bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetic defects in congenital solitary kidney rats cause low nephron numbers and predispose to severe renal damage
Authors: Wang Xuexiang , Johnson Ashley C , Lee Jonathan , Solberg‐Woods Leah , Garrett Michael R
Published: June 2021

The occurrence of people born with single kidney is ~1:1000. The clinical outcome for these patients and potential predisposition for decline in renal function and hypertension is controversial. Our lab recently developed a unique rat model that are born with single kidney (50–75% offspring) and demonstrate low nephron number. The model is currently being utilized to address whether early development (in utero) with a single kidney increases susceptibility to renal injury and hypertension compared to two kidney control and/or uninephrectomized rats. At an early age, renal hemodynamic data shows no hyperfiltration for spontaneous one kidney rats, whereas nephrectomized rats exhibit significantly elevated GFR compared to two kidney control. At advanced age (18 months), spontaneous one kidney rats demonstrate the highest proteinuria, histological kidney injury and lowest GFR. We are currently investigating early kidney developmental changes using molecular genomic approaches (whole transcriptome and miRNA analysis using NGS) and whole organ kidney culture to identify the mechanism of low nephron number. In summary, congenital single kidney rats exhibit more severe renal damage compared to two kidney control (and nephrectomized rats) and genes involved in nephrogenesis are likely implicated and program the model to develop significant kidney disease with age. Support by UMMC IRSP and Hearin Foundation.
Is there truly a sympathetic component to aldosterone hypertension?
Authors: Lohmeier Thomas , Liu Boshen , Georgakopoulos Dimitrios
Published: June 2021

Studies in rodents suggest that sympathetic activation may contribute to aldosterone (Aldo) hypertension (HT), but evidence in non‐rodent animals and in humans is equivocal. The goal of this study was to use complementary techniques to determine whether sympathetic activation contributes to HT induced by high circulating levels of Aldo. After control measurements, Aldo was infused IV for 14 days at 12μg/kg/day in 9 dogs maintained on a constant Na intake (~65 mmol/day). This increased plasma Aldo (control= 4.3±0.4 ng/dL) ~ 10‐fold and mean arterial pressure (MAP) from 103±2 to 121±5 mmHg. In addition, Aldo infusion was associated with marked polydipsea, increases in plasma osmolality (293±1 to 297±1 mOsm/kg H20) and vasopressin (0.86±.32 to 1.73±0.34 pg/mL), hypokalemia (4.2±0.1 to 2.3±0.1) and suppression of PRA to undetectable levels. Despite these responses, there were no significant changes in plasma [norepinephrine], and depressor responses to ganglionic blockade with hexamethonium (5 mg, IV) were actually attenuated during Aldo HT, suggesting suppression of sympathetic activity. Finally, in 2 dogs, bilateral renal denervation failed to attenuate Aldo HT. In these dogs, MAP was 122 and 142 before and 122 and 144 two weeks after bilateral renal denervation. Thus, the failure of both global and renal specific measures to demonstrate sympathetic activation indicates that classic renal mineralocorticoid mechanisms account for the initial progression of HT produced by pathophysiological circulating levels of Aldo. HL‐51971
Nicotine reduces blood pressure in mouse model of systemic lupus erythematosus
Authors: Mathis Keisa W.
Published: June 2021

Stimulation of the cholinergic anti‐inflammatory pathway through nicotinic acetylcholine receptors (α7nAchR) can suppress systemic inflammation by a mechanism involving vagus nerve activation. Evidence supports an important role for immune system activation and inflammation in the pathogenesis of hypertension. Hypertension and renal injury are prevalent in human patients with the chronic autoimmune inflammatory disorder, systemic lupus erythematosus (SLE), and some evidence suggests that these patients have impaired vagal function. Based on this, we hypothesized that the cholinergic anti‐inflammatory pathway is impaired in SLE and that activation of the pathway via nicotine would reduce blood pressure in mice with SLE. Female SLE (NZBWF1) and control (NZW) mice were infused with nicotine or vehicle for 7 days (2 mg/kg subcutaneous) after which time blood pressure was assessed. Blood pressure (mmHg) was higher in vehicle‐treated SLE mice compared to controls (135±7 vs. 114±4). Blood pressure in nicotine‐treated SLE (119±9) and control (114±1) mice was similar to vehicle‐treated controls. These data implicate impairment of the cholinergic anti‐inflammatory pathway in the development of SLE‐associated hypertension and suggest that it may be an important pathway to target clinically in patients with chronic inflammatory diseases associated with hypertension. AHA4350019, 5T32HL10532, HL051971
Sympathetic vascular transduction following spontaneous MSNA bursts is augmented in young black men
Authors: Credeur Daniel Paul , Holwerda Seth W , Fairfax Seth T , Keller David M , Fadel Paul J
Published: June 2021

Young blacks exhibit an augmented vasoconstrictor response to α‐adrenergic agonists compared to whites. Whether this increase in α‐adrenergic receptor sensitivity in blacks evokes greater vasoconstriction to spontaneous bursts of muscle sympathetic nerve activity (MSNA) is unknown. Thus, in fourteen young normotensive black (N=7) and white men (N=7), MSNA (microneurography) and femoral artery blood flow (duplex Doppler ultrasound) were continuously measured for 20 minutes under resting conditions. Signal averaging was used to characterize beat‐by‐beat changes in leg vascular conductance (LVC) for 15 cardiac cycles following each individual MSNA burst. Variations in MSNA burst pattern (single vs. multiple successive bursts) and burst size (height) were considered. Blacks demonstrated a significantly greater reduction in LVC following single (blacks: −11.2±1.7 vs. whites: −4.6±1.24%; P<0.05) as well as multiple MSNA bursts (blacks: −19.4±2.0 vs. whites: −13.2±1.6; P<0.05). In addition, increases in burst height evoked graded reductions in LVC in both groups, with blacks having significant greater reductions in LVC. These preliminary findings suggest an augmented sympathetic vascular transduction in young normotensive blacks in response to spontaneous bursts of MSNA.

