bluuebunny/crossref_metadata_2025 huggingface dataset:
The role of GPR91 in the Akita model of diabetic nephropathy (DN)
Authors: Gevorgyan Haykanush Anush , Riquier-Brison Anne DM , Lam Lisa , Peti-Peterdi Janos
Published: June 2021

The importance of local tissue metabolism, succinate accumulation and signaling via its receptor GPR91 in the activation of the intra‐renal renin‐angiotensin system (RAS) in DN has been established by our laboratory using the streptozotocin (STZ) model. However, the genetic Akita (A) mouse model of type I diabetes is an excellent alternative to study DN due to lack of the toxic side effects of STZ. In this study we used wild type A (n=6) and GPR91−/− A (A−/−, n=5) mice at 18‐wks of age. Although both groups develop robust hyperglycemia, we observed noticeable decreases in water intake, food intake and urine volume in A−/− mice. Urinary albumin excretion showed 1.69±0.7 fold increase in A group compared to WT and only 1.04±0.2 increase in in A−/− mice. Immunohistochemical analyses of TGFβ found strong glomerular labeling in A but not A−/− kidneys. Whole kidney Western blots demonstrated higher TGFβ content in A mice (1.27±0.3) than A−/− (0.68±0.1). A mice revealed higher cortical renin content (1.37±0.2) than A−/− (1.0±0.1). A significant difference in medullary renin was observed between A (1.34±0.2) and A−/− mice (0.8±0.1). We concluded that GPR91 plays an important role in the pathogenesis of DN.
The renin–angiotensin system (RAS) in adaptive thermogenesis
Authors: Farah Livia Soares , Bianco Antonio Carlos , Ribeiro Miriam Oliveira
Published: June 2021

It has been reported that the inhibition of angiotensin converter enzyme (ACE) by captopril reduces body weight gain in rats. Recent studies have demonstrated that interscapular brown adipose tissue (iBAT) produces angiotensin II (ANG II), which facilitates sympathetic neurotransmission (SN). In our study we evaluated the role of iBAT in mediate the effect of captopril on body weight. For that we treated rats with 2mg/day of captopril (CAP) in the drinking water for 4 weeks. Captopril treated rats gained less weight when compared to control (105±14 vs. 82±7 CAP; p<0.01). When exposed to cold (4°C), controls and captopril treated rats kept their body temperature within normal values (35.1±1 vs. 36.3±1°C CAP). Food intake was similar for both groups, suggesting an increase in metabolic efficiency in rats treated with Captopril. Interestingly, increases in iBAT temperature after 20′ of Norepinefrine i.v. infusion was significantly higher in Captopril treated rat when compared to controls (1.54±0.1 vs. 2.14±0.2°C CAP, p>0.05). In conclusion, our data suggest that Captopril reduces body weigh by activating facultative thermogenesis through brown adipose tissue.
A unique model for evaluating obesity cardiomyopathy: Can less mean more?
Authors: Nelson Michael D. , Victor Ronald G. , Szczepaniak Edward , Simha Vinaya , Garg Abhimanyu , Szczepaniak Lidia S.
Published: June 2021

Cardiac steatosis and lipotoxicity is one proposed mechanism of obesity cardiomyopathy. Congenital generalized lipodystrophy (CGL) provides a unique opportunity to explore this relationship. Due to a generalized lack of adipocytes, we hypothesized that CGL patients would have elevated myocardial triglyceride content compared to controls. We further hypothesized that CGL patients would have reduced left ventricular (LV) function. Five patients with CGL and 5 healthy controls were recruited. Patients and controls underwent cardiac magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) to measure LV end‐diastolic and end‐systolic volumes and myocardial steatosis, respectively. LV mass was computed as the product of end‐diastolic myocardial volume and myocardial density. LV ejection fraction was calculated as a ratio between stroke volume and end‐diastolic volume. Left ventricular concentricity was calculated as a ratio between LV mass and LV end‐diastolic volume. Intra‐myocardial triglyceride levels were elevated in CGL patients compared to healthy controls (p< 0.05). LV systolic function was similar between CGL patients and healthy controls. CGL patients had elevated LV concentricity index compared to controls (P < 0.05). These data support a growing body of basic research, suggesting a mechanistic link between steatosis and lipotoxicity.
T lymphocytes infiltrating the kidney of Dahl SS rats are activated and differentiated
Authors: Rudemiller Nathan , Lund Hayley , Guo Chuanling , Flister Michael , De Miguel Camen , Mattson David L.
Published: June 2021

Previous studies demonstrated that immunosuppression attenuates Dahl SS hypertension and renal disease (Am J Physiol 298:R1136). Experiments in this study examined the infiltration of T lymphocytes in the kidney and other organs and characterized the cytokine profile of the infiltrating cells in the kidney. No significant changes in T cell number were observed in the spleen, brain, blood, or aorta of Dahl SS rats fed the low (0.4% NaCl) or the high salt (4.0% NaCl) diet (n=4–7). A significant increase in T cells was observed in the liver (from 0.8±0.1 to 2.0±0.3 × 105 cells/gram) and in the kidney (from 3.1±0.6 to 6.2±0.9 × 105 cells/gram) of Dahl SS rats fed high salt. A gene expression analysis with real‐time PCR was performed on RNA isolated from circulating T cells and T cells isolated from the kidneys of 12 Dahl SS rats (4 pooled samples of 3 individual rats each) fed high salt for three weeks. The infiltrating T cells in the kidney expressed significantly greater (by 5–54 fold) mRNA for proliferation factors (Il‐2, Il‐4, and IFN‐γ) and for cytokines and genes related to Th1 cells (TNF, Il‐10, CCr5, Csf2), Th2 cells (Il4, Il6, Il10), and Th17 cells (Il‐6 and TGFβ) compared to circulating T cells. This study shows that the inflammatory state of T cells infiltrating the kidney, which could have implications in renal disease and hypertension, is starkly different from that of the peripheral T cells.
T lymphocytes promote autoimmune‐associated hypertension
Authors: Mathis Keisa W. , Wasson Katie L. , Masterson C. Warren , Ryan Michael J.
Published: June 2021

