bluuebunny/crossref_metadata_2025 huggingface dataset:
Regulation of hypoxic responses by Sprouty2 in hepatocellular carcinoma
Authors: Hicks Kristin C , Patel Tarun B
Published: June 2021

Sprouty2 (Spry2) is an inhibitor of receptor tyrosine kinases (RTKs) and decreases in its levels have been documented in breast, lung, liver, and prostate cancer. Decreases in Spry2 levels have been correlated with poor patient prognosis in hepatocellular carcinoma (HCC). During tumor growth, cells experience hypoxic periods and adapt to these by altering a variety of cellular processes mainly through the Hypoxia Inducible transcription Factors (HIFs). HIFs are composed of an α subunit (HIF1α or HIF2α) and a β subunit (HIF1β a.k.a. aryl hydrocarbon receptor nuclear translocator (ARNT)). In hypoxia, the HIFα subunits are stabilized, translocate to the nucleus, and dimerize with ARNT. The HIFs have been shown to be upregulated in cancers correlating to poor patient prognosis. We have shown that Spry2 protein levels are elevated in hypoxia. The objective of our study was to determine the role of Spry2 in regulating hypoxic responses in HCC cells as it could have an important impact in tumor growth and progression. We found that endogenous Spry2 by modulating ARNT protein levels alters the transcriptional activity of HIF. Thus, our data suggest that Spry2 regulates HIF activity in HCC cells.

Supported by NIH grant GM 073181
Cannabinoid 1 receptor as therapeutic target in preventing chronic epilepsy
Authors: Di Maio Roberto
Published: June 2021

Objective

Temporal lobe epilepsy (TLE) is the most common form of refractory epilepsy. Cannabinoid type 1 (CB1) receptor regulates neuronal excitability and has been shown to mediate the anticonvulsant effects of cannabinoids in acute animal models of seizure. However, the potential of cannabinoids for preventing chronic epileptic damage as consequence of an inciting event (acute seizure) remains under explored.

Methods

We investigated the neuroprotective role of the CB1 receptor against the hippocampal damage occurring in a chronic pilocarpine (PILO) model of TLE in rats. We simulated a therapeutic approach by administering the CB1 agonist WIN55.212–2 to rats for 15 days, beginning 24 hours after the induction of acute epileptic syndrome with PILO.

Results

After 6 months, the animals treated with WIN55.212–2 for 15 days following PILO administration, showed (relative to vehicle control): decreased epileptic behavior, reduced abnormal fiber sprouting in the dentate gyrus, preservation of GABA‐ergic neurons, less oxidative injury through NADPH oxidase activation, and normalization of CB1 receptor distribution and expression. Taken together, these data suggest that novel compounds targeting CB1 receptor or downstream signaling pathways may be efficacious in the prevention of chronic TLE.

Interpretation

Given the prevalence and severity of TLE, there is great potential of such compounds.
Effects of 5‐fluorouracil and scopolamine in three variations of the autoshaping procedure
Authors: Walker Ellen Ann
Published: June 2021

The experiments in the present study examined whether 5‐ fluorouracil (5‐FU) or scopolamine would be more disruptive to the Instrumental or Pavlovian conditioning aspects of the autoshaping procedure. We compared the effects of saline, scopolamine or 5‐FU in the traditional autoshaping procedure with both Pavlovian and Instrumental components (autoshaping‐PI) to an autoshaping procedure with predominantly Instrumental contingencies (autoshaping‐I) and an autoshaping procedure with predominantly Pavlovian associations (autoshaping‐P). Mice were presented a tone on a variable‐time schedule, with the tone remaining on for 6 sec or until a nose‐poke response occurred (autoshaping‐PI; autoshaping‐I). If no response was made during the tone, the dipper was presented at the end of the 6 sec period (autoshaping‐PI; autoshaping‐P). Mice acquired autoshaped responses in all three procedures, although acquisition occurred more quickly in autoshaping‐PI. Responding in the autoshaping‐PI and autoshaping‐I procedures revealed more disruptions after injections of 5‐FU and scopolamine demonstrating either the increased sensitivity of these two procedures to drug effects.
Use of novel pharmacological agents to assess the role of Rho GTPase regulatory proteins in cognition
Authors: Martinez Luis , Tejada‐Simon Maria Victoria
Published: June 2021

Cognitive impairment is thought to persist in certain neurological disorders due to abnormal neuroplasticity, the ability of neuronal connections to undergo experience‐dependent modifications. These changes, used to process and accommodate new information, require reorganization of the actin cytoskeleton within dendritic spines, a process that is known to be regulated by the Rho family of small GTPases. In an effort to further define the involvement of small GTPases in cognition, the Rho GTPase Rac1 was pharmacologically targeted in vivo using novel agents that inhibit its activation. Mice treated intraperitoneally with different concentrations of Rac1 inhibitors were subjected to specific behavioral tasks that examine contextual (associated fear conditioning) and spatial memory (Morris water maze). Performance on these tasks is dependent on the neuronal integrity of the hippocampus. Overall, our results showed that mice treated with a Rac1 inhibitor displayed attenuated learning and impaired recall. These results complement previous findings in our laboratory demonstrating, through in vitro electrophysiology studies, that either genetic deletion or pharmacological inhibition of Rac1 in the mouse hippocampus disrupts several forms of long‐term synaptic plasticity. Taken together, these results support a role of Rac1 in cognitive function and memory formation.
Hyperplasticity in the amygdala of rat Model by prenatal exposure to valproic acid
Authors: Lin Hui‐Ching , Chan Yun‐Han , Chen Po See
Published: June 2021