Supported by NIH Grant R15HL096103 (DMK) and RO1 HL093167 (PJF)
Sympathetic nerve activity and systemic arterial pressure during obstructive sleep apnea in conscious rats
Authors: Sukeguchi Chie , Miki Kenju
Published: June 2021

This study investigated sympathetic nerve activity responses in obstructive sleep apnea. Wistar male rats were chronically instrumented for measurements of renal (RSNA) and lumbar (LSNA) sympathetic nerve activity, as well as for an electroencephalogram, electromyogram, and electrocardiogram and systemic arterial pressure, and with a tracheal balloon for induction of apnea. At least 3 days after the surgery, rats were subjected to sleep apnea by inflating the tracheal balloon for 40 seconds during non‐rapid eye movement sleep. RSNA and LSNA increased immediately after the onset of tracheal balloon inflation and by approximately 3 – 4 fold compared with those at pre‐inflation levels. Heart rate decreased in a biphasic manner; it began to fall gradually during the initial several seconds and then it fell abruptly by approximately 200 beats/min. Systemic arterial pressure increased because of the tracheal balloon inflation. We conclude that sympathetic nerve activity plays a critical role in increasing systemic arterial pressure during obstructive sleep apnea in rats. JSPS Grant.
Severe hemorrhage attenuates cardiopulmonary chemoreflex (CCR) control of renal and adrenal sympathetic nerves via adenosine operating in the nucleus of the solitary tract (NTS)
Authors: Minic Zeljka , Li Cailian , O'Leary Donal S , Scislo Tadeusz J.
Published: June 2021

During severe hemorrhage adrenal sympathoactivation is accompanied by paradoxical renal sympathoinhibition and bradycardia. Adenosine operating in the NTS via A1 and A2a receptors contributes to the paradoxical effects and it inhibits CCR mediated sympathoinhibition (Am J Physiol 291: H2453‐H2461, 2006; 303: R539‐R550, 2012; FASEB J 25: 844.3, 2011). Therefore, we hypothesized that adenosine released into the NTS during severe hemorrhage will attenuate the CCR and that blockade of NTS adenosine receptors will reverse this attenuation. We compared renal and adrenal sympathetic responses evoked by right atrial injections of 5HT3 receptor agonist, phenylbiguanide (PBG, 1–8 μg/kg), under control conditions, during hemorrhage and during hemorrhage after blockade of NTS adenosine receptors with bilateral microinjections of 8‐(p‐sulfophenyl) theophylline (1 nmol/100 nl) in urethane/chloralose anesthetized rats. Hemorrhage significantly attenuated the CCR responses and blockade of NTS adenosine receptors reversed this attenuation. We conclude that adenosine released into the NTS contributes to the paradoxical posthemorrhagic effects but it also serves as a negative feedback regulator of the CCR. HL‐67814
Muscle metaboreflex‐induced increases in ventricular performance are limited in hypertension due to exaggerated coronary vasoconstriction
Authors: Spranger Marty D. , Sala‐Mercado Javier A. , Kaur Jasdeep , Abu‐Hamdah Rania , O'Leary Donal S.
Published: June 2021

Increases in myocardial O2 consumption mainly occur via increases in coronary blood flow (CBF) inasmuch as cardiac O2 extraction is high even at rest. However, during exercise sympathetically‐mediated constrictor tone can limit increases in CBF. Increased sympathetic activity during exercise may stem from muscle metaboreflex activation (MMA). Sympathetic activity is often elevated even at rest in subjects with hypertension (HTN). We tested whether HTN causes exaggerated coronary vasoconstriction during mild exercise with MMA which thereby limits increases in CBF and ventricular performance. MMA was achieved by reducing hindlimb blood flow in canines by ~60% during treadmill exercise before and after α1‐adrenergic blockade. Experiments were repeated in the same animals following the induction of HTN (2K1C) which increased mean arterial pressure from 94.7 ± 3.8 to 130.5 ± 11.0 mmHg. In HTN, MMA‐induced increases in CBF, left ventricular dP/dtmax and cardiac output were markedly reduced to only 37 ± 12%, 26 ± 16% and 17 ± 8% of the changes observed in control. In this setting, α1‐blockade nearly restored these responses to the levels observed when normotensive. We conclude that in HTN, exaggerated MMA‐induced increases in sympathetic activity functionally vasoconstrict the coronary vasculature impairing increases in CBF which limits O2 delivery and ventricular performance. NIH HL‐ 55743 and HL‐095819
Lack of interaction of co‐existing TRPV1 and CB1 receptors indicates differential control of separate basal and synchronous glutamate release mechanisms in the solitary tract nucleus
Authors: Andresen Michael C. , Hofmann Mackenzie E. , Fawley Jessica A.
Published: June 2021

Cannabinoid receptor agonists structurally parallel TRPV1 agonists. Both endocannabinoids and endovanilloids resemble membrane derived phospholipids. In spinal dorsal horn, TRPV1 and CB1 co‐localize. Can single compounds crossover to activate both receptors? Can activation of one receptor influence function of the other? Support for both CB1/TRPV1 interactions exists. Here we examined CB1 and TRPV1 receptor activation in synaptic transmission by solitary tract afferents (ST) in horizontal slices of the solitary tract nucleus (NTS). The CB1 selective compound ACEA (10 μM) depressed excitatory postsynaptic current amplitude (ST‐EPSCs) in 78% of ST afferents (TRPV1+, capsaicin sensitive, −38 ± 7% n=14 vs. TRPV1‐ −37 ± 10%, n= 7) but CB1‐ afferents were uncommon. In addition, CB1 activation increased synaptic failures only in TRPV1+. ACEA did not alter spontaneous EPSCs frequency or amplitude. WIN 55,212–2 (10 μM) had identical actions. ACEA actions were blocked by CB1 antagonist AM251 (10 μM). CB1 activation or block failed to alter temperature driven changes in basal or spontaneous EPSCs that depend on TRPV1. The results indicate widespread presynaptic CB1 and suggest that CB1 selectively targets action potential dependent glutamate release likely by inhibiting voltage dependent calcium channels (decreasing release probability) and action currents that depolarize terminals (synaptic failures).
Auditory and visual stimuli evoke a synchronised activation of sympathetic, respiratory and motor outputs after disinhibition of midbrain colliculi
Authors: Muller‐Ribeiro Flavia C.F. , Fontes Marco P. , Dampney Roger A.L. , Goodchild Ann K.
Published: June 2021