Evidence suggests that essential hypertension has autoimmune origins. We tested whether T cells, important mediators of autoimmunity, contribute to the prevalent hypertension in the autoimmune inflammatory disorder systemic lupus erythematosus (SLE). Female mice with active SLE (NZBWF1 with albuminuria ≥ 100 mg/dL) or control mice (NZW) were treated with the T cell receptor antibody (anti‐CD3) or anti‐IgG for 3–4 weeks (intranasally; 25 μg/week). Renal cortical expression of CD3 antigen was increased in SLE mice (p=0.03) and splenic CD3+CD4+ T cells were increased in SLE mice compared to controls (19 ± 2% vs. 14 ± 1% gated, p=0.04). Spleen weight was increased in SLE mice compared to controls, and was normalized by anti‐CD3. Blood pressure (mmHg) was increased in SLE mice compared to controls (155 ± 3 vs. 112 ± 4, p=0.001), and was lower in SLE mice (137 ± 6; p=0.01) but not controls (117±2) after anti‐CD3. Albuminuria (mg/mg creatinine) was increased in SLE mice compared to controls (500 ± 290 vs. 0.05 ± 0.01; p=0.01), and was reduced in SLE mice (50 ± 37; p=0.03) but not controls (0.06 ± 0.01) after anti‐CD3. Renal cortical IL‐17 receptor expression was increased in SLE mice compared to controls, and was reduced after anti‐CD3. These data suggest that T cell‐mediated inflammation contributes to hypertension and renal injury during SLE. Support: AHA4350019 (KWM), HL085907, HL092284, HL085907S1 (MJR), HL051971 (UMMC‐Phys)
Neuronal regulation of systemic osmotic stress response mechanisms in C. elegans
Authors: Lee Elaine , Manion Dacie , Ditano Jennifer , Strange Kevin
Published: June 2021

In C. elegans, 2 osmosensory ASH neurons function to detect hypertonic stress (HS) environments and activate a behavioral avoidance response. osm‐9 and osm‐12 encode proteins essential for ASH function. Worms lacking OSM‐9 or OSM‐12 activity exhibit defective HS avoidance. A central question in stress physiology is how animals exposed simultaneously to multiple acute and chronic stressors activate appropriate defense mechanisms that maximize survival as well as reproductive fitness. We tested the role of ASH neurons in HS resistance. osm‐9 and osm‐12 mutants showed greatly enhanced survival during HS compared to wild type (WT) worms. Improved HS survival is due to decreased water loss, more rapid volume recovery, constitutive glycerol accumulation and/or activation of proteostasis mechanisms to cope with HS‐induced protein damage. Mutant and WT worms exhibited similar water loss and volume recovery during HS. No constitutive activation of the glycerol accumulation pathway was detected in osm‐9 and osm‐12 mutants. These results suggest that signals from ASH neurons normally suppress the activity of proteostasis mechanisms in a cell non‐autonomous fashion. Neuronal control of proteostasis allows animals to regulate the allocation of cellular resources in manner that coordinates and maximizes their physiological requirements with their need to cope with acute and chronic environmental stressors.
Proteostasis in C. elegans is maintained during extreme osmotic stress by reduced translation with resultant increases in molecular chaperone capacity
Authors: Burkewitz Kristopher , Choe Keith , Lee Elaine , Deonarine Andrew , Strange Kevin
Published: June 2021

Hypertonic stress (HS) causes rapid and extensive protein damage in C. elegans. Degradation of damaged proteins is essential for survival during HS. Acclimation to mild HS suppresses protein damage and increases survival under more extreme conditions. Three processes are expected to suppress protein damage in acclimated animals, 1) accumulation of the chemical chaperone glycerol, 2) upregulation of proteasome/lysosome activity, and/or 3) increases in molecular chaperoning capacity. Protein damage is unaffected by glycerol accumulation or increased protein degradation activity. Thus, changes in molecular chaperone capacity must be responsible for suppressing protein damage. Mild HS inhibits protein synthesis ~70%, which is expected to increase chaperone availability. Consistent with this idea, chronic or acute inhibition of protein synthesis by RNAi of essential translation genes or by cycloheximide suppresses HS‐induced protein aggregation 50–80%. Increases in protein production by dietary changes increase aggregation 70–180%. Our results demonstrate directly for the first time that inhibition of protein synthesis protects extant proteins from damage brought about by an environmental stressor, and challenge the widely held view that chemical chaperones are accumulated during HS to protect protein structure.
The role of carbonic anhydrase II on HCO3−‐initiated transport through the SLC4A4 transporter NBCe1A
Authors: Moss Fraser John , Boron Walter F
Published: June 2021

The carbonic anhydrase II (CAII) metabolon hypothesis predicts that CAII binds to the electrogenic Na/HCO3 co‐transporter (NBCe1), enhancing its transport of HCO3−‐related species. In 2006, using a short I–V protocol (60 ms steps from the reversal potential Erev, 20 mV increments), the Boron Lab (BL) showed that neither pure CAII injected into Xenopus oocytes expressing NBCe1 nor CAII fused to the NBCe1 C‐terminus (Ct) changed NBCe1 slope conductance (G). Yet in 2007, using long I–V protocol steps and a holding potential (Vh) tens of mV from Erev, Becker & Deitmer (BD) concluded the opposite. We hypothesized that the CAII in the cytosol (rather than that attached to the membrane) enhanced transport in the BD protocol by dissipating long‐distance HCO3− and pHi gradients—an effect that should be accentuated by long Vh steps far from Erev. We found G was unchanged even after increasing Vh steps to 30 s, whether CAII was injected in the cytosol or fused to the NBCe1‐Ct. In fact, replicating the entire BD protocol detected no significant CAII‐mediated change in peak NBCe1 current (ΔI) or G. Even when NBCe1 was driven to near‐maximal activity by setting Vh at 0 mV, CAII did not enhance the recovery rate from CO2‐induced acidification, NBCe1 ΔI or G. These data contradict the metabolon hypothesis. We continue to test if the source of CAII protein or if NBCe1A expression levels can yield systematic differences in the effect of CAII on G.
Immunocytochemical techniques identify Na+‐coupled HCO3− transporters (NCBTs) in chemosensitive neurons of the Medullary Raphé
Authors: Coley Austin Avery , Ruffin Vernon A. , Hopfer Ulrich , Boron Walter F.
Published: June 2021