Amygdala is the brain area involved in socio‐emotional behavior. However, the role of amygdala in autism remains inconclusive. Here, we used 28–35 days valproate (VPA)‐induced rat model of autism to observe the autistic phenotypes and evaluate their synaptic characteristics in the lateral nucleus (LA) of the amygdala. The VPA‐treated offspring demonstrated less social interaction and increased anxiety. Slice preparation and electrophysiological recordings of the LA were performed in the VPA‐treated offspring. The magnitude of long term potentiation (LTP) was significantly higher in the VPA‐exposed than saline‐exposed offsprings. These results indicated the enhancement of synaptic plasticity in the LA might contribute to autism in the VPA‐treated offspring.
The malin‐laforin compex modulates glycogen catabolism through its effect on glycogen phosphorylase
Authors: Dukhande Vikas V , Paasch Bradley , Gentry Matthew S
Published: June 2021

Autosomal recessive mutations in genes encoding malin or laforin cause Lafora disease (LD). LD is a progressive myoclonic epilepsy characterized by pathological aberrant glycogen accumulations in brain and various other organs. Laforin is a novel glycogen phosphatase essential for maintaining glycogen in a soluble form. Malin, a RING‐type E3 ubiquitin ligase, is known to ubiquitinate proteins involved in glycogen metabolism. However, malin's role in LD pathogenesis is not clear. In this study, we aimed to discover malin substrate and define their function in LD pathogenesis. We used malin knockout and laforin knockout mice, immunoprecipitation, western analysis, immunocytochemistry, glucan‐binding assay, PAS staining and enzyme activity measurements. We discovered glycogen phosphorylase (GP) as a novel substrate of malin. GP is vital in catabolism of cellular glycogen. The expression of GP was decreased in liver and skeletal muscle tissues from malin knockout mice. Our data suggest that malin does not target GP for proteasomal degradation. Instead, we found that malin interacted with GP and increased nuclear localization of GP. Malin also altered the glucan‐binding and phosphorylation of GP. Thus, our data show that GP is a novel substrate of malin. Further exploration into the mechanisms of this regulation will unravel insights in glycogen metabolism, physiological functions of the malin‐laforin complex and will aid in betterment of LD pharmacotherapy.

Supported by NIH grants R00NS061803, R01NS070899 and University of Kentucky College of Medicine startup funds to MSG and AHA Great Rivers Affiliate Postdoctoral fellowship to VVD.
Stable anti‐ulcer gastric pentadecapeptide BPC 157 also for multiple sclerosis: Counteraction of cuprizone brain injuries and motor disability
Authors: Sikiric Predrag , Suran Jelena , Drmic Domagoj , Kolenc Danijela , Klicek Robert , Brcic Luka , Aralica Gorana , Seiwerth Sven , Rucman Rudolf
Published: June 2021

Like other agents for ulcerative colitis (i.e., natalizumab), stable gastric pentadecapeptide BPC 157 may affect multiple sclerosis (MS). BPC 157 is anti‐ulcer peptide (GEPPPGKPADDAGLV, M.W. 1419) stable in human gastric juice, successful for inflammatory bowel disease, phase II, wound treatment, no toxicity or side effects reported, LD1 not achieved,effective alone without carrier. Along with these, particularly for MS, direct healing of the severely injured muscles and severely injured nerves and brain injuries with BPC 157, not mentioned with DMDs, may define a particular way not relied on immunomodulation. Cuprizone diet 2.5% was combined with BPC 157 (/kg) in drinking water, μg or 10ng, 0.16μg or 0.16ng/ml/12ml/day/rat till the sacrifice after 4 days, with the additional ig (/kg) cuprizone 1g and BPC 157 μg or 10ng once daily. Several times higher cuprizone diet and regimen exaggerate and speed up the damaging process; nerve damage in various brain areas, most prominent damage in corpus callosum, laterodorsal thalamus, nucleus reunions, anterior horn motor neurons. Consistently less nerve damage was in all damaged areas in all BPC 157‐cuprizone rats, also in those areas that were most affected. Consistently, BPC 157 counteracts cerebellar ataxia, rescues impaired forelimb function. Thereby, we advocate BPC 157 also in MS therapy. Supported by Grant 108–1083570‐3635, Croatia
Single nucleotide polymorphisms and subcellular localization of glucocorticoid receptor‐associated signaling proteins associated with cortisol‐mediated stress response
Authors: Knox Rachel S. , Biette Kelly M. , Fox Olivia , Brasseale Nancy , Murphy Patrick J. M.
Published: June 2021

The goal of our study was to identify correlations between interpersonal genetic variability in healthy volunteers and alterations in the glucocorticoid receptor‐associated signaling pathway. Salivary cortisol (CORT) concentrations were measured in healthy volunteers (n=66) before and after CORT stimulation in order to determine variability in circulating CORT levels and CORT response. Statistical analysis revealed inter‐participant CORT variability in concentration or percent change were not exclusively correlated to participant demographics, health history, or glucocorticoid receptor (GR) single nucleotide polymorphisms (SNPs). Interestingly, nuclear localization of the GR‐associated molecular chaperone protein hsp90 correlated with an elevated pre‐stimulus CORT concentration. To investigate the observed CORT response‐hsp90 localization association further, coding and proximal coding regions of the constitutively expressed hsp90 gene (HSP90AA) were sequenced to identify a possible genetic linkage. Eight hsp90 SNPs were identified, four of which were not reported in the NCBI database. All participants expressed at least two of the hsp90 SNPs, with some participants exhibiting as many as five. These data demonstrate inter‐person differences in CORT response, nuclear hsp90 concentrations, and GR‐associated SNPs are readily detected and potentially interrelated.
G protein‐coupled receptor‐1 is an active signaling receptor for the adipokine chemerin
Authors: Rourke Jillian L , Shanmugam Muruganadan , Sinal Christopher J
Published: June 2021

BACKGROUND

Chemerin, is an adipose‐derived signaling protein (adipokine) that regulates adipocyte differentiation and metabolism through activation of chemokine‐like receptor‐1 (CMKLR1). Chemerin also binds to G protein‐coupled receptor 1 (GPR1), suggesting that this protein may be a second chemerin signaling receptor.