The midbrain colliculi are involved in cardiovascular and behavioural responses to threatening stimuli. Disinhibition of this area can evoke spontaneous synchronized sympathetic and respiratory responses in anesthetized rats. This raised the question as to whether naturalistic stimuli can generate such responses. In urethane‐anesthetized, paralysed, vagotomised and artificially ventilated rats, abrupt sound (n=7) or bright light (n=5) generated synchronised bursts of activity in sympathetic (309 ± 30 or 217 ± 38%), phrenic (60 ± 18 or 43 ± 15 bpm), and sciatic (17 ± 4 or 15 ± 3%) nerve activities and blood pressure (30 ± 4 or 31 ± 8 mmHg) after disinhibition of ventral sites in superior and inferior colliculi. The onsets of the sympathetic, respiratory and sciatic responses following auditory or light stimuli occurred within a time period of <50 msec. Simultaneous multi‐sensory stimulation evoked an enhanced response. Blockade of the dorsomedial hypothalamus (DMH), dorsolateral periaqueductal grey (dlPAG) or cuneiform nucleus using muscimol did not affect the responses. Our findings show that neurons in the colliculi can generate synchronised cardiovascular, respiratory and behavioural responses triggered by a sudden auditory or visual stimulus. The output pathways from the colliculi do not depend on synapses within the DMH, dlPAG or cuneiform nucleus.

Supported by NHMRC 1028183 and 570912 and by Capes.
Elderly blacks have a similar sympathetic neural responsiveness but greater pressor response to cold stress than elderly whites
Authors: Okada Yoshiyuki , Edwards Jeffrey , Jarvis Sara S. , Best Stuart A. , Meier Rhonda L. , Bivens Tiffany B. , Vongpatanasin Wanpen , Levine Benjamin D. , Fu Qi
Published: June 2021

Muscle sympathetic nerve activity (MSNA) response to cold stress has been reported to be greater in young blacks than whites. We evaluated whether elderly blacks have greater sympathetic activation during cold stress than age matched whites. MSNA (microneurography), blood pressure (BP), and heart rate (HR) were measured during supine baseline, 2‐min cold pressor test (CPT), and 3‐min of recovery in 10 elderly blacks [age 65 ± 4 (SD) yrs] and 30 elderly whites (69 ± 7 yrs). Mean BP increased during CPT (P < 0.001), while it was greater in blacks than whites at 2 min of CPT (114 ± 21 vs 103 ± 13 mmHg, P = 0.042). HR increased during CPT (P < 0.001) without a group difference (72 ± 7 vs 65 ± 10 mmHg, P = 0.113). MSNA burst frequency, burst incidence, and total MSNA increased during CPT (all P < 0.001). None of these variables showed racial differences during baseline (burst frequency: 51 ± 12 vs 48 ± 10 bursts/min, burst incidence: 77 ± 16 vs 80 ± 15 bursts/100beats, and total MSNA: 781 ± 266 vs 759 ± 241 units) or CPT (63 ± 10 vs 61 ± 10 bursts/min, 88 ± 10 vs 93 ± 8 bursts/100beats, and 1311 ± 636 vs 1476 ± 720 units; all P >; 0.050). Thus, elderly blacks have a greater pressor response to cold stress than elderly whites, which is not attributable to sympathetic neural response. Other vasoconstriction systems (i.e., sympathetic vascular transduction and/or non‐adrenergic vasoconstrictor mechanisms) must be involved.

Supported by NIH R01 HL091078.
An inverse relationship between testosterone and muscle sympathetic nerve activity: an investigation in lean healthy females
Authors: Madison Erin Elizabeth , Miner Jonathon Cory , Miner Jennifer Ann , Kaplan Paul F. , Minson Christopher Todd
Published: June 2021

It has been documented that women with polycystic ovary syndrome (PCOS) have an elevated level of testosterone, which coincides with having an increased resting level of sympathetic activity, as measured by muscle sympathetic nerve activity (MSNA). However, there is a lack of research into the effect of moderate testosterone levels on sympathetic activity in lean, healthy females. In order to isolate the impact of testosterone in these women, the level of estrogen and progesterone were controlled via hormonal suppression in 8 healthy, lean women using a gonadotropin hormone antagonist (GnRHa) protocol. After four continuous days of estradiol and progesterone suppression, each subject came into the lab for testing of total testosterone level via blood sample, and MSNA via microneurography of the peroneal nerve. The study day testosterone level in these lean women ranged from 10.3 – 36.4 ng/dL, and the average burst frequency at rest was 2 – 22 bursts/min. A negative correlation was found between total testosterone levels and MSNA (R2=48.26, p=0.056). Although a positive correlation has previously been documented in women with increased levels of the hormone testosterone, this study showed a negative relationship between these two variables under normal ranges of testosterone.

Supported by NIH Grant HL081671.
The neurosteroid metabolite of progesterone, 3α‐OH‐dihydroprogesterone (3α‐OH‐DHP), is required for attenuated baroreflex mediated sympathoexcitation in pregnancy
Authors: Phaup J. Glenn , Hasser Eileen M. , Heesch Cheryl M.
Published: June 2021