The medullary raphé of the medulla oblongata contains chemosensitive neurons that sense Δ[CO2]. Serotonergic (5HT) neurons increase their firing rate in response to increases in arterial [CO2]. The GABAergic response is a decreased firing rate. Both contribute to increased pulmonary ventilation. The neuronal electroneutral Na/HCO3− cotransporters (nNCBTs) respond to intracellular acid loads (e.g. caused by increased [CO2]) by actively moving HCO3− into cells to restore the intracellular pH. We hypothesized that nNCBTs are less abundant in serotonergic and GABAergic neurons than in non‐chemosensitive neurons. A reduced nNCBT presence would be expected to slow pHi recovery, sustaining the fall in pHi and maintaining altered firing rates in 5HT and GABA neurons. An immunocytochemistry protocol was developed to distinguish cell types using monoclonal primary antibodies against MAP2, a neuronal marker; TPH, a 5HT neuronal marker; and GAD2, a GABA neuronal marker. The cell type–specific staining was colocalized with polyclonal antibodies against the nNCBTs: NBCn1, NBCn2, and NDCBE. Quantitative image analysis techniques were applied to distinguish positively stained cells from the background signals. Contrary to the hypothesis, the fraction of nNCBT‐positive cells was >80% in all cell populations for all 3 nNCBTs, and not substantially different for TPH+ vs TPH− neurons or GAD2+ vs GAD2−.
microRNA 29b is upregulated in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension and inhibits K+ channel expression and function
Authors: Pohl Nicole Monique , Yamamura Aya , Yamamura Hisao , Makino Ayako , Yuan Jason X.-J.
Published: June 2021

Downregulated voltage‐gated K+ (Kv) channels have been demonstrated in pulmonary artery smooth muscle cells (PASMC) from patients with idiopathic pulmonary arterial hypertension (IPAH). It is, however, unclear how Kv channels are downregulated in IPAH PASMC. To determine if the downregulation of K+ channels in IPAH‐PASMC could be due to miRNA regulation, we analyzed and compared human miRNA expression profile using a miRNA PCR array. Among 89 miRNAs screened, 6 miRNAs were significantly upregulated in IPAHPASMC, including miR‐29b. Real time PCR experiments confirmed the upregulation of miR‐29b in IPAH‐PASMC compared to normal PASMC. Transfection of miR‐mimic miR‐29b in normal PASMC decreased KCNA5 protein levels, while inhibition of miR‐29b in IPAH‐PASMC rescued KCNA5 protein expression. In addition, patch clamp experiments revealed that whole‐cell K+ currents are decreased in miR‐29b mimic transfected normal human PASMC, indicating that the increase in miR‐29b may be responsible for the decreased Kv currents in IPAH‐PASMC. Consistently, transfection of miR‐29b inhibitor in IPAH‐PASMC rescued whole‐cell K+ currents. In conclusion, upregulated miRNA 29b in IPAH‐PASMC might be responsible for the inhibited Kv channel expression and function and may therefore provide a new therapeutic target to treat patients with IPAH.
The urea transporter UT‐A1 is phosphorylated at serines 486 and 499 downstream of cyclic AMP production
Authors: Thai Tiffany L , Gumina Diane L , Ilori Titilayo O , Klein Janet D , Blount Mitsi A , Sands Jeff M
Published: June 2021

Urea permeability in the renal inner medullary collecting duct (IMCD) is important for urine concentration. Membrane insertion of the urea transporter UT‐A1 is regulated downstream of vasopressin (AVP)‐induced production of cyclic AMP (cAMP). UT‐A1 is phosphorylated downstream of AVP at two sites: serines 486 and 499, both of which are required for membrane insertion. We tested the hypothesis that cAMP production is required for phosphorylation of UT‐A1 at serines 486 and 499. Mouse IMCD3 cells stably transfected with UT‐A1 were treated with the adenylyl cyclase agonist forskolin and levels of phosphorylation of S486 and 499 were tested by western blot. Forskolin‐induced phosphorylation of both serines was observed with maximum effect at 5 min. Rats were fed a diet of 33% lithium for 7 days as a model of long‐term adenylyl cyclase inhibition. Kidneys were harvested and inner medullas were treated with forskolin and analyzed for total UT‐A1 as well as phosphorylation of 486 and 499. Baseline levels of UT‐A1 and phosphorylation at both sites was reduced. Whereas a strong increase in S486 and S499 phosphorylation was observed in response to forskolin in control rats, little response was seen in animals fed lithium. Overall, we conclude that cAMP produced by adenylyl cyclase results in phosphorylation of UT‐A1 serines 486 and 499 in the inner medulla. This work was supported by NIH grants T32 DK007771 and RO1 DK41707.
DIDS causes Src‐dependent inhibition of Na,K‐ATPase in the ciliary epithelium and slows aqueous humor secretion by the intact porcine eye
Authors: Delamere Nicholas A , Mandal Amritlal , Shahidullah Mohammad
Published: June 2021

Aqueous humor secretion is the result of net ion transport that involves the coordinated operation of Na,K‐ATPase in the nonpigmented ciliary epithelium (NPE) with several other basolateral transporters and channels in the ciliary epithelium bilayer. Here, we report on the ability of DIDS to reduce aqueous humor secretion and present studies to examine the mechanism.

In an intact pig eye preparation the rate of aqueous secretion was significantly reduced by DIDS added to the blood‐side perfusate. In primary cultured NPE, DIDS inhibited ouabain‐sensitive 86Rb uptake and activated Src family kinase (SFK) and ERK1/2 as judged by Western blot analysis. PP2, a SFK inhibitor, prevented the DIDS effect on 86Rb uptake, SFK and ERK1/2. DIDS was found to reduce cytoplasmic pH in BCECF‐loaded NPE. Parallel studies showed PP2 eliminates activation of SFK and ERK1/2 that occurs upon pH reduction.