OBJECTIVE

To investigate differential signaling and expression of chemerin receptors

METHODS

Receptor signaling and tissue expression were assessed using a panel of in vitro luciferase‐ reporters and quantitative PCR, respectively.

RESULTS

GPR1 and CMKLR1 exhibit unique expression profiles which converge with the highest levels of each observed in white adipose tissue (WAT). CMKLR1 is highest in lipid‐filled adipocytes, while GPR1 expression predominates in the WAT stromal vascular fraction. Unlike CMKLR1, chemerin activation of GPR1 does not induce MAPK signaling, but does exhibit higher potency for arrestin recruitment. In common, chemerin activation of both CMKLR1 and GPR1 enhances serum‐response factor (SRF) – mediated gene transcription via a RhoA dependent pathway.

CONCLUSIONS

These results demonstrate that GPR1 is an active chemerin receptor and identify RhoA/SRF as a novel chemerin signaling axis. Differential receptor expression suggests that GPR1 may play a distinct role in chemerin function in WAT.

Supported by CIHR & Killam Trusts
Angiotensin II‐independent Activation of AT1 Receptors in Skeletal Muscle Arterioles
Authors: Hong Kwangseok , Nourian Zahra , Reim Sean , Yu Xichun , Li Hongliang , Yang Yan , Jackson William F , Benbrook Alexandra , Kem David C , Hill Michael A
Published: June 2021

Studies demonstrate activation of the angiotensin type 1 receptor (AT1R) by mechanical stimuli (Zou et al, 2004; Mederos y Schnitzler et al, 2008) and activating antibodies (Lukitsch et al, 2012). Such activation occurs independently of the receptor's classical agonist, angiotensin II. The purpose of the present study was to determine if the AT1R can be similarly activated in skeletal muscle arterioles. Using pressure myography, the effects of AT1R blockers were assessed on myogenic tone and pressure‐diameter relationships of isolated rat first order cremaster muscle arterioles. The AT1R antagonist Losartan (10−6M) had no apparent effect on steady‐state myogenic tone or pressure‐diameter relationships. In contrast, an alternate inverse agonist, Candesartan (10−7 and 10−6M), showed a partial but concentration‐dependent inhibition of myogenic reactivity as shown by a rightward shift in the pressure‐diameter relationship. AT1R activation, with concentration‐dependent vasoconstriction, was demonstrated with rabbit polyclonal antibodies (ab) directed to the second extracellular loop of the receptor. The ab‐induced constriction was inhibited by Candesartan (10−6M). Collectively the data support novel modes of activation of the AT1R in arterioles from skeletal muscle but also highlight possible variation in efficacy of AT1R blockers in inhibiting these responses. (Supp NIH HL092241).
Altered electrical reactivity of endothelial tubes with aging: Role of mitochondria and Ca2+‐activated K+channels
Authors: Behringer Erik J , Segal Steven S
Published: June 2021

Hyperpolarization of endothelial cells (ECs) is integral to coordinating smooth muscle cell relaxation throughout resistance networks and endothelial function is compromised with aging. Aberrations in mitochondrial function may contribute to endothelial dysfunction however little is known of such effects with aging in the microcirculation. We tested (P<0.05) whether disrupting the proton gradient across the inner mitochondrial membrane (IMM) altered plasma membrane potential (Vm) in endothelial tubes (width, 60 μm; length, 1–3 mm) freshly isolated (bath pH, 7.4; 32°C) from superior epigastric arteries of Young (4–6 mo) and Old (24–26 mo) male C57BL/6 mice. The IMM proton ionophore FCCP (10 nM‐10 μM) depolarized Young ECs progressively (from resting Vm of −33±2 mV to −17±2 mV; n=5). In contrast, FCCP (10 nM‐10 μM) hyperpolarized Old ECs progressively (from −42±4 to −67±3 mV; n=5) which was abolished by SKCa/IKCa channel blockade with apamin (Ap, 300 nM) + charybdotoxin (ChTx, 100 nM). In Old, 300 nM FCCP hyperpolarized ECs by −12±1 mV and this reversed to 9±1 mV depolarization with Ap+ChTx (n=4). Despite no effect on Vm of Young, BKCa channel activation (NS1619, 30 μM) hyperpolarized Old ECs by −13±1 mV (n=6). We hypothesize that changes in endothelial function with aging reflects mitochondrial dysregulation of SKCa/IKCa channels with acquired BKCa channel activity. (R01HL086483, R37HL041026, F32HL110701).
Role of L‐type calcium channels in acute alcohol intoxication‐induced enhancement of lymphatic calcium transient magnitude.
Authors: Souza‐Smith Flavia M. , Breslin Jerome W. , Molina Patricia
Published: June 2021

We previously demonstrated that acute alcohol intoxication (AAI) increases the magnitude of Ca2+ transients in pumping lymphatic vessels (LV). We tested whether sarcolemmal Ca2+ entry or Ca2+ release from the SR contribute to the AAI‐induced increase in Ca2+ transient magnitude. AAI was produced by intragastric administration of 30% alcohol to conscious, unrestrained rats through surgically implanted catheters. Isovolumic administration of water (vehicle) served as control. Mesenteric LVs were isolated, cannulated and loaded with Fura‐2 AM to measure changes in intracellular Ca2+. Measurements were performed at a basal intraluminal pressure of 2 cm H2O and during pressure steps to 6 and 10 cm H2O. Nifedipine (ND‐10−7 M) was used to block L‐type Ca2+ channels and xestospongin C (XC‐10−6 M) was used to inhibit SR Ca2+ release. The results show that ND reduced Ca2+ transient magnitude of LV from AAI and vehicle groups at all pressures studied, and also reduced contraction frequency (CF) of LV from the vehicle group. Treatment with XC did not significantly change any of the Ca2+ parameters in either LVs from the AAI or vehicle group. These data suggest that L‐type Ca2+ channels contribute to the AAI‐induced increase in lymphatic Ca2+ transient magnitude. We speculate that alcohol may activate a nifedipine‐insensitive pathway to maintain CF.
Preconditioning with Bay 60–2770, An Activator of Soluble Guanylate Cyclase, Attenuates Myocardial Infarction in eNOS Knockout Mice
Authors: Wang Meifang , Wang Derek Z , Jones Allan W , Zuidema Mozow Y , Wang Walter Z , Korthuis Ronald J
Published: June 2021

