Previously, we have shown that male mice exposed to maternal separation and early weaning (MSEW)—a mouse model of early life stress—display increased mean arterial pressure compared with controls when fed a high-fat diet. As the stimulation of sensory nerves from fat has been shown to trigger the adipose afferent reflex, we tested whether MSEW male mice show obesity-associated hypertension via the hyperactivation of this sympathoexcitatory mechanism. After 16 weeks on high-fat diet, MSEW mice displayed increased blood pressure, sympathetic activation, and greater depressor response to an α-adrenergic blocker when compared with controls ( P <0.05; n=8), despite no differences in adiposity and plasma leptin. The acute infusion of capsaicin in epididymal white adipose tissue (1.5 pmol/μL of capsaicin, 8 μL/per site, 4 sites, bilaterally) increased the total pressor response in MSEW mice compared with controls (110±19 versus 284±33 mm Hg×30 minutes; P <0.05; n=8). This response was associated with neuronal activation in OVLT, posterior paraventricular nucleus of the hypothalamus, and RVLM ( P <0.05 versus control; n=6–7). Renal denervation abolished both the acute and chronic elevated mean arterial pressure in obese MSEW mice. Moreover, selective sensory denervation of epididymal white adipose tissue using resiniferatoxin (10 pmol/µL solution; n=6) decreased mean arterial pressure in obese MSEW mice only ( P <0.05 versus control). Obese MSEW mice displayed increased epididymal white adipose tissue levels of both tryptophan hydroxylase (Tph1) mRNA expression and its synthesis product serotonin (8.3±1.9 versus 16.6±1.7 ug/mg tissue; P <0.05 versus control). Thus, afferent sensory signals from epididymal white adipose tissue may contribute to the exacerbated fat–brain–blood pressure axis displayed by obese male mice exposed to early life stress.

Experimental stimulation of sensory nerves from white adipose tissue contributes to the increased sympathetic activation associated with obesity‐induced hypertension as part of a mechanism called adipose afferent reflex (AAR). Previously, we have demonstrated that male mice exposed to maternal separation and early weaning (MSEW), a mouse model of early life stress display increased mean arterial pressure (MAP) and sympathetic activation compared to control mice when fed a high fat diet (HF). Moreover, MSEW‐HF males display exacerbated blood pressure responses to the acute stimulation of the AAR in epididymal white adipose tissue (eWAT) compared to control mice. Thus, the aim of this study was to investigate the contribution of the renal nerves in the chronic increase in MAP and in the acute AAR stimulation in MSEW males fed HF. Male C57BL/6J mice pups were separated from the dams from postnatal day (PD) 2 to PD 16 and weaned early on PD 17. Control litters remained undisturbed with the dams and were weaned on PD 21. After weaning, Control and MSEW mice were placed on HF (60% Kcal from fat) for six months. Two weeks before the experiments a set of mice (n=4) were implanted with radiotelemetry. At six months, MAP was recorded at baseline and 6 days after bilateral renal denervation (RDNX, 10% phenol in ethanol). MSEW mice fed a HF displayed significantly increased chronic baseline MAP compared to controls (122±2 vs. 114±2 mmHg respectively, p<0.05). RDNX decreased MAP in both groups and abolished the differences between groups (MSEW: 112±2 vs. Control: 110±2 mmHg, p<0.05). Another set of mice was subjected to sham or RDNX surgery 4 days prior to the acute AAR stimulation with capsaicin (CAP; 0.5 nmol/ul; 4 sites; 4ul/min; 2 min; bilateral; n=7) to determine the MAP response. Mice were anesthetized, fitted with carotid catheters, and epididymal white adipose tissue (eWAT) was exposed for saline and CAP infusions. In these experiments, only in obese sham MSEW displayed increased MAP in response to CAP (baseline: 98±1 vs. CAP: 104±2 mmHg, p<0.05). RDNX significantly decreased baseline MAP in Control (baseline sham: 93±1 vs. baseline RDNX: 78±4 mmHg, p<0.05) and in MSEW (baseline sham: 98±1 vs. baseline RDNX: 80±3 mmHg, p<0.05) obese male mice. Moreover, CAP infusion in eWAT failed to increase MAP after RDNX in obese MSEW male mice (baseline RDNX: 80±3 vs. CAP RDNX: 75±6 mmHg). Taken together these results show that the renal nerves display an important contribution to the mechanism by which male mice exposed to early life stress in combination with an obesogenic diet present an overactivation of the fat‐brain‐blood pressure axis in response to capsaicin.