The ability of DIDS to suppress aqueous humor secretion is consistent with inhibition of Na,K‐ATPase in the NPE following a drop in cytoplasmic pH and SFK and ERK1/2 activation.

Funding: NIH Grant EY06915
The trade‐off between maturation and growth during accelerated vertebrate development
Authors: Mueller Casey A , Augustine Starrlight , Kooijman Sebastiaan A. L. M , Kearney Michael R , Seymour Roger S
Published: June 2021

Developmental energetics are crucial to a species’ life history but are poorly understood mechanistically. Traditional energy and mass budgeting does not distinguish between costs of growth and maturation, making it difficult to account for accelerated development. We apply Dynamic Energy Budget theory (DEB), which exclusively considers maturation costs, to interpret empirical data on the energetics of development through metamorphosis in two related frogs, Crinia georgiana and Pseudophryne bibronii. Dry mass (2.74 in C. georgiana, 2.68 mg in P. bibronii) and energy content (70 and 65 J) of fresh ova were similar between the species. However, development to metamorphosis was 1.7 times faster in C. georgiana while P. bibronii produced nine times the dry mass at metamorphosis. DEB theory explains these differing patterns through a trade‐off in ontogenetic energy allocation between maturation and growth. P. bibronii partitioned energy in a constant fraction (0.31) throughout development whereas C. georgiana increased the fraction (0.14 to 0.39) of energy allocated to maturation over growth between hatch and the onset of feeding. The change in developmental energy allocation in C. georgiana may result from a selective pressure to increase development rate. Funding from the University of Adelaide.
Prolonged Fasting Increases Nrf2 Nuclear Accumulation and DNA Binding Ability in Postweaned Northern Elephant Seals
Authors: Vazquez-Medina Jose Pablo , Soñanez-Organis Jose Guadalupe , Viscarra Jose Abraham , Tift Michael S , Forman Henry Jay , Crocker Daniel E , Ortiz Rudy Martin
Published: June 2021

Northern elephant seals naturally experience prolonged periods of absolute food and water deprivation (fasting) while breeding, molting and weaning. Prolonged fasting in elephant seals promotes insulin resistance, activates the renin‐angiotensin system and increases NADPH oxidase activity and protein expression without increasing oxidative damage or inflammation. Prolonged fasting also stimulates the antioxidant system of the elephant seal likely resulting in the prevention of oxidative damage. The redox‐senstive transcription factor Nrf2 is the main regulator of the adaptive response to oxidative stress. Nrf2 translocates into the nucleus and increases the transcription of antioxidant genes in response to intracellular oxidant production. To test the hypothesis that Nrf2 is activated in response to prolonged fasting in elephant seals, angiotensin II (Ang II) and TGF‐β circulating concentrations, along with NADPH oxidase 4 (Nox4) mRNA expression and Nrf2 nuclear content and DNA binding ability, were measured in blood and muscle samples collected from seals fasting for 1, 3, 5 and 7 weeks. Ang II and TGF‐β increased gradually through the fasting and where 40‐fold (Ang II) and 2‐fold (TGF‐β) higher at wk 7 than at wk 1. Nox4 was 5‐fold higher at wk 7 than at wk 1 while Nrf2 nuclear content and DNA binding ability were 2‐fold higher. Results show that prolonged fasting activates the adaptive response to oxidative stress in elephant seals and suggest that Ang II and TGF‐β stimulate Nox4, which may potentially mediate a hormetic protective response in fasting seals.
Sphingolipid Ratios in Marine Mammal Skeletal Muscle Cell Membranes:Protection from Oxidative Stress?
Authors: Young Kathryn , Trumble Stephen
Published: June 2021

Living cells are often exposed to dramatic changes in environmental conditions and adapt to these changes by expression of inducible enzyme or signal transduction pathways that provide an appropriate response to the environmental signals. In marine mammals, these extreme changes in environment (a stress stimulus) may be induced by deep or prolonged diving. However, the physiological systems activated by stress stimuli often not only protect the cell but may also prove damaging. This dual pathway is evident in the sphingolipids, ceramides and sphinosine‐1‐phosphate (S‐1‐P), which are known to harm (apoptosis) and protect cells (from oxidative stress), respectively. Regulation of these seemingly contradictory functions must operate on multiple pathways to ensure an adequate homeostasis. Therefore, the ratio of ceramides to sphingosine‐1‐phosphates, in particular, may be a useful prediction of stress at the cellular level. Sphingolipids in marine mammal tissue have never before been characterized; therefore, it was our goal to establish ratios of ceramides to S‐1‐P in several species of diving mammals. We analyzed muscle tissue from 5 species of marine mammals (Pusa hispida, Phoca largha, Callorhinus ursinus, Eumetopia jubatus, and Odobenus rosmarus) as well as a control (Mus musculus) and found significantly different ratios of ceramides to S‐1‐P in marine mammals correlating with diving capacity. These differences may indicate that increased oxidative stress endured by cell membranes necessitate increased S‐1‐P as a protective mechanism. Additionally, marine mammals typically donˈt suffer from the metabolic diseases often associated with increased ceramide production, lowering the ceramide:S‐1‐P ratio that much further. This first comparative account of sphingolipid ratios in marine mammals may link levels of oxidative stress to dive depth and duration, an important environmental variable in the life history of all marine mammals.
In vitro effects of acute exposure to hyperglycemic conditions on endothelium‐dependent vasodilation in mourning doves (Zenaida macroura)
Authors: Jarrett Catherine L , Sweazea Karen L
Published: June 2021