Exogenous 3á‐OH‐DHP mimics effects of pregnancy on arterial baroreflex (BX) control of renal sympathetic nerve activity (RSNA). Current experiments chronically blocked endogenous 3á‐OH‐DHP formation (Finasteride administration) and evaluated BX function in NP and near‐term pregnant (P, d21) rats. Either vehicle (Veh) or Finasteride (Fin, 25mg/kg) was given daily (s.cu.) for 7 days. RSNA electrodes were implanted on d5 of treatment. On d7, reflex changes in RSNA to increases (phenylephrine, iv) and decreases (nitroprusside, iv) in mean arterial pressure (MAP) were obtained in conscious rats and data fit to a logistic curve. As seen previously, in Veh treated P rats maximum RSNA was decreased (Veh‐P, 155 ± 14%; Veh‐NP, 335 ± 44% baseline), and baroreflex curve midpoint was shifted to lower MAP (Veh‐NP, 100 ± 4; Veh‐P, 84 ± 5 mmHg). Chronic Finasteride had no effect on BX function in Veh‐NP rats, but restored maximum RSNA in P rats (Fin‐P, 309 ± 25% baseline) to NP levels, while curve midpoint remained low (Fin‐P, 74 ± 3 mmHg). Thus, 3á‐OH‐DHP, likely through positive modulation of GABAA receptors in the RVLM, contributes to attenuated BX sympathoexcitation in P rats. NIH R01 HL091164 (CMH)
Effects of the reversible acetylcholinesterase inhibitor pyridostigmine on vascular reactivity of spontaneously hypertensive rats (SHR)
Authors: Lataro Renata Maria , Silva Marcondes A. B. , Tostes Rita C. A. , Salgado Helio Cesar
Published: June 2021

We investigated the chronic (16 weeks) effect of pyridostigmine (PYR; 1.5 mg/kg/day) on mesenteric vascular reactivity of SHR. Responses to phenylephrine (Phe), sodium nitroprusside (SNP) and acetylcholine (Ach) were determined with and without endothelium and in the presence of vehicle or L‐NAME plus indomethacin. Electrical stimulation (EE)‐induced contractions were determined in the presence of vehicle, atropine or atropine plus L‐NAME. Ach‐induced relaxation was similar between groups. However, Ach‐induced vasodilation, in the presence of L‐NAME plus indomethacin, was augmented in SHR as compared to WKY and SHR+PYR. No differences in Phe‐induced contraction or SNP‐induced relaxation were observed among groups. EE‐induced contractions, in the presence of atropine, were significantly augmented in endothelium‐intact SHR, but not in SHR+PYR. L‐NAME augmented EE‐induced contractions in WKY and SHR+PYR, but not in SHR. No differences in EE‐induced contraction were observed under atropine plus L‐NAME. In conclusion, PYR improves vascular dysfunction in SHR, which is associated with decreased bioavailability of nitric oxide and is compensated by other endothelium‐derived factors. Financial support:FAPESP.
Promising applications of nano‐encapsulated (−) epigallocatechin gallate during adipogenesis in 3T3‐L1 cells
Authors: Bapat Priyanka Vinayak , Sun Ming , Wang Shu
Published: June 2021

Background

Overexpression of thermogenic genes in white adipose tissue can ameliorate obesity. Many studies reported acute thermogenic effects of epigallocatechin gallate (EGCG) in white adipocytes. However, its low stability limits its therapeutic applications. Objective of this study was to synthesize biocompatible nanoencapsulated EGCG (Nano‐EGCG) to increase stability and enhance biological effects.

Methods

Stability of Nano‐EGCG at 37°C was measured using HPLC system. 3T3‐L1 cells were treated at differentiation and maturation with native and Nano‐EGCG at 10 μM. Triglyceride content in mature adipocytes was measured by an enzymatic method. The gene expression levels were measured using real‐time PCR.

Results

Nano‐EGCG was more stable than native EGCG in vitro. Nano‐EGCG significantly (p<0.05) reduced triglyceride content in mature adipocytes as compared to EGCG. Moreover, when preadipocytes were induced to differentiate with Nano‐EGCG, the expression levels of peroxisome proliferator activated receptor gamma coactivator1‐alpha (PGC1α), and uncoupling protein family (UCP 1,2,3) were significantly increased as compared to native EGCG.

Conclusion

Nanoencapsulation significantly increased EGCG stability. Nano‐EGCG at low concentrations significantly reduced triglyceride content in mature adipocytes and increased the expression of thermogenic genes during differentiation.

Grant Funding Source: TTU research incentive funding
Identifying markers of insulin resistance in the plasma proteome
Authors: Jamnik Joseph , Garcia‐Bailo Bibiana , Borchers Christoph , El‐Sohemy Ahmed
Published: June 2021

Type 2 diabetes (T2D) represents a significant global health problem. The plasma proteome comprises numerous high‐abundance proteins that may be associated with the development of T2D. The objective was to determine the association between HOMA‐IR, a common measure of insulin resistance, and 54 plasma proteins belonging to disease‐associated pathways in a population of healthy young adults (n=488) from the Toronto Nutrigenomics and Health Study. Protein concentrations were measured by a multiple reaction monitoring HPLC‐MS/MS assay. General linear models were used to assess the association between plasma proteins and HOMA‐IR. Analyses were adjusted for age, sex, ethnicity, waist circumference, physical activity, and contraceptive use among women. Hemopexin, complement factor H, heparin cofactor II, afamin, α1‐anti‐trypsin, α2‐HS‐glycoprotein, apolipoprotein L1, complement C3, serum amyloid P‐component, vitronectin, and transthyretin were significantly (p<0.0009) positively correlated with HOMA‐IR. Adiponectin, an anti‐inflammatory protein, was inversely correlated with HOMA‐IR. These results suggest that a number of plasma proteins may be useful biomarkers of insulin resistance and T2D risk. Research support from the Advanced Foods and Materials Network.
Low‐dose cadmium (Cd) exposure up‐regulates glutathione (GSH) synthesis in cultured choroid plexus
Authors: Villalobos Alice , Young Robin , Stuart Samantha Francis
Published: June 2021

