Acute Stress Induces Contrasting Changes in AMPA Receptor Subunit Phosphorylation within the Prefrontal Cortex, Amygdala and Hippocampus

Exposure to stress causes differential neural modifications in various limbic regions, namely the prefrontal cortex, hippocampus and amygdala. We investigated whether α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) phosphorylation is involved with these stress effects. Using an acute inescapable stress protocol with rats, we found opposite effects on AMPA receptor phosphorylation in the medial prefrontal cortex (mPFC) and dorsal hippocampus (DH) compared to the amygdala and ventral hippocampus (VH). After stress, the phosphorylation of Ser831-GluA1 was markedly decreased in the mPFC and DH, whereas the phosphorylation of Ser845-GluA1 was increased in the amygdala and VH. Stress also modulated the GluA2 subunit with a decrease in the phosphorylation of both Tyr876-GluA2 and Ser880-GluA2 residues in the amygdala, and an increase in the phosphorylation of Ser880-GluA2 in the mPFC. These results demonstrate that exposure to acute stress causes subunit-specific and region-specific changes in glutamatergic transmission, which likely lead to the reduced synaptic efficacy in the mPFC and DH and augmented activity in the amygdala and VH. In addition, these findings suggest that modifications of glutamate receptor phosphorylation could mediate the disruptive effects of stress on cognition. They also provide a means to reconcile the contrasting effects that stress has on synaptic plasticity in these regions. Taken together, the results provide support for a brain region-oriented approach to therapeutics

Randomised, Double-Blind, Placebo-Controlled, Dose-Escalating Phase I Study of QGC001, a Centrally Acting Aminopeptidase A Inhibitor Prodrug

International audienceBackground and Objectives Inhibition of brain aminopeptidase A (APA), which converts angiotensin II into angiotensin III, has emerged as a novel antihypertensive treatment, as demonstrated in several experimental animal models. QGC001 (originally named RB150) is a prodrug of the specific and selective APA inhibitor EC33, and as such it is the prototype of a new class of centrally acting antihypertensive agents. Given by the oral route in hypertensive rats, it enters the brain and generates EC33, which blocks the brain renin-angiotensin system activity and normalises blood pressure. The aim of the present study was to evaluate the safety, pharmacokinetics and pharmacodynamic effects of QGC001 in humans. Design and Methods Fifty-six healthy male volunteers were randomly assigned to receive in double-blind and fasted conditions single oral doses of 10, 50, 125, 250, 500, 750, 1,000 and 1,250 mg of QGC001 (n = 6/dose) or placebo (n = 2/dose). We measured plasma and urine concentrations of both QGC001 and EC33 by liquid chromatography-tandem mass spectrometry, plasma renin concentrations (PRC), plasma and free urine aldosterone (PAldo and UAldo), plasma copeptine (PCop), and plasma and urine cortisol (PCort and UCort) concentrations, and supine systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR) at various time points. Results All doses of QGC001 were clinically and biologically well-tolerated. Peak plasma concentrations (C-max) of QGC001 and EC33 increased linearly with the dose, with a median time to reach C-max (t(max)) of 1.5 h for QGC001 and 3.0 h for EC33. The median plasma elimination half-life of QGC001 was 1.6 h consistently throughout doses. Urinary excretion of QGC001 and EC33 was below 2 % of the administered dose. When compared with placebo, QGC001 did not significantly change PRC, PAldo, UAldo, PCop, PCort or UCort. No significant change was observed for supine HR, SBP and DBP in any treatment group. Conclusion Single oral administration of QGC001 up to 1,250 mg in healthy volunteers was well-tolerated. Following oral administration, QGC001 is absorbed via the gastrointestinal tract and converted partially into its active metabolite EC33 in plasma. As in animal experiments, in normotensive subjects QGC001 had no effect on the systemic renin-angiotensin-aldosterone parameters and on PCop concentrations, a marker of vasopressin release. In normotensive subjects, a single dose of QCG001 had no effect on SBP, DBP or HR. These data support further evaluation of multiple oral doses of QGC001 in human volunteers and its clinical efficacy in hypertensive patients

Phosphorylation states at multiple AMPAR subunits after acute stress.