Normal avian plasma glucose levels are 1.5–2 times greater than mammals of similar size. In mammals, hyperglycemia leads to endothelial dysfunction and impaired vasodilation. The hypothesis was that endothelium‐dependent vasodilation of isolated avian arteries would not be impaired following acute exposure to high glucose. Isolated small resistance saphenous arteries (80–150μm, inner diameter) were cannulated and pressurized in a vessel chamber then incubated with a ringer solution containing either normal or high glucose (20mM vs. 30mM) for 1h at 41°C. Vessels were then pre‐constricted to 50% of resting inner diameter with phenylephrine (PE) and exposed to increasing doses of acetylcholine (ACh; 10−9 to 10−5 M, 5 mins per step). Percent reversal of PE‐induced tone was measured by tracking the inner diameter with edge‐detection software. Contrary to our hypothesis, ACh‐induced vasodilation was impaired with acute exposure to high glucose (p=0.013). The impairment was not related to increased osmolarity since vasodilation of arteries exposed to a combination of 20mM glucose and 10mM mannitol was not different from controls (p=0.898). Pre‐exposure of high glucose arteries to the superoxide dismutase mimetic tiron (10mM) improved vasodilation (p<0.05). Therefore, dove arteries do not appear to have endogenous mechanisms to prevent impaired vasodilation resulting from high glucose
The effects of Apelin‐13 on cardiovascular system can be mediated by modulating the excitability of SFO neurons
Authors: Dai Li , Smith Pauline M , Ferguson Alastair V
Published: June 2021

Apelin is an adipocyte derived hormone involved in the regulation of water balance, food intake, and the cardiovascular system via actions in the central nervous system. The subfornical organ (SFO) is a circumventricular organ with well document roles in body fluid homeostasis, cardiovascular control and more recently, energy balance. Owing to the lack of the normal blood‐brain barrier, SFO enables detecting circulating signal molecules from blood and cerebrospinal fluid. In this study, we have examined the effect of apelin on blood pressure (BP) and heart rate (HR) of anesthetized SD rats by microinjection of apelin into SFO in vivo and on the excitabilities of dissociated SFO neurons in vitro. We found that microinjection of apelin into SFO decreased BP (‐ 1492.3 ± 357.1 mmHg*sec, n=5) and HR (−32.4 ± 10.39 beats, n=5). Whole cell current clamp recording revealed that 100 nM aplein caused a hyperpolarization in 41 % of the tested SFO neurons (−9.9 ± 1.3 mV, n=21), while 33 % of neurons were depolarized by apelin administration (6.8 ± 0.9mV, n=17). Our research suggests that circulating apelin can directly affect BP and HR via modulating the excitability of SFO neurons.
Neonatal Bladder Inflammation Reduces Stress‐Induced Opioidergic Inhibition of Visceromotor Responses to Urinary Bladder Distension in Adult Rats
Authors: Ness Timothy J , DeWitte Cary R , Robbins Meredith T
Published: June 2021

Objective

The present study sought to determine the effect of the opioid antagonist naloxone on stress‐induced analgesia and stress‐induced hyperalgesia (SIH) mechanisms in adult rats which experienced bladder inflammation as neonates.

Methods

Female rat pups (P14–16) were anesthetized and treated with intravesical 1% zymosan (ZY− rats). Control pups were only anesthetized (AN− rats). As adults, rats were separated into acute footshock (‐FS‐; 15 min, 1 mA, 1 S duration) or non‐footshock (‐NFS‐) groups & further subdivided into naloxone‐treated (1 mg/kg; ‐NAL) and saline‐treated (−SAL) subgroups. These rats were anesthetized immediately following FS or NFS treatments, drugs given i.p.and visceromotor responses (VMR) to phasic, graded urinary bladder distension (UBD) determined.

Results

There was no significant effect of naloxone treatment on UBD‐related SIH observed in ZY‐rats (VMRs of ZY‐FS‐NAL rats = ZY‐FS‐SAL rats > ZY‐NFS‐NAL rats = ZY‐NFS‐SAL rats). In contrast, naloxone treatment revealed a reactive opioidergic suppression of SIH in AN‐rats (VMRs of AN‐FS‐NAL rats > other AN− groups).

Conclusions

These data support the assertion that neonatal bladder inflammation results in the failure of an opioidergic inhibitory system which is activated by acute stress, the results of which is magnification of the concurrent pronociceptive processes related to visceral SIH. Supported by DK51413
EXERCISE TRAINING INDUCES CHANGES IN THE RENIN‐ANGIOTENSIN SYSTEM TOWARDS ANG‐(1–7)/ACE2/MAS AXIS IN THE HYPOTHALAMUS OF RENOVASCULAR HYPERTENSIVE RATS
Authors: Cangussu Luiza Michelle , Oliveira Luzia M , Silva Jose R , Alzamora Andreia C , Santos Robson AS , Campagnole-Santos Maria J
Published: June 2021

In the present study we evaluated angiotensin peptide levels, mRNA expression of angiotensin receptors, AT1 and MAS, and the enzymes, ACE and ACE2 in the hypothalamus (Hy) in renovascular hypertensive rats (2K1C) submitted to exercise training (ET). One week after renal artery constriction, animals started ET (swimming 60 min/day, 5 days/week). After 4 weeks, mean arterial pressure and heart rate (telemetry method) of 2K1C‐ET (122±6 mmHg and 349±5 beats/min; n=5) were lower in relation to 2K1C (157±3 mmHg and 361±5 beats/min; n=4). 2K1C showed an increase in AT1 expression and Ang II levels accompanied by a reduction in Ang‐(1–7) levels in Hy, which significantly reduced the Ang‐(1–7)/ Ang II levels (16±2 vs 40±10 in Sham). No changes in ACE or ACE2 were observed. ET induced in Hy an increase in ACE (2.4±0.3 au vs 1±0.1 au in Sham), ACE2 (3.7±0.4 au vs 1±0.1 au in Sham) and Mas (3.2±0.4 au vs 1±0.1 ua in Sham). These changes were accompanied by a reestablishment of Ang peptides ratio in the Hy. Our data show that ET may induce selective changes in the brain RAS favoring the ACE2/MAS axis, at least in the hypothalamus, suggesting a central site involved in the anti‐hypertensive effects induced by ET.
Cardiovascular effects of Angiotensin peptides are differentially mediated at the CVLM of hypertensive rats
Authors: Sousa Graziele G , Castro Uberdan GM , Silva Marcelo E , Santos Robson AS , Campagnole-Santos MariaJose , Alzamora Andreia C
Published: June 2021