Our previous studies showed that soluble Guanylate Cyclase (sGC) activation protects against postischemic intestinal injury when used as a preconditioning stimulus in wild type, heme‐oxygenase‐1 KO and eNOS KO (eNOS −/−)mice. The aim of this study was to determine whether a similar protective effect of sGC activation by Bay60–2770 (Bay 60) would occur in postischemic cardiac injury. Infarct size (% LV), assessed by 2,3,5‐triphenyltetrazolium chloride (TTC) staining, was significantly increased in eNOS −/− mice after 30 min left anterior descending coronary artery (LAD) ischemia and 24 hr reperfusion as compared with eNOS −/− sham mice (p < 0.001). Bay 60 treatment (300ug/kg, ip bolus, 24hr prior to LAD ischemia) significantly attenuated I/R‐induced increase in infarct size (p < 0.05) in eNOS−/− mice. Serum Troponin‐I (cTnI), a marker of myocardial injury, after 30 min LAD ischemia and at 4 hr reperfusion, was markedly elevated in eNOS −/− mice, compared with eNOS −/− sham mice (p < 0.01). Preconditioning with Bay 60 24hr prior to cardiac I/R did not prevent the increase in cTnI levels. These findings indicate that Bay 60 preconditioning 24hr prior to cardiac I/R limits the size of the evolving infarct area at 24 hr of perfusion independently of eNOS and factors leading to early release of cTnI. Supported by a grant from the NIH (AA‐014945). BAY 60–2770 was a gift from Bayer Schering Pharma AG.
Deleterious Effect of Acute Administration of Nitric Oxide Donor GSNO on Cardiac Ischemia Reperfusion Injury in a Consomic Rat Model
Authors: Cheng Qunli , Nabor Darren , Stowe David , Bienengraeber Martin , Lazar Jozef , Riess Matthias
Published: June 2021

Endothelial nitric oxide (NO) synthase in part mediates cardioprotection by ischemic preconditioning (IPC). We hypothesized that genetically determined resistance to IPC in a consomic rat model can be restored by preconditioning with the NO donor GSNO. 15 min after two 5‐min boluses of 20 μM GSNO or vehicle (Con) with 5 min washout in‐between, isolated hearts from Brown Norway (BN), Dahl Salt Sensitive (SS), and consomic (SS‐6BN) underwent 30 min global ischemia (I) and 120 min reperfusion (R). On R, systolic and developed left‐ventricular pressure (LVP), contractility, and coronary flow were decreased while diastolic LVP and infarct size were increased in all Con groups, more so in SS than BN or SS‐6BN. GSNO did not improve function or IS in any group, but worsened IR injury in all strains, in SS‐6BN more than in SS or BN. Concurrent fluorescent redox state measurements suggest mitochondrial dysfunction and incomplete washout of GSNO before IR. These findings agree with prior reports of deleterious NO‐donor effects on mitochondrial function and cardiac IR injury. Possible causes could be dose, timing/duration, and/or tissue compartmentalization of endogenous vs exogenously administered NO. NO can react with superoxide to form peroxynitrite and thereby may amplify IR injury. Rat chromosome 6 appears to play a role in mediating this damage. Research supported by Department of Veterans Affairs (CARA‐026–10F) and NIH.
Localization of fluorescent particles with nanometer precision using standard microscopy and a point spread function approach
Authors: Marki Alex , Ermilov Eugeny , Secomb Timothy W. , Pries Axel R.
Published: June 2021

Background

Resolution of optical microscopy is limited by wave optics to about 200 to 400 nm. However, diffraction patterns generated by individual fluorescent particles (point spread functions, PSF) contain location information with much higher precision. Here, we describe a user‐friendly approach for single fluorescence particle 3D localization and tracking with nanometer precision in a standard fluorescence microscope.

Methods

Images of fluorescent particles were obtained over a large range (about 7 μm) of vertical positions and analysed by whole pattern (WP) comparison with a PSF recorded for the specific setup and measurement of the outermost ring radius (ORR).

Results

For a vertical range of 4.5 μm, standard deviation (SD) from the predicted value, indicating validity, was 15.6 and 23.5 nm for WP and ORR. Underexposure or overlapping images led only to a slight increase in SD. The precision, determined as SD of the distance between two particles, ranged from 3 to 6 nm. Local viscosity may be determined from random particle movement with a precision of about 5% of the measured value.

Conclusion

With a validity in the range of 20 nm and a precision of about 4–5 nm obtained by standard fluorescent microscopy, the PSF approach offers a valuable tool for a variety of experimental investigations of particle/structure localizations and local viscosity, e.g. in studies of the endothelial surface layer.
Relations of aortic pulse wave velocity and augmentation index assessed by the new brachial cuff SphygmoCor Xcel with other arterial stiffness measures
Authors: Hwang Chueh‐Lung , Hwang Moon‐Hyon , Yoo Jeung‐Ki , Kim Han‐Kyul , Mackay Karen R. , Hemstreet Olivia , Vasconez Estefania , Jacobs Kimberley , Nichols Wilmer W. , Christou Demetra D.
Published: June 2021

