.

OBJECTIVE: The objective of this study was to examine the cross-sectional relationship between the expression of norepinephrine transporter (NET), the protein responsible for neuronal uptake-1, and indices of glycaemia and hyperinsulinaemia, in overweight and obese individuals. METHODS: Thirteen non-medicated, non-smoking subjects, aged 58 ± 1 years (mean ± standard error of the mean), body mass index (BMI) 31.4 ± 1.0 kg m-2, with wide-ranging plasma glucose and haemoglobin A1c (HbA1c, range 5.1% to 6.5%) participated. They underwent forearm vein biopsy to access sympathetic nerves for the quantification of NET by Western blot, oral glucose tolerance test (OGTT), euglycaemic hyperinsulinaemic clamp, echocardiography and assessments of whole-body norepinephrine kinetics and muscle sympathetic nerve activity. RESULTS: Norepinephrine transporter expression was inversely associated with fasting plasma glucose (r = -0.62, P = 0.02), glucose area under the curve during OGTT (AUC0-120, r = -0.65, P = 0.02) and HbA1c (r = -0.67, P = 0.01), and positively associated with steady-state glucose utilization during euglycaemic clamp (r = 0.58, P = 0.04). Moreover, NET expression was inversely related to left ventricular posterior wall dimensions (r = -0.64, P = 0.02) and heart rate (r = -0.55, P = 0.05). Indices of hyperinsulinaemia were not associated with NET expression. In stepwise linear regression analysis adjusted for age, body mass index and blood pressure, HbA1c was an independent inverse predictor of NET expression, explaining 45% of its variance. CONCLUSIONS: Hyperglycaemia is associated with reduced peripheral NET expression. Further studies are required to identify the direction of causality.

Ephedrine activates brown adipose tissue in lean but not obese humans.

AIMS/HYPOTHESIS: Brown adipose tissue (BAT) activation increases energy consumption and may help in the treatment of obesity. Cold exposure is the main physiological stimulus for BAT thermogenesis and the sympathetic nervous system, which innervates BAT, is essential in this process. However, cold-induced BAT activation is impaired in obese humans. To explore the therapeutic potential of BAT, it is essential to determine whether pharmacological agents can activate BAT. METHODS: We aimed to determine whether BAT can be activated in lean and obese humans after acute administration of an orally bioavailable sympathomimetic. In a randomised, double-blinded, crossover trial, we administered 2.5 mg/kg of oral ephedrine to nine lean (BMI 22 ± 1 kg/m²) and nine obese (BMI 36 ± 1 kg/m²) young men. On a separate day, a placebo was administered to the same participants. BAT activity was assessed by measuring glucose uptake with [¹8F]fluorodeoxyglucose and positron emission tomography-computed tomography imaging. RESULTS: BAT activity was increased by ephedrine compared with placebo in the lean, but unchanged in the obese, participants. The change in BAT activity after ephedrine compared with placebo was negatively correlated with various indices of body fatness. CONCLUSIONS/INTERPRETATION: BAT can be activated via acute, oral administration of the sympathomimetic ephedrine in lean, but not in obese humans.

The morning blood pressure surge is related to serum cholesterol.

A morning blood pressure surge (MBPS) may be either a mechanism for, or a marker of, increased cardiovascular events. This study has examined factors which may influence the morning surge: age, gender, metabolic factors, sympathetic function, blood pressure and arterial stiffness. Four measures of the MBPS were examined--sleep-trough surge, pre-awake surge, rate of blood pressure rise and a Power function. Subjects underwent ambulatory blood pressure monitoring, glucose tolerance test, central pulse wave velocity, sympathetic autonomic function tests (mental stress and sustained handgrip). MBPS was associated with age, hypertension, blood pressure variability and serum lipids. After adjustment for age and waist circumference, all four measures of MBPS remained positively associated with low-density lipoprotein (LDL) cholesterol. The novel finding of a significant relationship between measures of MBPS and LDL-cholesterol is an intriguing link between two major cardiovascular risk factors.

The effects of dietary weight loss with or without exercise training on liver enzymes in obese metabolic syndrome subjects.

AIM: Insulin resistance and visceral adiposity are predisposing factors for fatty liver disease. The main objectives of this study were (i) to compare the effects of caloric restriction (CR) alone or together with moderate-intensity aerobic exercise training (CR+EX) on liver enzymes, a surrogate marker of liver injury, in obese metabolic syndrome (MetS) subjects and (ii) to identify anthropometric, metabolic, cardiovascular and dietary predictors of changes in liver enzymes. METHODS: Sedentary men and women (n = 63), aged 55 ± 6 (s.d.) years with body mass index 32.7 ± 4.1 kg/m(2) and confirmed MetS, were randomized to 12-week CR, CR+EX or no treatment (Control). RESULTS: Weight loss averaged 7.6% in the CR and 9.1% in the CR+EX group (time effect, p < 0.001; group effect, p = 0.11); insulin sensitivity improved by 49 and 45%, respectively (both p < 0.001). Fitness (maximal oxygen consumption) increased by 19% in the CR+EX group only (p < 0.001). Alanine aminotransferase (ALT) levels decreased by 20% in the CR and 24% in the CR+EX group (time effect, both p < 0.001; group effect, p = 0.68); corresponding values for γ-glutamyltransferase (GGT) were -28 and -33%, respectively (time effect, both p < 0.001; group effect, p = 0.28). Reduction in abdominal fat mass (measured by DXA from L1 to L4) independently predicted ΔALT (r = 0.42, p = 0.005) and ΔGGT (r = 0.55, p < 0.001), whereas change in dietary saturated fat intake was independently associated with ΔALT (r = 0.35, p = 0.03). CONCLUSIONS: Reductions in central adiposity and saturated fat intake are key drivers of improvement in liver enzymes during lifestyle interventions. Exercise training did not confer significant incremental benefits in this study.