In the present study we investigated the effect of the non‐selective nitric oxide (NO) synthase inhibitor, L‐NAME, or the antioxidant vitamin C (Vit C) on the cardiovascular effects triggered by angiotensin (Ang) II and Ang‐(1–7) at the caudal ventrolateral medulla (CVLM) of renovascular hypertensive rats (2K1C). CVLM microinjection of L‐Name induced similar fall in blood pressure in Sham or 2K1C, while Vit C produced a fall in BP (− 19±4 mmHg, n=5) only in 2K1C rats. L‐NAME did not alter the hypotensive effect induced by CVLM microinjection of Ang II in Sham or 2K1C. However, L‐NAME increased the hypotensive effect of Ang‐(1–7) in Sham and did not alter its effect in 2K1C. In contrast, Vit C abolished the hypotensive effect of Ang II in the CVLM of both groups and did not change the hypotensive effect of Ang‐(1–7) in the CVLM of 2K1C (−12±1 mmHg, n=6) or Sham (− 12±1 mmHg, n=5). These data suggest that different mechanisms are involved in the cardiovascular effects of Ang peptides at the CVLM. Further, our results suggest that part of these mechanisms may be impaired in 2K1C hypertension.
Mitogen‐Activated Protein Kinase Mediates the Effects of Chemokine Stromal Cell‐Derived Factor‐1 (SDF‐1) on Cardiovascular Function and Sympathetic Drive in Rats
Authors: Wei Shun-Guang , Zhang Zhi-Hua , Yu Yang , Felder Robert B.
Published: June 2021

The chemokine SDF‐1 and its receptors are expressed by neurons and glial cells in cardiovascular autonomic regions of the brain. SDF‐1 expression increases in these regions in rats with heart failure (HF). Intracerebroventricular (ICV) administration of SDF‐1 elicits increases in mean blood pressure (MBP, mmHg), heart rate (HR, bpm) and renal sympathetic nerve activity (RSNA) that are exaggerated in rats with HF. SDF‐1 is known to activate mitogen‐activated protein kinase (MAPK) intracellular signaling. We hypothesized that p44/42 MAPK, which has recently been implicated in sympathetic activation in HF, mediates the sympathetic and hemodynamic responses to SDF‐1. In urethane anesthetized adult male rats, ICV injection of SDF‐1 (100 ng) induced significant (p<0.05) increases in MBP (27.6 ± 3.5), HR (55 ± 6) and RSNA (90.1 ± 6.8 %). Pretreatment with ICV PD98059 (1 μg), a selective p44/42 MAPK inhibitor, prevented these SDF‐1 induced responses. ICV minocycline (20 μg), an inhibitor of glial cell activation, significantly (p<0.05) attenuated but did not block the SDF‐1 induced increases in MBP (11.2 ± 2.4), HR (27 ± 4) and RSNA (38.8 ± 4.2 %). These data identify p44/42 MAPK signaling as an obligatory intracellular mechanism mediating the cardiovascular and sympathetic responses to SDF‐1. MAPK signaling in both neurons and glial cells may contribute to these responses. Supported by NIH RO1 HL073986 & HL096671.
Acute potentiating effects of tumor necrosis factor‐α (TNFα) on the responses of rat vagal pulmonary sensory neurons to capsaicin challenge
Authors: Hsu Chun-Chun , Geer Marcus , Lin You Shuei , Lee Lu-Yuan
Published: June 2021

TNFα is a proinflammatory cytokine, and plays an important role in the pathogenesis of allergic asthma. A recent study in our lab has demonstrated that a prolonged (24–48 h) treatment with TNFα increased the Ca2+ influx evoked by TRPV1 activators in vagal pulmonary sensory neurons (Am J Physiol 299: 483–492, 2010). The present study was carried out to test the acute effect of transient TNFα treatment on the TRPV1 sensitivity in isolated rat pulmonary sensory neurons. Our results showed: 1) A brief treatment with a low concentration of TNFα (1.44 nM; 9 min) induced a biphasic TRPV1 hypersensitivity in these neurons: the inward current evoked by capsaicin (average concentration 100 nM; 3 s) was enhanced both immediately following the TNFα treatment (Δ=280%), and after a long delay (peak at ~90 min; Δ=382%). 2) The immediate phase of the potentiating effect of TNFα was almost completely abolished by pretreatment with a COX‐2 inhibitor, NS‐398 (10 μM; 10 min), whereas the delayed phase was only partially attenuated. 3) In contrast, the same TNFα treatment did not generate any potentiating effect on the response to ATP (0.3 – 1.0 μM) challenge in these neurons. In summary, a brief treatment with TNFα induced both immediate and delayed potentiating effects on the TRPV1 sensitivity in isolated pulmonary sensory neurons, and a production of COX‐2 arachidonic acid metabolites is probably involved in the immediate sensitizing effect of TNFα. (Supported in part by NIH grant HL96914 & NSC98‐2917‐I038‐101 fellowship from ROC)
Role of group 1 metabotropic glutamate receptors (mGluR) in the hypothalamic paraventricular nucleus (PVN) in sympathetic and blood pressure control during water deprivation
Authors: Holbein Walter , Bardgett Megan , Andrade Mary Ann , Herrera-Rosales Myrna , Calderon Alfredo S , Toney Glenn M
Published: June 2021