Having shown in primary cultures of rat choroid plexus that Cd induced oxidative stress and stimulated apical choline uptake, we aim to elucidate the role of GSH in cellular adaptation to Cd exposure. Choroid plexus primary cultures were exposed for 12 h to 0 or 500 nM CdCl2. Immunoblot and qRT‐PCR analyses indicated heme oxygenase‐1 (HO‐1) gene and protein were induced 50‐fold at 12 h. Heat‐shock protein 70 (HSP70) gene and protein were maximally induced at 6 h, and metallothionein‐1 (MT‐1) mRNA was induced 50‐fold at 12 h. Catalytic and modifier subunits of glutamate‐cysteine ligase (GCL), the rate‐limiting enzyme in GSH synthesis, also were induced. GCLC protein peaked at 6 h, while GCLM peaked at 9 h; the respective genes were induced 5‐fold and 4‐fold at 12 h. We also assayed GSH and oxidized glutathione (GSSG) after 12‐h exposure to 0 or 250 nM CdCl2 100 μM buthionine sulfoximine (BSO), an inhibitor of GCL. Cd increased GSH by 2‐fold and GSSG by 30‐fold. BSO reduced GSH by 92% and further decreased GSH:GSSG ratios in Cd‐treated cells. Furthermore, BSO enhanced induction of HO‐1, HSP70, MT‐1, GCLC and GCLM, stimulation of choline uptake, and cytotoxicity in Cd‐treated cells. Thus, as the epithelium adapts to Cd exposure GSH synthesis is up‐regulated at transcriptional and translational levels. GSH may facilitate cell survival and temper stress‐induced changes in choline transport.
Nesfatin‐1 does not influence intracellular calcium concentrations in neurons of the nucleus of the solitary tract or the paraventricular nucleus
Authors: Mimee Andrea , Ferguson Alastair V.
Published: June 2021

Nesfatin‐1 (nes‐1) is a neuropeptide which exerts potent effects on food intake, cardiovascular parameters, and the stress axis. Immunohistochemical studies have shown expression of nes‐1 in the nucleus of the solitary tract (NTS) and paraventricular nucleus (PVN). To date there is minimal data describing the mechanism of action of nes‐1 in the brain. Using whole cell patch clamp recordings, we have previously shown bath applied nes‐1(10 nM) influences the membrane potential of 63% of NTS and 71% of PVN neurons. Here, as a first step in investigating the mechanism of nes‐1 actions, we examined the effect of nes‐1 on intracellular Ca2+ concentrations ([Ca2+]i) in dissociated NTS and PVN neurons loaded with the Ca2+ indicator Fura2‐AM. Bath application of nes‐ 1 at 10 and 100 nM increased [Ca2+]i in 4% (n=75) and 3% (n=84) of NTS neurons, respectively, as assessed by a minimum 20% increase in the ratiometric value of [Ca2+]i. Furthermore, 9% (n=125) of PVN neurons showed an increase in [Ca2+]i in response to 10 nM nes‐1. Our findings show nes‐1 does not influence [Ca2+]i in NTS or PVN neurons. A large discrepancy also exists in the proportion of neurons reported as responsive to nes‐1 in our electrophysiological vs calcium imaging experiments, suggesting studies of nes‐1 effects on [Ca2+]i may not be an appropriate method of evaluating neuronal nes‐1 activation patterns or mechanisms of action.

Funding: NSERC, FQRNT, HSFO.
Nesfatin‐1 influences the excitability of subfornical organ neurons
Authors: Kuksis Markus Arnis , Dai Li , Ferguson Alastair V
Published: June 2021

Nesfatin‐1, a centrally acting anorexigenic peptide, is produced in several brain areas involved in metabolic function and has been implicated in appetite and thirst reflexes. The present study was thus undertaken to determine the specific role of nesfatin‐1 in appetite and thirst regulation mediated by the subfornical organ (SFO). We first used RT‐PCR and were able to confirm the presence of nesfatin‐1 in SFO. We then used whole‐cell patch clamp recordings to investigate the influence of nesfatin‐1 on the membrane potential of dissociated SFO neurons. Approximately 68.8% (42 of 61) of neurons tested showed a response to nesfatin‐1 (10 nM and 1 nM). Of the responding neurons, 50% depolarized by a mean depolarization of 10.6 ± 1.64 mV (n=21) and 50% hyperpolarized by a mean hyperpolarization of −8.8 ± 2.2 mV (n=21). No effect was observed at a concentration of 100 pM (n=3). This study has demonstrated that nesfatin‐1 has the ability to change the membrane potential of SFO neurons, and therefore can be identified as a potential modulator of SFO function.

Supported by the Canadian Institutes of Health Research
Insulin and glucose stimulation of vasopressin (VP) and oxytocin (OT) release requires glucokinase (GK) and PI3 kinase (PI3K)
Authors: Sladek Celia D , Song Zhilin , Stevens Wanida
Published: June 2021

Neurons in the supraoptic nuclei (SON) produce OT and VP and express insulin receptors (InsR) and GK. Since OT is an anorexic agent and GK and InsR are hallmarks of cells that function as ‘metabolic sensors’, we evaluated the effect of glucose, Ins, and their downstream effector, KATP channels, on OT/VP release from rat hypothalamo‐neurohypophyseal system explants, and the effect of blocking GK and PI3K (mediates Ins‐induced mobilization of glucose transporter, GLUT4) on Ins (3ng/ml) and Ins+glucose‐induced OT/VP release. Ins increased OT/VP release and the response was sustained when glucose was increased from 2 to 6 mM (p≤0.002). The OT/VP responses to Ins+glucose were blocked by the GK inhibitor, alloxan (4mM; p≤0.002), and the PI3K inhibitor, wortmannin (10nM; OT: p=0.03; VP: p≤0.002). Closing of KATP channels with 200nM glybenclamide increased OT/VP release (OT: p<0.003; VP: p<0.05). These results suggest that Ins activation of PI3K increases GK‐mediated ATP production inducing closure of KATP channels and stimulation of OT/VP release. The findings are consistent with SON OT/VP neurons functioning as ‘metabolic‐sensors’ to participate in appetite regulation. Supported by NIH R21 HD072428.
Behavioral Profile Assessment in Offspring of Swiss MiceTreated During Pregnancy and Lactation with Caffeine.
Authors: Silveira Anderson Luiz Bezerra , Melo Roberto Laureano , Seara Fernando Azevedo Cruz , Almeida Cláudio Silva , da Rocha Fabio Fagundes , Santos Alex Sander Nunes , Marinho Bruno Guimarães , Reis Luís Carlos , Cortes Wellington Silva
Published: June 2021