<p>Immunoblots and histograms of the ratio between the phosphorylated form and total amount of the protein (white bars: no stress group, cross hatched bars: stress group). In the mPFC (A), stress decreased Ser831-GluA1 phosphorylation (p<0.05) and increased Ser880-GluA2 phosphorylation (p<0.05). In the amygdala (B), stress increased Ser845-GluA1 phosphorylation (p<0.05) and decreased Tyr876-GluA2 and Ser880-GluA2 phosphorylation (p<0.05. In the DH (C), stress decreased Ser831-GluA1 phosphorylation (p<0.05). In the VH (D), stress increased Ser845-GluA1 phosphorylation (p<0.05) and decreased Ser880-GluA2 phosphorylation (p<0.05). Immunoblots (100 kDa) illustrate the phosphorylated form of the protein (left: control, right: stress). The number of animals per group is 10 for controls and 11 for stress. Unpaired, two-tailed, Student's T-test. *, p<0.05, compared with the corresponding control group.</p

Summary of changes in AMPAR phosphorylation of GluA1 and GluA2 subunits after stress in the four regions studied.

<p>Summary of changes in AMPAR phosphorylation of GluA1 and GluA2 subunits after stress in the four regions studied.</p

Adrenal adaptation in potassium-depleted men: role of progesterone?

International audienceBACKGROUND:In rodents, the stimulation of adrenal progesterone is necessary for renal adaptation under potassium depletion. Here, we sought to determine the role of progesterone in adrenal adaptation in potassium-depleted healthy human volunteers and compared our findings with data collected in patients with Gitelman syndrome (GS), a salt-losing tubulopathy.METHODS:Twelve healthy young men were given a potassium-depleted diet for 7 days at a tertiary referral medical centre (NCT02297048). We measured by liquid chromatography coupled to tandem mass spectroscopy plasma steroid concentrations at Days 0 and 7 before and 30 min after treatment with tetracosactide. We compared these data with data collected in 10 GS patients submitted to tetracosactide test.RESULTS:The potassium-depleted diet decreased plasma potassium in healthy subjects by 0.3 ± 0.1 mmol/L, decreased plasma aldosterone concentration by 50% (P = 0.0332) and increased plasma 17-hydroxypregnenolone concentration by 45% (P = 0.0232) without affecting other steroids. CYP17 activity, as assessed by 17-hydroxypregnenolone/pregnenolone ratio, increased by 60% (P = 0.0389). As compared with healthy subjects, GS patients had 3-fold higher plasma concentrations of aldosterone, 11-deoxycortisol (+30%) and delta 4-androstenedione (+14%). Their post-tetracosactide progesterone concentration was 2-fold higher than that of healthy subjects and better correlated to plasma potassium than to plasma renin.CONCLUSION:The increase in 17-hydroxypregnenolone concentration after mild potassium depletion in otherwise healthy human subjects suggests that 17 hydroxylation of pregnenolone prevents the increase in progesterone observed in potassium-depleted mice. The unexpected over-response of non-mineralocorticoid steroids to tetracosactide in GS subjects suggests that the adrenal system not only adapts to sodium depletion but may also respond to hypokalaemia

P-glycoprotein influences urinary excretion of aldosterone in healthy individuals

International audienceObjectivesP-glycoprotein (P-gp), the product of the ABCB1 gene, is involved in the transport of aldosterone and cortisol in adrenal cells in vitro but its physiological role in humans remains controversial. Our objective was to test the influence of P-gp polymorphisms on aldosterone.MethodsWe evaluated plasma aldosterone concentration (PAC), urinary aldosterone, and blood pressure in a cohort of white normotensive men at baseline on diets unrestricted for sodium and potassium and after a 5-day treatment with 500 mg b.i.d. clarithromycin, a P-gp inhibitor. Included were 20 homozygous wild-type (P-gp0), 20 heterozygous (P-gp1), and 20 individuals with combined 2677G>T/A-3435C>T loss-of-function polymorphism of the ABCB1 gene (P-gp2).ResultsAt baseline, PAC, urinary aldosterone, urinary free cortisol to urine creatinine ratios, and blood pressure did not differ in the three genotypes. After clarithromycin administration, the urinary aldosterone to creatinine ratio increased by an average of 30% in the entire cohort (P < 0.001, n = 60). Increases were pronounced in P-gp1 (+40%; P = 0.014) and P-gp2 individuals (+50%; P = 0.020) but lesser and were NS in P-gp0 individuals (+10%; P = 0.259). PAC also increased from baseline after clarithromycin treatment in all individuals (+19%, P = 0.050); however, the increase in PAC was NS when the three genotypes were analyzed separately.ConclusionIn our experimental conditions, the interaction between P-gp inhibition and the ABCB1 genotype, suggests that aldosterone is indeed a physiological endogenous substrate of P-gp in humans and that P-gp interferes with the net equilibrium between aldosterone secretion and elimination processes in humans.Clinical Trial Registration-URL http//www.clinicaltrials.gov. Unique identifier NCT01627665