A number of methods are currently available for measuring arterial stiffness. However, it is not known whether differences exist between measures made on the left vs. right side of the body. Recently, the new SphygmoCor Xcel has been developed that measures aortic pulse wave velocity (AoPWV) and augmentation index (AIx). The first objective of this study was to use the SphygmoCor Xcel to examine the relation between AoPWV and AIx measured on the left vs. right side of the body. The second objective was to assess the relation of AoPWV and AIx (SphygmoCor Xcel) compared with carotid arterial compliance, distensibility and beta stiffness index (high resolution ultrasonography and applanation tonometry). Twenty‐six adults participated in this study (16 men, 25 to 75 y). AoPWV and AIx measured on the left vs. right side of the body were not significantly different (mean difference: −0.15±0.1 m/s, P=0.2 and −0.29±0.7 %, P=0.7, respectively) and were strongly related (AoPWV: r=0.94, P<0.0001 and AIx: r=0.98, P<0.0001). AoPWV and AIx were positively related with beta stiffness index (r=0.44, P=0.04 and r=0.42, P=0.04, respectively), but were not related with arterial compliance or distensibility (P>;0.5). In conclusion, arterial stiffness measures from the left and right side of the body are similar and highly correlated. Compared to other measures, AoPWV and AIx are only significantly related with the beta stiffness index.
Renin/prorrenin receptor expression on leukocytes associated with an experimental model of preeclampsia
Authors: Sanchez‐Guerrero Erika , Hernandez‐Campos Maria Elena , Trinidad‐Gomez Margarita Itzel , Lopez‐Sanchez Pedro
Published: June 2021

Renin/prorrenin receptor (RPR) is an important component of the renin‐angiotensin system; this system plays a relevant role in blood pressure regulation and electrolyte balance. RPR is a single‐spanning membrane protein that binds both renin and its inactive precursor, prorrenin. Binding to RPR increases the catalytic activity of renin and induces non‐proteolytic activation of prorrenin, leading to angiotensin‐dependent responses. The presence of RPR on immune cells and if this presence plays a role during preeclampsia is unknown. The aim of this work was to determine the expression of RPR on leukocytes from spleen during normal pregnancy and pregnancy complicated with preeclampsia in rats. We used female Wistar rats groups: 1) non‐pregnant rats, 2) pregnant rats, 3) non‐prengnant rats whit subrenal aortic coarctation, 4) pregnant rats with subrenal aortic coarctation (preeclamptic rats). We used flow cytometry, inmunoblot and RT‐PCR to determine RPR expression upon leukocytes from spleen. We found similar, basal expression of RPR in groups 1–3; however, preeclamptic rats expressed this receptor in a lower amount. These suggest a possible role of RPR in PIH. Supported by SIP‐IPN 20120122 and 20120929 grants
COMP‐Ang1 prevents structural and functional damage of the retina in a mouse model of central retinal artery occlusion
Authors: Olson Paul Robert , Cahoon Judd Michael , Flood Maggie Marie , Zhang Xiao Hui , Ambati Balamurali Krishna
Published: June 2021

The retina is the most metabollicaly active tissue in the body. Central retinal artery occlusion (CRAO) is a leading cause of blindness. CRAO results in retinal edema, changes in morphology, and loss of electrical function. We hypothesize that by stabilizing retinal blood vessels, the effects of CRAO can be mitigated. Cartilage Oligomatrix Protein Angiopoietin1 (COMP‐Ang1), promotes vascular stability through the Tie‐2 receptor mediated increase in adherens junctions. We tested whether COMP‐Ang1 could prevent the pathologies of CRAO. C57 mice were treated with intravitreal injections of PBS, or viral vector expressing GFP or COMP‐Ang1. Vessel occlusion in mice was accomplished by excitation of intravenously injected rose bengal by laser (588 nm) focused on the central retinal artery. COMP‐Ang1 decreased retinal edema, as evidenced by cross‐sections taken 1 day after laser, and maintained retinal morphology when compared to mice treated with control. COMP‐Ang1 improved both electrical retinal function, as assesed by scotopic electrical retinogram (ERG) (b‐wave amplitude 120 uV compared to 25uV control) and visual acuity, as determined by optokinetic tracking response (OptoMotry) (0.134 +/− 0.08 cycles/degree compared to 0.075 +/− 0.02 cycles/degree, control). These results demonstrate that promoting vascular stability may be beneficial to patients with CRAO and other retinal ischemic diseases.
Skeletal muscle remodeling in myofiber VEGF‐deficient exercise trained mice
Authors: Delavar Hamid , Rich Benjamin , Wagner Peter D. , Breen Ellen C.
Published: June 2021

VEGF plays an essential role in regulating angiogenesis and clearance of apoptotic cells. A loss of skeletal myofiber expressed VEGF inhibits exercise‐induced angiogenesis. We hypothesized that skeletal myofiber VEGF deficient mice would increase fibroblast proliferation in hind limb muscles; especially following repeated bouts of exercise.

Methods

Using an HSA‐Cre‐ERT2 X VEGFLoxP (skmVEGF−/−) mouse model, VEGF levels are reduced by >;80% in skeletal muscles of adult mice compared to control littermates (WT). 8 week exercise trained (EX) and untrained (UT) mice were administered BrdU daily during the final week. Proliferating cells (BrdU+) and fibroblasts (vimentin+) were detected by confocal microscopy.

Results

Total BrdU+ cells/myofiber increased in UT and EX skmVEGF−/− mice compared to the WT‐UT and WT‐EX groups, respectively, in the plantaris (WT‐UT, 6%, skmVEGF−/−UT, 32%, WT‐EX, 37%, skmVEGF−/−EX, 47%) and gastrocnemius (WT‐UT, 2%, skmVEGF−/−UT, 43%, WT‐EX, 25%, skmVEGF−/−EX, 43%). Exercise training, alone, also increased BrdU incorporation in WT muscles. Gastrocnemius sections from exercise trained skmVEGF−/− mice displayed fibrotic foci containing numerous vimentin+/BrdU+ cells. These data suggest that VEGF may prevent muscle injury and fibrosis during exercise training.
Functional Vasodilation is Impaired in Arterialized Capillaries following Ischemia
Authors: Cutts Joshua Patrick , MacGabhann Feilim , Cardinal Trevor
Published: June 2021