The objective was to assess the behavioral profile in offspring of Swiss mice treated during pregnancy and lactation with caffeine. For this purpose, two groups (n=6 each, and BW~35g) of female mice were treated during pregnancy and lactation by: tap water and caffeine solution at a concentration of 0,3mg/mL through oral route. The offsprings obtained, by completing 70 days of life, were underwent a behavioral battery test. Statistical analysis was performed by student t test and the different significance adopted was p<0.05. According to our results, it was not found any significant differences in the tail suspension and rotarod test as well. However, in elevated plus maze test, programmed animals had a lower percentage of entries in open arms (48.01±2.16 vs. 32.19±4.07, P<0.05) and higher number of entries in closed arms (15.57±3.13 vs. 12.77±2.31, P<0.05). Moreover, in the light‐dark box test there was significantly increase in number of transition in treated animals (23.00±1,03 vs. 29.46±1.796, P<0.05). In the meantime, in the von Frey test it was observed that programmed animals had a pattern hypernociception (p<0.0001). We can conjecture that caffeine treatment causes change in central adenosine signaling and, therefore, may have led a cytoarquitetonic alterations in limbic and cortical regions associated with anxiety‐like behavior and modulation of nociceptive responses.

Support: FAPERJ & CNPq.
Perinatal exposure to low levels of carbon monoxide impairs neurobehavior in adult mice
Authors: O'Neill John Timothy , Kehl Margaret L , Johnson Kristopher P , Cravedi Kevin , Barry Erin S , Grunberg Neil E
Published: June 2021

Maternal smoking is associated with several adverse neurological outcomes including increased risk for SIDS and attention deficit disorder. Perinatal exposure to carbon monoxide (CO) disrupts development of brain structures and associates with neurological disorders.

OBJECTIVE

to uncover age and sex differences in neurobehavioral phenotype of perinatally CO‐exposed mice. Pregnant mice were exposed to 0, 75, and 150 ppm of CO approximating levels found in smokers from embryonic day 0 until weaning. We utilized the Open Field Test (OFT), Acoustic Startle Response/Prepulse Inhibition (ASR/PPI), and the Neurobehavioral Severity Scale Revised (NSS‐R) to evaluate changes in neurological and behavioral functioning in rodents. Animals were evaluated at 14 and 60 days of life (do).

RESULTS

At 14 do: (1) NSS‐R increased (0.33±0.22, 1.56±0.19 and 1.27±0.23 at 0, 75 and 150 ppm respectively) indicating decreased sensory‐motor function after exposure to CO, with females showing greater sensitivity; (2) males and females exposed to CO showed decreased startle with and without pre‐pulse.

RESULTS

at 60 do: (1) OFT‐horizontal activity decreased without sexual preference; (2) CO‐exposed animals showed some enhanced startle with PPI. We conclude that perinatal exposure to CO is associated with mild neurological impairment in 14 do mice that persists to 60 days.

Funding: Flight Attendant Medical Research Institute
Ursolic acid potentiates muscarinic receptor‐induced ANP secretion in perfused beating rat atria
Authors: Kim Hye Yoom , Kwon Oh Jeong , Kho Joung Hyun , Tan Rui , Cho Kyung Woo , Kang Dae Gill , Lee Ho Sub
Published: June 2021

Ursolic acid has recently been reported to increase both atrial natriuretic peptide (ANP) secretion and mechanical dynamics in rabbit atria. The present study was designed to clarify the regulatory effects of ursolic acid on the ¥â‐adrenergic or muscarinic receptor‐mediated changes in ANP secretory and contractile function allowing measurement of atrial dynamics such as pulse pressure, stroke volume, and cAMP efflux in isolated perfused beating rabbit atria. Pretreatment with ursolic acid significantly attenuated the isoproterenol (¥â‐adrenergic agonist)‐ induced decrease in ANP secretion and increases in cAMP levels and atrial dynamics. Interestingly, ursolic acid concentration‐dependently accentuated the acetylcholine‐induced increase of ANP secretion and decrease of pulse pressure in the presence of isoproterenol (p < 0.001). These findings indicate that acetylcholine‐induced increase of ANP secretion is potentiated by ursolic acid, furthermore, acetylcholine‐induced decrease of atrial dynamics is also potentiated by ursolic acid, suggesting that ursolic acid regulates muscarinic receptor‐mediated secretory and contractile responses in perfused beating rabbit atria. Thus, this implicates for the beneficial effects of ursolic acid in the regulation of cardiovascular and body fluid homeostasis.
Protease‐activated receptor ‐3 signals independent of protease‐activated receptor‐1 to regulate endothelial nitric oxide synthase in human endothelial cells
Authors: Tillery Lakeisha Carrie , Motley‐Johnson Evangeline
Published: June 2021

Protease‐activated receptor‐3 (PAR‐3) is widely recognized as a non‐signaling elusive member of a G protein‐coupled receptor family known to regulate the endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) signaling pathway via phosphorylation of its regulatory sites. Failure of the eNOS/NO pathway leads to endothelial dysfunction, a precursor to cellular induced hypertension. Using human coronary artery endothelial cells (HCAECs) as a model for human endothelium, we demonstrated that PAR‐3 is highly co‐expressed with PAR‐1 and ‐2 and can be activated using a synthetic peptide ligand, SFNGGP‐NH2. We hypothesize that PAR‐3 can signal independently of PAR‐1 and ‐2 to regulate eNOS via site specific phosphorylation and induce proi‐nflammation via the NFkB pathway. Analysis using eNOS specific antibodies showed that 50uM of SFNGGP lead to the phosphorylation of the positive site of eNOS at ‐Ser1177 and the negative site ‐Thr495 in a time‐dependent manner. Pretreatment of HCAECs with 10uM SCH79797, a selective PAR‐1 inhibitor, did not block the increase in phosphorylation at both eNOS regulatory sites stimulated by SFNGGP. We created a stable PAR‐1 deficient cell line of HCAECs using lentiviral particles to further establish the signaling potential of PAR‐3 in the presence of thrombin and the PAR‐3 activating peptide. Our data suggests that PAR‐3 is able to regulate eNOS/NO signaling via site specific phosphorylation that is independent of PAR‐1 activation and cause pro‐inflammation with the secretion of cytokines. Understanding the signaling pathways responsible for eNOS‐induced NO and cytokine production will lead to a better understanding of endothelial dysfunction and vascular inflammation in cardiovascular diseases.
L‐Leucine Inhibits Nitric Oxide Synthesis in Endothelial Cells
Authors: Yang Ying , Meininger Cynthia , Kelly Katherine , Wu Zhenlong , Bazer Fuller , Wu Guoyao
Published: June 2021