Adipose tissue homeostasis is a complex processes influenced by many mechanisms that become dysregulated with obesity. We have previously shown that Maternal Separation and Early Weaning (MSEW), a mouse model of early life stress, exacerbates high fat diet (HF)‐induced fat deposition in female mice, but not male mice. In addition we have shown that female MSEW mice fed a HF show increased mineralocorticoid receptor (MR) expression in gonadal white adipose tissue (gWAT), which has been shown to promote adipose tissue differentiation. Therefore, the aim of this study was to determine the effect of MSEW combined with chronic HF on gWAT hypertrophy, the tissue source and circulating levels of aldosterone, and whether the pharmacological antagonism of the MR will prevent the exacerbated gWAT expansion in female MSEW mice. MSEW and control (C) mice were weaned onto HF (60 % Kcal from fat) for 20 weeks. Adipose tissue was collected from a small subset of mice to conduct collagenase digestion followed by magnetic activated cell sorting (MACS) for isolation of preadipocytes for RNAseq analysis (Novegene, Bejing, China). Another subset of MSEW and C mice were randomized to receive either vehicle (50% Ora swift in drinking water) or spironolactone (100 mg/kg/day in vehicle) treatment for 2 weeks. RNAseq analysis revealed an upregulation of genes involved in lipoprotein particle binding and protein‐lipid complex binding (Pcsk9/Trem2/Lpl/Stab2/Ldlr, P<0.05), and low‐density lipoprotein particle binding (Pcsk9/Trem2/Stab2/Ldlr, P<0.05). Female MSEW mice fed a HF for 20 weeks showed increased adiposity compared to C (42.81 ± 1.62 vs. 33.50±0.97 % body weight, respectively, P<0.05). Furthermore, the MSEW females fed a HF displayed increased gWAT whitening determined by reduced mitochondrial DNA density (0.8±0.09 vs. 1.08±0.07 MitoDNA/gDNA, P<0.05). Female MSEW mice fed a HF showed increased aldosterone levels compared to C counterparts (733.12 ± 89.22 vs. 475.95 ± 36.50 pg/mL respectively, P<0.05). Adrenal‐derived media explant Aldo was increased in obese female MSEW mice compared with C (83.55±10.44 vs. 38.31±7.65 pg/mg tissue, P<0.05) while it was similar in media from gWAT explants. Chronic MR antagonism induced a greater fat mass loss in female MSEW mice compared to both vehicle‐treated MSEW females (‐2.07±0.88 vs. 3.11±1.48 treatment delta adiposity, P<0.05) and spironolactone‐treated C (‐2.07±0.88 vs. 0.62±1.30 treatment delta adiposity, P=0.06). Overall, MSEW‐induced exacerbated adiposity was associated with decreased mitochondrial DNA, increased pro‐obesogenic gene expression, and increased serum aldosterone. Chronic MR antagonism reduced fat mass in female MSEW mice, suggesting that MSEW might enhance adipose tissue differentiation via MR activation and as well as increased LPL‐induced lipid uptake, a MR downstream effector gene. Taken together, these data show that MSEW exacerbates the adrenal gland‐derived aldosterone production, whereas the increased gWAT MR expression may contribute to adipocyte hypertrophy.

Early life stress (ELS) is associated with increases in body mass index and systolic blood pressure in adult life, leading to obesity and obesity‐induced hypertension, major risk factors for cardiovascular disease (CVD). Previously, we have shown that Maternal Separation and Early Weaning (MSEW), a model that mimics the effects of ELS observed in humans, exacerbates hypertensive stimuli responses in male and female mice. The aim of this study was to investigate the status of the water and electrolyte homeostasis (WEH) at baseline in mice exposed to MSEW. Therefore, we determined water intake and excretion, plasma and urinary electrolytes, glomerular filtration rate (GFR) and sodium transporters expression along the nephron in lean mice. MSEW litter pups were separated from dams from postnatal day (PND) 2 to 16, followed by early weaning on PND 17. Litters that remained undisturbed and were weaned on PND 21 served as controls. At weaning, mice were placed on a low‐fat diet (LF, 10% Kcal from fat) for 12 weeks. Single mouse metabolic cages were used to measure water intake and urine excretion. GFR was measured using a transcutaneous technique. MSEW males fed a LF showed an increase in water intake compared to controls (4.14 ± 0.32 vs. 3.4 ± 0.36 mL/day respectively, P<0.05), urine excretion (1.01 ± 0.16 vs. 0.61 ± 0.07 mL/day, P<0.05), and potassium excretion (12.05 ± 2.48 vs. 7.29 ± 1.49 µmol/day/g BW, P<0.05). MSEW mice showed no differences in plasma sodium (165.16 ± 2.44 vs. 161.34 ± 2.67 mmol/L), plasma potassium (5.42 ± 0.19 vs. 5.86 ± 0.16 mmol/L) or sodium excretion compared to controls (3.65 ± 0.72 vs. 3.24 ± 1.43 µmol/day/g BW, respectively). In addition, MSEW males showed increased GFR compared to controls (1.12 ± 0.05 vs. 0.90 ± 0.03 mL/min/100g BW, P<0.05). Renal cortex NHE3 expression was increased in MSEW compared to control mice (7.74 ± 2.22 vs. 1.2 ± 0.25 2^ddCt, respectively, P<0.05) while NCC expression was decreased compared to controls (0.38 ± 0.13 vs. 1.25 ± 0.32 2^ddCt respectively, P<0.05). Yet, no significant changes between MSEW and control mice were found in renal transporters NKCC2 (0.91 ± 0.26 vs. 1.23 ± 0.29 2^ddCt, respectively), or in the ENaC subunits ENaCα (0.97 ± 0.09 vs. 1.12 ± 0.21 2^ddCt), ENaCβ (1.14 ± 0.13 vs. 1.14 ± 0.27 2^ddCt), and ENaCγ (0.88 ± 0.13 vs. 1.08 ± 0.17 2^ddCt). Lean female MSEW mice showed no significant differences in water intake, urine excretion, urinary potassium, GFR, or renal transporters expression. Both increased NHE3 and reduced NCC expression have been linked to a greater potassium waste. These results show that the MSEW effects on WEH in lean mice are sex‐specific, where only male mice display hyperfiltration and alterations in specific renal transporters expression. Chronic hyperfiltration may result in kidney damage and over time, chronic kidney disease. This could be aggravated in animals exposed to a secondary stressor such as a high‐fat or high‐salt diet. Ongoing studies of plasma and urine osmolarity may provide insights into the potential me...

Visceral adiposity has been largely implicated in increased sympathetic activation, and pathogenesis and target organ damage associated with the development of obesity‐induced hypertension. The experimental stimulation of afferent excitatory signals from adipose tissue has been shown to contribute with the increased sympathetic activation associated with obesity‐induced hypertension as part of a sympathoexcitatory mechanism called the adipose afferent reflex (AAR). Previous studies from our lab have shown that male mice exposed to maternal separation and early weaning (MSEW), a mouse model of early life stress, display increased mean arterial pressure (MAP) compared to control mice when fed a high fat diet (HF) that correlates with increased neuronal activation in non‐endocrine neurons in the posterior paraventricular nucleus of the hypothalamus. Moreover, MSEW‐HF males display exacerbated blood pressure responses to the acute stimulation of the AAR in epididymal white adipose tissue (eWAT) compared to controls. Therefore, the aim of this study was to determine the contribution of the afferent sensory nerves innervating the eWAT in the chronically increased blood pressure displayed by obese MSEW male mice. Male C57BL/6J mice pups were separated from the dams from postnatal day (PD) 2 to PD 16 and weaned early on PD 17. Control litters remained undisturbed with the dams and were weaned on PD 21. After weaning, Control and MSEW mice were placed on HF (60% Kcal from fat) for four months. At 14 weeks, mice (n=8) were implanted with radiotelemetry for chronic MAP measurements. After two weeks of recovery, MAP baseline was recorded during five consecutive days followed by eWAT denervation. For this procedure, eWAT was exposed and infused with vehicle (0.6% ethanol in saline; 4ul; 8 sites; bilateral) or resiniferatoxin (RTX, 10 pmol/ul, 8 sites, 4ul; bilateral). Then, MAP was continuously recorded for 4 days. RTX procedure in eWAT was validated using a GFP reporter mouse for sensory neurons, B6.129P2(Cg)‐Calcatm1.1(EGFP/HBEGF)Mjz/Mmnc, and the absence of a green signal in the RTX‐treated areas confirmed the afferent denervation. Sympathetic index was evaluated before and after denervations by a single injection of a ganglion blocker (mecamylamine; 5mg/kg; ip). MSEW mice fed a HF displayed significantly increased baseline MAP compared to controls (120±2 vs. 110±2 mmHg respectively, p<0.05). Ganglion blockade induced a greater MAP reduction in MSEW compared to controls (Delta MAP: ‐20±2 vs. ‐12±2 mmHg, respectively, p<0.05). Sham denervation did not change MAP from baseline in both groups. However, RTX denervation only decrease MAP in MSEW‐HF mice (baseline: 120±2 vs. RTX: 112±2 mmHg, p<0.05) and abolished the baseline MAP differences between groups. MAP decrease in response to ganglion blockade after RTX denervation was similar in both groups (MSEW: ‐14±4 vs. Control: ‐15±2 mmHg). Taken together these results show that afferent sensory nerves from epididymal white adipose tissue contribute to the exacerbated obesity‐indu...