Compared to WKY rats, sympathetic nerve activity (SNA) and elevated arterial pressure (AP) in spontaneous hypertensive rats (SHR) is supported by enhanced glutamatergic drive of group 1 mGluR (mGluR1 & mGluR5) in the PVN. Whether enhanced group 1 mGluR activity is a genetically determined feature of sympathetic control unique to SHR or is a mechanism of PVN activation common to other hypertensive states is undetermined. To investigate this, euhydrated (EU) and 48 h water deprived (WD) rats were studied, as dehydration also acutely increases AP by increasing glutamatergic drive of the PVN. In conscious EU rats, injection of the group 1 mGluR agonist DHPG (2 nmol in 200 nl) increased AP (n=3, p<0.05). This effect was attenuated by prior PVN injection of the mGluR1 antagonist LY‐367385 (4 nmol in 200 nl; n=3, p<0.05), but not the mGluR5 antagonist MPEP (4 nmol in 200 nl; n=2). Injections of LY and MPEP alone were each without affect in EU (n=2) and WD rats (n=3). Similar results were obtained in EU (n=3) and WD (n=2) rats under anesthesia in which renal SNA was also unaffected by blockade of PVN group 1 mGluR. Thus, whereas activation of PVN group 1 mGluR is sympathoexcitatory and contributes to sympathetic support of ongoing AP in SHR, the latter appears not to be a common mechanism of PVN driven support of SNA or elevated AP, as no tonic role for PVN group 1 mGluR activity was observed either in EU or 48 h WD rats. HL102310 and HL088052 (GMT)
Carotid body denervation does not affect CO2 sensitivity in multiple rat strains
Authors: Mouradian Gary Charles , Miller Justin , Muere Clarissa , Forster Hubert V. , Hodges Matthew R.
Published: June 2021

O2 and CO2/pH are detected by the carotid bodies (CBs) and central respiratory chemoreceptors, respectively. CB activity appears to modulate central chemoreceptor activity, suggesting chemoreceptor interdependence. Here, we studied the effects of CB denervation in CO2‐insensitive Brown Norway (BN) and CO2‐sensitive Dahl Salt‐sensitive (SS) and Sprague Dawley (SD) rats by chronic measurements of ventilation (VE) and blood gases at rest, and during hypercapnia (7% CO2) or hypoxia (12% O2) before and for 21 days (d) after CBD. Peak eupneic hypoventilation (Δ PaCO2) occurred 1–2 d post‐CBD in all strains, increasing 11.0 (SS), 9.4 (BN) and 7.0 (SD) mmHg. Resting PaCO2 returned to control by 12 d post‐CBD (BN and SD), but remained elevated (+3.9 mmHg) in SS rats. SS and SD rats showed high sensitivity (VE ~300% of control) and BN rats a low sensitivity (VE ~150% of control) to hypercapnia pre‐CBD, which was unaffected by CBD. In addition, CO2 sensitivity (measured as ΔVE/ΔPaCO2) was also unaffected by CBD in all strains, despite a near‐complete loss of the hypoxic ventilatory response and attenuation of the ventilatory response to i.v. NaCN. As seen in other species, these data suggest that CBD in rats leads to eupneic hypoventilation and elimination of the hypoxic ventilatory response, but the lack of effect on CO2 sensitivity does not support the concept of chemoreceptor interdependence. Supported by NIH HL097033.
Functional expression of calcium sensing receptor in rat vagal pulmonary sensory neurons
Authors: Gu Qihai , Kagira Martin , Gilbert Carolyn
Published: June 2021

The calcium sensing receptor (CaSR), first isolated from bovine and human parathyroid, is a G‐protein‐coupled receptor that monitors the systemic, extracellular, free ionized‐calcium level in organs involved in systemic Ca2+ homeostasis. Recent studies have shown that CaSR is also expressed in many tissues that are not directly related to the plasma Ca2+ regulation, such as central and peripheral nervous systems where the function of this receptor remains to be defined. In this study, we examined the functional expression of CaSR in rat vagal pulmonary sensory neurons. Our immunohistochemical experiments demonstrated the expression of CaSR in these sensory neurons, as well as in the trachea and lungs. Whole‐cell patch‐clamp recordings were performed in isolated pulmonary sensory neurons. The results showed that pretreatment with NPS 2390 or NPS 2143, a selective CaSR antagonist, significantly enhanced the capsaicin‐evoked whole‐cell inward current. These data suggest that CaSR is likely involved in the integration of primary pulmonary sensory inputs under physiologic and/or pathophysiologic conditions. Funded partially by AHA 0835320N and MUSM startup funds.
“Full term” prenatal nicotinic exposure (fPNE) produces a depressed hypoxic ventilatory response (dHVR) in postnatal rats
Authors: Xu Fadi , Zhuang Jianguo
Published: June 2021

Maternal smoking is the key risk factor and hypoxemia is an acute precursor for SIDS. An existing SIDS rat pup model induced by a traditional PNE (tPNE) covering the last 2/3 of the gestation period shows an excessive mortality (15%) during 60 min severe hypoxia. We tested if fPNE consisting of the periods immediately before pregnancy and during pregnancy and lactation would depress HVR, leading to a higher mortality than tPNE. After detecting the metabolism and collecting blood sample, awake P11‐14 rats pretreated with vehicle (Ctrl), tPNE, and fPNE were individually placed in a plethysmograph and exposed to 5% O2 for 60 min with the rectal temperature at ~34.0°C. We found that fPNE induced a dHVR 38 ± 4% (P < 0.01) lower than tPNE and Ctrl pups without effects on baseline VE, HR, metabolism, body weight and temperature, blood gases, and pH. Dramatic bradycardia associated with apneas and gasps occurred 35 ± 3.3 min after hypoxia, which led to a lethal VE arrest 42 ± 2.7 min after hypoxia with heart beat persisting for additional several min in 10/16 fPNE pups (63% mortality). Neither the dHVR nor the cardiorespiratory failure (death) was observed in all Ctrls and 6/8 tPNE pups during this period. Two remaining tPNE pups died (25% mortality) ~54 min after hypoxia following a less dHVR. Our results suggest that fPNE with a higher mortality and worse dHVR is a more suitable SIDS model than tPNE (Supported by HL 107462 and ALA RG‐191095‐N).
Some Morphologic and Growth Features of Juxta‐alveolar Smooth Muscle Cell Lines from High Altitude Hypoxia Sheep
Authors: John Theresa Adebola
Published: June 2021

OBJECTIVES

We derived smooth muscle cells (SMCs) from lung terminal parenchyma and studied the effect of high altitude hypoxia (HAH) on their monolayer morphology and growth profiles.