Ischemic disease is the leading cause of morbidity and mortality in the developed world. Patients with a well‐developed collateral network experience less severe symptoms following ischemic events, but not all patients have well‐developed collateral networks. Stimulating arterialization of capillaries into functional collaterals could minimize the impact of ischemic events in patients with poorly‐developed collateral networks. Stable arterialized capillaries develop following occlusion, but the reactivity of these vessels is unknown. Ischemic events, such as angina, occur with increased metabolic demand, so effective collaterals must be capable of dilating and increasing blood flow to support tissue oxygen demand. To examine the ability of arterialized capillaries to regulate blood flow, we ligated the lateral cranial feed artery of the right spinotrapezius muscle and measured functional vasodilation at days 7 and 21 following arterial occlusion. Vasodilation in arterialized capillaries was absent at day 7 compared to arterioles with a similar resting diameter. (2% ± 2% vs. 29% ± 12% p< 0.05) Vasodilation was restored by day 21.(167% ± 17% vs 81% ± 13% p>; 0.05) Studies are currently underway to examine the mechanism of impaired vasodilation at day 7 and the hemodynamic effects of vasodilation at day 21 following arterial occlusion.
Elevation of extracellular NaCl increases secretion of von Willebrand Factor from endothelial cells
Authors: Dmitrieva Natalia I. , Burg Maurice B.
Published: June 2021

Osmoregulation maintains normal systemic osmolality at 270–290 mosmol/kg. However, it is higher in some organs: 330–335 in spleen, liver and thymus, 400–550 in intervertebral discs, 400 in synovial fluid, and up to 1700 in human renal medulla, where it powers concentration of the urine. During pathological hypernatremia, serum osmolality can rise to 340. Hypernatremia is a known risk factor for thrombosis. Von Willebrand factor (vWF) plays a key role in thrombosis by mediating platelet adhesion to vascular walls. We now find that, in tissue culture of Human Umbilical Vein Endothelia Cells, elevating medium osmolality to 320–380 mosmol/kg by adding NaCl reversibly increases both vWF mRNA and the rate of vWF secretion. In mice, vWF is higher in endothelial cells of the kidney medulla, where NaCl is high, than in kidney cortex, where it is not, as measured by immunostaining. Further, water restriction increases blood vWF. High blood vWF is a recognized risk factor for cardiovascular disease. We suggest that disturbed water balance and hypernatremia increase vWF, resulting in increased risk of thrombosis. Supported by the Intramural Program of NHLBI
Serum S100B is associated with stress, adiposity and elevated blood pressure
Authors: Cannon Joseph G. , Sloan Gloria J. , Webb R. Clinton , Cortez‐Cooper Miriam
Published: June 2021

This study addressed the hypothesis that chronically elevated blood pressure injures vascular cells and overly‐stresses intracellular matrix, causing the release of damage‐associated molecular pattern molecules (DAMPs) that initiate and sustain low‐level inflammation, further elevating blood pressure. S100B is a DAMP released by several types of cells, including adipocytes, that can affect vascular function, activate monocytes, and has been associated with stress. We measured carotid systolic blood pressure (cSBP), serum S100B and cortisol concentrations, monocyte activation (CD16 expression) and percent body fat (DEXA) in normotensive and prehypertensive (but otherwise healthy) men (n = 14) and women (n = 22), 20–40 years of age. Subjects with above‐median S100B concentrations had significantly higher cSBP than those with below‐median S100B (P = 0.01), and this difference was significantly greater in men (P = 0.03). Monocyte CD16 expression correlated with serum S100B in the prehypertensive subjects (R = 0.67, P = 0.01). In addition, high S100B concentrations were associated with high cortisol (P = 0.03) and % body fat (P = 0.05). These data are consistent with the concept that S100B contributes to the low‐level inflammation associated with elevated blood pressure. Moreover, S100B may represent a mechanistic link between adiposity and increased blood pressure. Supported by NIH HL093663.
Melatonin combined with treadmill exercise synergistically promotes neurogenesis and reduce apoptosis in focal cerebral ischemic rats
Authors: Lee Minkyung , Lee Seunghoon , Hong Yunkyung , Kim Koo , Hong Yonggeun
Published: June 2021

Stroke potently stimulates neuronal stem cell neurogenesis. However, only a small fraction of new born cells survives. The aim of this study, we determine whether melatonin and/or exercise could be induced to the contribution of apoptotic cell death as well as promoted to functional consequence followed by neurogenesis in ischemic brain. Sprague‐Dawley (SD) rats were randomly divided into control (Con), MCAo+Vehicle (MCAo), MCAo+Exercise (MCAo+Ex), MCAo+Melatonin (MCAo+MT), Melatonin combined with Exercise in MCAo (MCAo+MT+Ex). Following the MCAo, the SD rats were treated with MT (10 mg/kg) at 7:00 and 19:00, trained with Ex (20 m/min, 6 days/week) at 17:00 and 22:00. We confirmed that MT+Ex was significantly improved the neurological symptom in behavioral assessment (p < 0.01). Histological data showed that the loss of neural cells on the primary motor cortex was significantly reduced in MT+ME. Moreover, the expression of Nestin was significantly enhanced in MT and Ex compared to MCAO (p < 0.01). At day 28, Pro‐apoptosis marker Bax was decreased (p < 0.05), whereas anti‐apoptosis marker Bcl2 was increased in MT and Ex group (p < 0.01). These results suggest that MT and/or Ex intervention enhanced generation of neuronal cells and mediated reduction of apoptosis, as well as improvement of neurological dysfunction.
Ventilatory pattern variability predicts long‐term survival following cardiac arrest and resuscitation in rats
Authors: Xu Kui , Kuang Youzhi , Jacono Frank J. , Dick Thomas E. , LaManna Joseph C.
Published: June 2021