Concentrations of L‐leucine (physiological isomer) are elevated in the plasma of obese humans as well as both diet‐induced and spontaneously obese rats in association with vascular insulin resistance. Because reduced availability of nitric oxide (NO) has been implicated to play a key role in mediating the metabolic syndrome in obese subjects, this study was conducted to test the hypothesis that elevated levels of L‐leucine may inhibit NO synthesis by endothelial cells (EC). Bovine venular EC were cultured for 48 h in Dulbecco's modified Eagle's medium containing 0.2 mM L‐arginine, 0.5 mM L‐glutamine and 0.1, 0.5 or 2 mM L‐leucine, L‐valine, L‐isoleucine, or D‐leucine. L‐Leucine dose‐dependently increased (P < 0.05) glutamine:fructose aminotransferase (GFAT) activity and the synthesis of glucosamine‐6‐P (a competitive inhibitor of glucose‐6‐P dehydrogenase, an enzyme in the pentose cycle), while inhibiting NO production by EC (P < 0.05). L‐Valine, L‐isoleucine, and D‐leucine had no effect on GFAT activity or NO synthesis in these cells (P >; 0.05). Compared with 0.1 mM L‐leucine, 0.5 and 2 mM L‐leucine reduced pentose cycle activity and intracellular concentrations of NADPH (a cofactor for NO synthase) in a dose‐dependent manner. These results suggest that L‐leucine inhibits NO production by endothelial cells through glucosamine‐mediated reduction of NADPH generation via the pentose cycle. Supported by AHA.
Circulating Porphyromonas gingivalis lipopolysaccharide induces left ventricular dysfunction through MMP‐9 regulation of inflammation
Authors: DeLeon Kristine Y , de Castro Bras Lisandra E , Zhang Jianhua , Lindsey Merry L
Published: June 2021

Periodontal disease (PD) has been correlated to myocardial infarction (MI) however the mechanism whereby PD influences remodeling of the left ventricle (LV) is not understood. Matrix metalloproteinase (MMP)‐9 regulates LV remodeling by coordinating the inflammatory and fibrotic responses. We hypothesized that low circulating levels of P. gingivalis‐lipopolysaccharide (Pg‐LPS) would induce an LV inflammatory response regulated by MMP‐9. Using echocardiography, we compared LV function in 4–7 month old wild type (WT) and MMP‐9 null mice (n≥6/group) exposed to 1 or 28 days of Pg‐LPS (1 μg/g body weight) or saline. LV function was not affected at d1. By d28, the ejection fraction of WT+LPS significantly decreased to 57±1% compared to 67±1% for saline (p<0.05). This decrease was attenuated in null+LPS (63±1%), implicating MMP‐9 as a key regulator of cardiac function. WT+LPS LV MMP‐9 expression increased 3 fold at d1 compared to d28 and saline (both p<0.05). IL‐1α plasma levels increased at d28 in WT+LPS (187±32 pg/mL) compared to saline (110±14 pg/mL, p<0.05) and null+LPS mice (102±12 pg/mL; p<0.05). LV genes, Ccl6, Ccl9, and C3, doubled in WT+LPS at d1 compared to saline, but returned to baseline by d28. In contrast, the inflammatory response in null mice was sustained until d28. In conclusion, Pg‐LPS induced significant changes in the inflammatory response in an MMP‐9 dependent manner causing a decrease in LV function.
Lysyl oxidase inhibition in the volume overloaded heart prevents adverse collagen remodeling, apoptosis, and cardiac dysfunction
Authors: El Hajj Milad C , Voloshenyuk Tetyana G , Claudino Mario A , Bradley Jessica M , Gardner Jason D
Published: June 2021

Heart failure is the most prevalent and costly disease in the U.S. Our goal was to determine if the over‐activation of lysyl oxidase (LOX), a collagen cross‐linking enzyme, accelerates cardiac disease and failure. LOX is elevated in human failing hearts, but it is not known if LOX plays a causative role in disease. Using the aortocaval fistula (ACF) rat surgical model of volume overload, we assessed the role of LOX activity in progression of heart failure over 14 wks. LOX activity was inhibited by BAPN (100 mg/kg/d) at 2 wks post‐surgery. Echocardiography was used to evaluate cardiac function and progression of LV remodeling. LOX expression and activity, collagen content, cross‐linking, and typing were determined in LV homogenates. Fixed sections of mid‐LV were assessed for apoptosis by TUNEL. ACF surgery caused significant ventricular dilatation (43% increase) and dysfunction (26% decreased, %FS). LOX protein expression was increased (65%) with concomitant increases in LOX activity. These increases in LOX were associated with significantly elevated collagen concentration, cross‐linking, and type I/III. LOX inhibition prevented ACF‐induced changes in LV collagen, and led to maintenance of systolic function. LOX inhibition also attenuated LV dilatation and prevented apoptosis, but did not reduce LV hypertrophy. These data indicate that LOX inhibition is cardioprotective in the volume overloaded heart.
Nicotine Reduced Post‐Infarct Inflammation and Improved Cardiac Output during Exercise in Conscious Mice
Authors: Lujan Heidi L. , Wang Richard , Blumberg Amanda J. , DiCarlo Stephen E.
Published: June 2021