Brain leptin resistance in human obesity revisited.

Leptin is a 16 kDa peptide predominantly produced by adipocytes. Leptin and its receptor are known to be involved in the regulation of energy balance. The data from animal studies as well as our own observations of leptin overflow from the brain suggest that the central nervous system is a site of leptin synthesis. Using simultaneous arterio-venous blood sampling we here confirm that leptin is released from the brain into the internal jugular vein, and that release is greater in overweight men and in females compared to lean men, 467.3 ng/min+/-160.4 and 1426 ng/min+/-769.3 vs 80.0 ng/min+/-29.3, respectively (P<0.05). Furthermore, we have examined the gene expression of leptin and its receptor isoforms by reverse transcription-polymerase chain reaction (RT-PCR) in human cadaver hypothalami across a broad range of adiposity. Leptin gene expression was detected in a number of donors; the presence of detectable leptin mRNA was related to the mode of death rather than BMI or gender. We have also demonstrated gene expression of the three leptin receptor isoforms in the human hypothalamus. No relation was observed between the levels of hypothalamic expression of the long signaling form of the leptin receptor and BMI. In summary, this study indicates that it is very difficult to explain human obesity on the basis of central nervous system "leptin resistance", in that leptin is released in the brain, and at a higher level in the obese, and brain leptin receptor gene expression is not impaired in obesity.

Estrogen increases prepulse inhibition of acoustic startle in rats.

Epidemiological studies have shown gender differences in the age of onset and symptoms of schizophrenia. Because sensorimotor gating mechanisms are deficient in schizophrenia, we studied the effect of administration of estrogen on prepulse inhibition of startle in rats, an animal model of sensorimotor gating. Rats were tested in an automated startle apparatus for their responses to random combinations of 115-dB sound pulses and prepulses of various intensity. Startle responses were reduced by increasing intensities of prepulses, indicating prepulse inhibition. Repeated administration of startle pulses caused gradual habituation of startle responses. Ovariectomy did not induce significant changes in either habituation of the startle response or prepulse inhibition of startle. Treatment with 17beta-estradiol caused an increase in percentage prepulse inhibition at all prepulse intensities at 18 h, but only at higher prepulse intensities at 30 min after injection. Habituation of startle responses was not affected. The enhancing effect of estradiol on prepulse inhibition was mimicked by testosterone, but not by dihydrotestosterone. Estradiol treatment increased prepulse inhibition similarly in controls or after disruption of prepulse inhibition induced by treatment with apomorphine or dizocilpine (MK-801). Our results may help to explain gender differences in schizophrenia and some of the beneficial clinical effects of estrogen treatment in this disease.

Cardiovascular and behavioural responses to psychological stress in spontaneously hypertensive rats: effect of treatment with DSP-4.

We used a model of psychological stress combining exposure to an open-field novel environment, radio-telemetric measurement of blood pressure and heart rate, and behavioural tracking analysis of behavioural parameters. All rats showed significant increases in blood pressure and heart rate for the duration of open-field exposure, with spontaneously hypertensive rats (SHR) showing markedly greater pressor responses and tachycardia when compared to either Wistar-Kyoto (WKY) or Sprague-Dawley rats (SD rats). Behavioural responses in the open-field were unrelated to the magnitude of cardiovascular responses. Open-field exposure on 4 consecutive days induced similar pressor responses and tachycardia on each day. By contrast, behavioural responses were reduced from the second day of open-field exposure. Treatment of SHR and WKY rats with DSP-4, to deplete central noradrenaline levels, did not affect cardiovascular responses in SHR, whereas WKY rats showed a trend towards inhibition. However, in WKY rats, but not SHR, DSP-4 treatment caused a marked reduction in behavioural activity in the open-field. In conclusion, these data show that: (1) SHR display marked cardiovascular hyperreactivity to psychological open-field stress when compared to two normotensive rat strains; (2) unlike behavioural responses, cardiovascular stress responses do not habituate upon repeated stress exposure; and (3) noradrenergic projections from the locus coeruleus do not appear to play a major role in cardiovascular stress responses in SHR or WKY rats, although they may be involved in behavioural responses in WKY rats.

Cardiovascular responses to open-field stress in rats: sex differences and effects of gonadal hormones.

We studied sex differences in cardiovascular responses to stress using a new radio-telemetry model in which freely-moving Spontaneously Hypertensive rats (SHR) are exposed to open-field novelty stress. This model allowed simultaneous assessment of cardiovascular and behavioural responses to psychological stress. Female SHR in the diestrous stage of their estrous cycle had markedly greater pressor and tachycardic responses to open-field exposure when compared to either female rats not in diestrous or male SHR. Treatment of ovariectomized SHR with estrogen alone had no significant effect on cardiovascular reactivity, while a combined treatment of estrogen and progesterone slightly, but significantly attenuated their pressor response to open-field stress. In addition, treatment of castrated male rats with testosterone significantly enhanced their pressor responses to stress when compared to values obtained before treatment. None of the hormone treatments had any significant effect on heart rate responses to stress. Neither at different stages of the estrous cycle nor after hormone treatments were there any marked changes in behavioural responses in the open-field, making it unlikely that the differences in cardiovascular stress responses were caused by changes in behavioural activity. These data demonstrate differences in cardiovascular stress responses that seem to be dependent on the stage of the estrous cycle. They suggest that particularly androgens, such as testosterone, may enhance pressor responses to stress. On the other hand, a combination of estrogen and progesterone, rather than estrogen alone, may have a small attenuating effect on cardiovascular reactivity.