Ventilatory pattern variability (VPV) reflects brainstem function with respect to cardio‐respiratory regulation. We applied linear and nonlinear tools to assess VPV following cardiac arrest and resuscitation with the goal of predicting long‐term survival. Spontaneous breathing (plethysmography) was recorded for 1h for VPV analysis at baseline and after cardiac arrest (12 min) and resuscitation in male Wistar rats. VPV was similar before cardiac arrest but differed between survivors (survived for 4d, n = 15) and non‐survivors (died before 4d, n = 12) after resuscitation. The nonlinear complexity index (NLCI), a measure of deterministic structure in ventilatory patterns, was similar between survivors and non‐survivors (0.08 ± 0.03 vs.0.07 ± 0.03 bits) before cardiac arrest. On day 0 and day 1 post‐resuscitation NLCI was significantly increased (~3 folds) in the non‐survivors while remained unchanged from baseline in the survivors. Traditional measures such as respiratory rate and coefficient of variation of respiratory cycle length cannot distinguish the difference in ventilatory pattern between the survivors and non‐survivors. The characteristic changes in VPV early in recovery distinguished the survivors from non‐survivors, suggesting that VPV could be a predictor of long‐term survival following cardiac arrest and resuscitation. Supported by NS 38632, HL 087377, VA Research Service (I01BX000873).
Inhibition of glial glutamate transporter GLT1 in the nucleus tractus solitarii attenuates baroreflex control of sympathetic nerve activity and heart rate
Authors: Yamamoto Kenta , Hosford Patrick S. , Ramage Andrew G. , Franzke Michelle , Mifflin Steve
Published: June 2021

Astrocyte glutamate transporter (GLT‐1) removes neuronally released glutamate from the extracellular space. We examined the effect of inhibiting GLT1 in the nucleus tractus solitarii (NTS) on mean arterial pressure (MAP), renal sympathetic nerve activity (RSNA) and heart rate (HR) in Inactin anesthetized and artificially ventilated Sprague‐Dawley rats. In experiment 1, it was found that dihydrokainate (DHK; inhibitor of GLT1, 5mM, 100nl) injections into the NTS (n=4) decreased MAP (49±12mmHg), RSNA (84±15%) and HR (34±5bpm). Application of kynurenate (KYN; glutamate receptor antagonist, 5mM, 30μl) to the dorsal surface of the brainstem (n=4) blocked responses to NTS injections of DHK with MAP now only decreasing by 14±12mmHg, RSNA by 26±27% and HR by 10±8bpm (p<0.001). The DHK response returned after 30min. In experiment 2, the effect of DHK on arterial baroreflex was examined using i.v. infusions of phenylephrine and nitroprusside. NTS infusions of DHK (5mM, 40nl/min; n=6) reduced baroreflex response range (maximum ‐ minimum) of RSNA by 91±2% and HR by 83±5% (p<0.001). These results indicate that inhibition of GLT1 decreases MAP, RSNA and HR by indirect activation of ionotropic glutamate receptors and that the astrocytic glutamate transporter in the intermediate NTS plays an important role in the maintenance and regulation of cardiovascular function. Supported by HL‐088052
Caveolin‐1 phosphorylation on tyrosine 14 affects neuronal growth
Authors: Rohrback Suzanne E , Patel Hemal H , Patel Piyush M , Head Brian P
Published: June 2021

We have previously shown that neuron‐targeted caveolin‐1 (Cav‐1, SynCav1 vector) promotes dendritic growth and arborization. The mechanism by which Cav‐1 promotes neuronal growth is not completely understood. Cav‐1 is phosphorylated on tyrosine 14 (Y14) by Src tyrosine kinase during cell migration in non‐neuronal cells and during NMDAR activation in primary neurons, leading us to hypothesize that Cav‐1 phosphorylation is a key for receptor signaling resulting in neuronal growth. To test our hypothesis we assessed the effects of a non‐phosphorylatable mutant of Cav‐1 (SynCav1‐mutP) on neuronal signaling and growth. Levels of MAP2 and tau (dendritic and axonal markers) were assessed by Western blot and found to be increased 2–3 fold in wild type mouse primary neurons after incubation with SynCav1 for 21 days as compared to SynGFP transfection. Interestingly, SynCav1‐mutP transfection resulted in only a 2‐fold MAP2 increase. Confocal microscopy staining for MAP2 showed a similar trend, with a 2‐ fold and 1.5‐fold increase in dendritic branching, length, area, and volume following SynCav1 or SynCav1‐mutP treatment, respectively. These results indicate that Cav‐1 phosphorylation on Y14 may in part regulate pro‐growth signaling in primary neurons. (Supported by VA CDA‐2, VA Merit, and NINDS)
Angiotensin (1–7) and Mas receptor activation inhibits Aldosterone induced ROS inflammatory responses in neurons of the paraventricular nucleus (PVN).
Authors: Hay Meredith , Xue Baojian , Johnson Ralph Fred , Beltz Terry G , Johnson Alan Kim
Published: June 2021

It is now well recognized that the renin angiotensin system includes two separate enzymatic pathways that provide a physiological counterbalance of two related peptides acting at distinct receptors. The well described ACE ‐AngII‐AT1R system is thought to be physiologically opposed and balanced by the ACE2‐Ang‐(1–7)‐Mas receptor system. In the brain and other tissues, Ang II activation of AT1 receptors increases ROS related inflammation while Ang (1–7) and Mas receptor activation decrease ROS and inflammation. The present study investigated the effects of Ang (1–7) aldosterone (ALDO) induced increases in ROS production in PVN cells in vitro. PVN neurons were isolated from 22 day old female SD rats, dissociated and cultured on round coverslips and maintained in HBSS media for at least 48 hours prior to visualization by confocal microscopy. All cells were preincubated in 10nM ALDO for 48 hours and then 1.0 μM Ang (1–7) or 1.0 μM Ang (1–7) + 10.0 μM A779 for 2 hours prior to testing with acute 1.0 μM Aldo. To measure ROS, changes in the ethidium fluorescence were measured which is directly proportional to the oxidation of dihydroethidium (DHE) by ROS. The cells were loaded with DHE (20 μM) added to the cell culture media for 1h and then placed in a chamber for microscopic visualization of fluorescence. Perfusion with 1.0 μM ALDO increased ROS (Δ=113.4±28%) in the PVN neurons. In cells preincubated in Ang (1–7), ALDO increase in ROS was significantly inhibited (Δ=6.4±27%). Preincubation in Ang (1–7) + A779, restored the ALDO increase in ROS (Δ=105.9±9%). These results suggest that Ang (1–7) can inhibit ROS inflammatory effects of ALDO and this effect involves Ang (1–7) activation of the Mas receptor. (Supported by: HL‐14388, HL‐98207, and MH‐80241).
Sympathoexcitation and Pressor Response Induced by Central Amygdala‐injected Ethanol Requires Activation of Local NMDA Receptors
Authors: Chen Qing‐Hui , Gui Le , Larson Robert A , Chapp Andrew , Keim Alexander , Gu Mingjun
Published: June 2021