Myocardial ischemia/reperfusion and infarction (I/R/I) have complex and lethal effects on the myocardium due, in part, to the inflammatory and reparative processes. Although inflammatory mediators are essential, an excessive inflammatory response extends the injury. Thus, there is a critically fine balance between the beneficial effects of the inflammatory response and the potential of extending the myocardial injury. The management of this balance should center on endogenous modulators of the inflammatory and reparative processes because physiological mechanisms may be the most effective and innocuous strategy. Accordingly, activation of the Cholinergic Anti‐Inflammatory Pathway (CAP) was hypothesized to be an important strategy. The CAP is an endogenous mechanism by which acetylcholine, via the α7 subunit of the nicotinic acetylcholine receptor, interacts with the innate immune system to modulate and restrain the inflammatory cascade. Thus, we hypothesized that application of the nicotine patch would reduce post‐infarct macrophage infiltration and improve cardiac function during exercise in conscious mice. We implanted a Doppler ultra‐sonic flow probe on the ascending aorta and coronary artery occluder for repeated measurements of cardiac output in conscious mice, at rest and during exercise, before and during I/R/I. The nicotine patch attenuated post‐infarct leukocyte recruitment (1005 ± 296 versus 3210 ± 460 cells/mm2), reduced infarct size (66 ± 6 versus 38 ± 11 %), and improved maximum cardiac output during exercise (13.2 ± 1.2 versus 16.4 ± 1.6 ml/min) following I/R/I in conscious mice.
Thromboxane A2 mediates apoptosis in cardiomyocytes via IP3
Authors: Touchberry Chad D , Elmore Christopher J , Srinivas Shubra , Wacker Michael J
Published: June 2021

We have shown that the inflammatory mediator thromboxane A2 (TxA2) can induce ventricular arrhythmias that were associated with increased intracellular calcium in cardiomyocytes. We were able to prevent both the arrhythmias and the changes in intracellular calcium by inhibiting inositol trisphosphate (IP3) signaling. In this investigation, we sought to determine if TxA2 receptor (TxA2R) activation would trigger apoptosis in isolated adult mouse cardiomyocytes. Our hypothesis is that TxA2R‐stimulation induces IP3 formation and subsequent IP3 receptor activation triggers apoptosis. We found that TxA2Rs were expressed in the ventricles and that there was a concentration dependent increase in cell death (Trypan blue and MTT assays) following treatment with the TxA2 mimetic U46619 (0.1–10 μM). We have confirmed that TxA2‐induced death was apoptotic via cleaved caspase‐3 and TUNEL staining. Treatment of cells with the TxA2R antagonist SQ29548 and the IP3 inhibitors Gentamycin and 2‐APB eliminated the increase in apoptosis by U46619. In conclusion, our data suggests that: 1) TxA2‐induced cell death is mediated in part by apoptosis and 2) the IP3 pathway is a novel mediator of apoptosis in cardiomyocytes. These findings may provide important therapeutic targets for inflammatory‐induced cardiac apoptosis that can lead to heart failure. (AHA 11SDG5330016 to MJW)
Identification of novel scleraxis gene targets in cardiac myofibroblasts
Authors: Bagchi Rushita , Roche Patricia , Schweitzer Ronen , Czubryt Michael
Published: June 2021

Cardiac fibrosis is a significant clinical problem yet lacks effective therapies. We have shown that the transcription factor scleraxis is a key regulator of collagen 1α2 gene expression in cardiac myofibroblasts. We now identify novel scleraxis gene targets via gain‐/loss‐of‐function. Scleraxis over‐expression in primary cardiac myofibroblasts decreased the expression of matrix remodeling enzymes matrix metalloproteinases 9 and 11 and tissue inhibitor of metalloproteinases 3. Expression of the proteoglycans versican and decorin were decreased following scleraxis overexpression, yet we observed no change in decorin and a loss of versican in scleraxis null hearts. The expression of thrombospondin 4 and the myofibroblast marker α‐smooth muscle actin (αSMA) was increased by scleraxis overexpression and strongly down‐regulated in scleraxis null hearts, and may thus represent direct scleraxis gene targets. Our results show that scleraxis exerts wide‐ranging effects on extracellular matrix‐related gene expression, with the effect on αSMA expression suggesting that scleraxis may contribute to the fibroblast‐to‐myofibroblast transition that accompanies cardiac fibrosis, further supporting our general hypothesis that scleraxis represents a novel therapeutic target for this disease.

Supported by the Canadian Institutes of Health Research & the St. Boniface Hospital Research Foundation.
Activation of proteases and changes in Na+‐K+ ATPase subunits in hearts subjected to ischemia‐reperfusion
Authors: Müller Alison L , Freed Darren H , Dhalla Naranjan S
Published: June 2021

Objective

To evaluate the role of proteases in sarcolemmal Na+‐ K+ ATPase subunits and activity these ischemia‐reperfusion (I/R) injury by subjecting rat hearts to different times of global ischemia and reperfusion.

Methods

Global ischemia was induced in rat hearts for 15, 30, and 60 minutes. I/R injury was performed by reperfusing hearts for 5, 10, 20, and 40 minutes after ischemia. Extent of degradation of Na+‐K+ ATPase subunits, α‐1, α‐2, β‐1, and β‐2 was measured in addition to activity assays for Na+‐K+ ATPase, calpain, and MMP‐ 2. Hearts were also perfused with MDL28170 or doxycycline to inhibit calpain or matrix metalloproteinase (MMP)‐2, respectively.

Results

Global ischemia depressed protein content of β‐1 and β‐2 subunits and activity of Na+‐K+ ATPase which was accompanied by augmented activities of both calpain and (MMPs) and depressed protein content of β‐1 and β‐2 subunits. Calpain and MMP‐2 activities increased whereas activity and protein content for Na+‐K+ ATPase subunits decreased upon reperfusion. MDL28170 was more effective in attenuating the I/R‐induced alterations in cardiac contracture, Na+‐K+‐ATPase activity than doxycycline.

Conclusions

These results show that activation of both calpain and MMP‐2 are involved in depressing Na+‐K+ ATPase activity and degradation of its subunits directly or indirectly in hearts subjected to I/R injury.

Funding

Manitoba Health Research Council.