Simultaneous Characterization of Metabolic, Cardiac, Vascular and Renal Phenotypes of Lean and Obese SHHF Rats

International audienceIndividuals with metabolic syndrome (MetS) are prone to develop heart failure (HF). However, the deleterious effects of MetS on the continuum of events leading to cardiac remodeling and subsequently to HF are not fully understood. This study characterized simultaneously MetS and cardiac, vascular and renal phenotypes in aging Spontaneously Hypertensive Heart Failure lean (SHHF +/? regrouping +/+ and +/cp rats) and obese (SHHF cp/cp , ''cp'' defective mutant allele of the leptin receptor gene) rats. We aimed to refine the milestones and their onset during the progression from MetS to HF in this experimental model. We found that SHHF cp/cp but not SHHF +/? rats developed dyslipidemia, as early as 1.5 months of age. This early alteration in the lipidic profile was detectable concomitantly to impaired renal function (polyuria, proteinuria but no glycosuria) and reduced carotid distensibility as compared to SHHF +/? rats. By 3 months of age SHHF cp/cp animals developed severe obesity associated with dislipidemia and hypertension defining the onset of MetS. From 6 months of age, SHHF +/? rats developed concentric left ventricular hypertrophy (LVH) while SHHF cp/cp rats developed eccentric LVH apparent from progressive dilation of the LV dimensions. By 14 months of age only SHHF cp/cp rats showed significantly higher central systolic blood pressure and a reduced ejection fraction resulting in systolic dysfunction as compared to SHHF +/?. In summary, the metabolic and hemodynamic mechanisms participating in the faster decline of cardiac functions in SHHF cp/cp rats are established long before their physiological consequences are detectable. Our results suggest that the molecular mechanisms triggered within the first three months after birth of SHHF cp/cp rats should be targeted preferentially by therapeutic interventions in order to mitigate the later HF development

Circulation

BACKGROUND: Myocardial infarction (MI) induces a repair response that ultimately generates a stable fibrotic scar. Although the scar prevents cardiac rupture, an excessive profibrotic response impairs optimal recovery by promoting the development of noncontractile fibrotic areas. The mechanisms that lead to cardiac fibrosis are diverse and incompletely characterized. We explored whether the expansion of cardiac fibroblasts after MI can be regulated through a paracrine action of cardiac stromal cells. METHODS: We performed a bioinformatic secretome analysis of cardiac stromal PW1(+) cells isolated from normal and post-MI mouse hearts to identify novel secreted proteins. Functional assays were used to screen secreted proteins that promote fibroblast proliferation. The expressions of candidates were subsequently analyzed in mouse and human hearts and plasmas. The relationship between levels of circulating protein candidates and adverse post-MI cardiac remodeling was examined in a cohort of 80 patients with a first ST-segment-elevation MI and serial cardiac magnetic resonance imaging evaluations. RESULTS: Cardiac stromal PW1(+) cells undergo a change in paracrine behavior after MI, and the conditioned media from these cells induced a significant increase in the proliferation of fibroblasts. We identified a total of 12 candidates as secreted proteins overexpressed by cardiac PW1(+) cells after MI. Among these factors, GDF3 (growth differentiation factor 3), a member of the TGF-β (transforming growth factor-β) family, was markedly upregulated in the ischemic hearts. Conditioned media specifically enriched with GDF3 induced fibroblast proliferation at a high level by stimulation of activin-receptor-like kinases. In line with the secretory nature of this protein, we next found that GDF3 can be detected in mice and human plasma samples, with a significant increase in the days after MI. In humans, higher GDF3 circulating levels (measured in the plasma at day 4 after MI) were significantly associated with an increased risk of adverse remodeling 6 months after MI (adjusted odds ratio, 1.76 [1.03-3.00]; P=0.037), including lower left ventricular ejection fraction and a higher proportion of akinetic segments. CONCLUSIONS: Our findings define a mechanism for the profibrotic action of cardiac stromal cells through secreted cardiokines, such as GDF3, a candidate marker of adverse fibrotic remodeling after MI. REGISTRATION: URL: https://www. CLINICALTRIALS: gov; Unique identifier: NCT01113268