METHODS

Lungs were obtained from adult sheep kept at 12, 470 ft altitude, PaO2 ~60 Torr for ~4 months and age matched control sheep kept at sea level. SMCs migrated from terminal lung parenchyma were cultured as monolayers, characterized by alpha‐smooth muscle actin immunofluorescence, and studied for monolayer morphologies and tritiated (3H) thymidine incorporation growth profiles.

RESULTS

HAH‐SMC morphologic varieties were: apoptotic, sub‐confluent, spindle‐form, hill and valley, swirl‐form, and rhomboid monolayers and control SMC varieties were swirl and hill‐and‐valley formations at confluence, jointly from vascular and bronchiolar termini. The normal growth pattern, typified by intrapulmonary 7th generation vein SMCs, showed mean peak 3H‐thymidine incorporation of 2829.46±294.24 counts per minute (cpm) at 72h. Comparatively, HAH‐SMCs profiles were varied: normal, subprolific, prolific, retarded, and senescent. At extremes, mean peak cpm by prolific cells were 7775.42±475.77 and by senescent cells were 26.71.

CONCLUSION

Chronic HAH caused morphologic and functional changes, possibly combined pathologic formations and adaptive responses, in juxta‐alveolar SMCs.

NHLBI MIRS HL‐059435 to T. A. John
Hypersensitivity of lung vagal C fibers induced by acute intermittent hypoxia in rats: role of reactive oxygen species and TRPA1
Authors: LAI CHING JUNG , SHEN MEI-YA , LUO YA-LING , YANG CHUNG-HUAN
Published: June 2021

Obstructive sleep apnea, manifested by intermittent hypoxia and excess production of reactive oxygen species (ROS) in the airways, is associated with hyperreactive airway diseases, but the mechanism remains unclear. Sensitization of lung vagal C fibers (LVCFs) contributes to the airway hypersensitivity. We investigated the mechanisms underlying the sensitization of LVCFs by acute intermittent hypoxia (AIH) for ten episodes of exposure to 30 s of hypoxic air (0, 5 or 10%O2) followed by 30 s of room air in anesthetized, open‐chest, and artificially ventilated rats. Our electrophysiological study revealed that the afferent response of LVCFs to intravenous capsaicin (a LVCF stimulant) was concentration‐dependently augmented by AIH. Similarly, the LVCF responses to α,β‐methylene‐ATP (α,β‐meATP; another LVCF stimulant) was augmented by AIH (0%O2). The AIH‐induced sensitization of LVCFs was prevented by pretreatment with dimethylthiourea (DMTU; a hydroxyl radical scavenger), N‐acetyl‐L‐cysteine (NAC; an antioxidant) or HC‐030031 [a transient receptor potential ankyrin 1 (TRPA1) receptor antagonist]. Contrastingly, AIH failed to alter the afferent response of LVCFs to mechanical stimulation by lung hyperinflation. Our results suggest that AIH sensitizes LVCFs to chemical stimulants in rats possibly via action of ROS and activation of TRPA1 receptors.
Chronic Intermittent Hypoxia (CIH) alters respiratory rhythmogenesis within the preBötzinger Complex
Authors: Garcia Alfredo J , Doi Atsushi , Malashchenko Tatiana , Wei Aguan , Kumar Ganesh , Prabhakar Nanduri , Ramirez Jan-Marino
Published: June 2021

CIH is an established model simulates many changes in cardio‐respiratory physiology associated with sleep disordered breathing. Here we show that CIH significantly altered cellular and molecular properties within the preBötzinger complex (preBötC), a central network critical for respiratory rhythmogenesis. In neonatal mice exposed to CIH (10days), lipid peroxidation was increased in the preBötC, and extracellular and intracellular recordings from brainstem slices of CIH treated mice revealed functional changes to respiratory rhythm generation. During fictive eupnea burst to burst variability in amplitude and frequency is significantly increased while during hypoxia, fictive gasping is strengthened as shown by larger population burst amplitudes and faster recoveries following reoxygenation. CIH affects the network response to noradrenergic and cholinergic modulation as evidenced by altered responses to (1) receptor agonists and (2) selective optogenetic stimulation of cholinergic and TH‐containing neurons. Moreover, both in vivo and in vitro data suggest respiratory rhythmogenesis after acute intermittent hypoxia can become destabilized following CIH exposure. These findings indicate that CIH alters functional properties of the central respiratory network which may contribute to the cardio‐respiratory changes found in sleep disordered breathing.
Colocalization of angiotensin converting enzyme 1 and FosB in the median preoptic nucleus (MnPO) following intermittent hypoxia
Authors: Faulk Katelynn E. , Knight W. David , Little Joel T. , Cunningham J. Thomas
Published: June 2021

Chronic Intermittent Hypoxia (CIH) is a model for the hypoxemia related to sleep apnea and is associated with hypertension and increased sympathetic tone. In rats, the MnPO demonstrates increased FosB expression following CIH and virally mediated inhibition of MnPO FosB attenuates sustained CIH hypertension. Our previous work identified Angiotensin Converting Enzyme I (ACEI) as a possible downstream target of MnPO FosB. The purpose of this study is to further characterize the expression of ACEI in the MnPO of CIH rats. Rats were exposed to CIH for 7 days through 3 minute periods of hypoxia (10% oxygen) and 3 minute periods of normoxia (21% oxygen) for 8 hours per day. Controls were not exposed to CIH but housed under similar conditions. Rats were then anesthetized and fixed through paraformaldehyde perfusion and their forebrains were later processed for immunohistochemistry. Antibodies used were rabbit polyclonal ACEI (1:500) and goat polyclonal FosB (1:5000). The sections were imaged using an Olympus DSU confocal microscope. In the MnPO, CIH exposure increased FosB positive cells (Control 2.3± 0.7; CIH 20.2±1.7; P<0.001) and colocalization of FosB with ACEI positive cells (Control 0.7±0.4; CIH 12.9±2; P<0.001) but failed to show any significant difference in the number of ACEI positive cells. These results confirm that ACEI positive cells in the MnPO are transcriptionally activated by CIH. P01 HL88052