The Central Amygdala (CeA) is involved in the sympathoexcitatory response to ethanol intake. However, the underlying neural mechanisms have not been determined. We tested the hypothesis that the increase in sympathetic nerve activity (SNA) in response to CeA‐injected ethanol requires activation of local NMDA receptors. In anesthetized rats, CeA ethanol (n=7) increased splanchnic SNA (SSNA, 121±21%), lumbar SNA (LSNA, 72±11%), and MAP (10±4mmHg) respectively. A cocktail containing KYN (7.2 nmol), a non‐selective glutamate receptor blocker, and ethanol attenuated responses (p<0.05; n=6) in SSNA (34±10%), LSNA (25±6%), and MAP (5±1mmHg) compared to that elicited by CeA ethanol alone (p<0.05). Similarly, responses to a cocktail containing AP5 (6 nmol), a NMDA receptor blocker, and ethanol in SNA and MAP were less than ethanol alone (p<0.05; n=6) and not statistically different from a cocktail containing KYN and ethanol. The symapthoexcitatory response to a cocktail containing NBQX (2.6 nmol), a non‐NMDA receptor blocker, and ethanol (n=6) were not different from CeA ethanol alone. CeA injection of KYN, AP5, and NBQX did not affect baseline parameters. Immunohistochemistry studies demonstrate abundant expression of NMDA NR1 receptors in CeA neurons with projections to the RVLM. This data indicates that sympathoexcitatory responses to CeA‐injected ethanol require action of local NMDA receptors. Support: AHA2640130 (QHC).
Electroacupuncture activated pathway from caudal ventrolateral medulla to rostal ventrolateral medulla
Authors: Guo Zhiling , Longhurst John C.
Published: June 2021

Our previous studies have shown that electroacupuncture (EA) at the Jianshi‐Neiguan acupoints (P5‐P6, overlying the median nerve) attenuates sympathoexcitatory responses through inhibition of neuronal activity in rostral ventrolateral medulla (rVLM). The caudal ventrolateral medulla (cVLM) modulates rVLM sympathetic nerve activity during baroreflex stimulation through a GABAergic mechanism. However, there is no information on the role of cVLM neurons in processing somatic nerve input during EA. Thus, the present study evaluated the potential for EA to activate cVLM neurons that project to the rVLM. Seven to ten days after unilateral microinjection of the retrograde tracer, rodamine‐conjugated microspheres (100 nl) into the rVLM, rats were subjected to 30 min of EA or control (needle placement without electrical stimulation) at P5‐P6 acupoints applied bilaterally. Perikarya containing the microsphere tracer were identified in the cVLM of both groups. Compared to three control animals, immunoreactivity of c‐Fos, an immediate early gene and neurons double‐labeled with c‐Fos and the tracer were more frequently found in the cVLM both rostrally and caudally (Bregma, −12.84 to −14.04 mm) in four EA‐treated rats. Neurons often colabeled with GAD67 mRNA detected with in situ hybridization and c‐Fos protein in the cVLM in two EA‐treated rats, but rarely in two controls. These results suggest that EA at P5‐P6 acupoints activates cVLM neurons, some of which are GABAergic. Furthermore, EA‐activated cVLM neurons frequently project directly to the rVLM (supported by NIH grants HL‐72125 and HL‐63313).
Chronic intermittent hypoxia (CIH, 3d) attenuates glutathione peroxidase expression (Gpx1) and function in the caudal nucleus tractus solitarii (cnTS)
Authors: Barr Stacy L. , Phaup J. Glenn , Kline David D. , Hasser Eileen M. , Heesch Cheryl M.
Published: June 2021

Hypoxia increases reactive oxygen species (ROS) in the nTS which likely contribute to changes in cardiorespiratory control. Previously we reported that 2 h of acute hypoxia increased mRNA for the ROS inactivating enzyme Gpx1 in the nTS. Current experiments evaluated GPX in CIH (3d). Gpx1 mRNA in the cnTS trended to decrease in 6 CIH (0.6 ± 0.1) versus 6 normoxic (N) male rats (1.0 ± 0.3). In inactin anesthetized, neuromuscular blocked, artificially ventilated and vagotomized N (6) and CIH rats (5), baseline cardiorespiratory values were similar. Microinjection of mercaptosuccinic acid (MCS, 30 nl; 0.05, 0.1, 0.5M), a GPX antagonist, in the cnTS produced similar graded responses. At 0.5M, MCS eliminated phrenic nerve discharge, and decreased mean arterial pressure ( N = −50 ± 5; CIH = −45 ± 5 mmHg) and heart rate ( N = −50 ± 5; CIH = −39 ± 9 bpm). However, decreases in splanchnic sympathetic nerve activity due to 0.5M MCS were attenuated in CIH (−68 ± 8%) versus N rats (−87± 4%), consistent with decreased Gpx1 mRNA. Data suggest that an initial compensatory increase in Gpx1 mRNA is reversed following 3 d CIH possibly contributing to attenuated sSNA responses to GPX blockade. NIH HL 98602