bluuebunny/crossref_metadata_2025 huggingface dataset:
Neuroprotective effects of hibernating woodchuck plasma and plasma fractions
Authors: Brown Stephen A. , Govindaswami Meera , Kindy Mark S. , Oeltgen Peter R.
Published: June 2021

We tested the ability of hibernating woodchuck plasma (HWP) and plasma fractions and summer active woodchuck plasma (SAWP) and plasma fractions to provide ischemic neuroprotection in mice after 1 hr of middle cerebral artery occlusion. C57BL/6 mice (n=5 per group) weighing 25 g were anesthetized and injected in the tail vein with either 200 μl of PBS, 200 μl HWP, 200 μl SAWP, 5 mg/kg of an affinity chromatography purified HWP or SAWP albumin fraction or with 5 mg/kg of an anion exchange 88 kDa hibernation‐specific fraction (HSF). Twenty‐four hr after ischemia, coronal brain sections were subjected to 2% triphenyl tetrazolium chloride staining. One mm thick serial sections were cut and the infarct area in each section was determined according to the formula (contralateral volume‐ipsilateral volume × 100/contralateral volume). Infarct volume with SAWP and SAWP albumin fraction treatment was not significantly different compared to the PBS control of 78.4 ±7 mm3(SD). Infarct volume with HWP treatment decreased 32% to 53 ±6 mm3(p<0.05) while infarct volume following treatment with HWP albumin fraction decreased 26% to 55 ±6 mm3 compared to the PBS control of 74 ±3 mm3. The 88 kDa HSF decreased infarct volume 32% to 49 ±4 mm3 (p<0.001). Our study indicates that HWP and HWP fractions, but not SAWP can provide ischemic neuroprotection. Supported by Office of Naval Research Grant (PRO) and VA Merit Review Grants (MSK, SAB).
Bicyclol protects mice brain from ischemia/reperfusion injury
Authors: Sun Lina , Shen Jia , Liao Yuanjin , Lin Shupeng , Bai Xiaoyin , Xia Qiang
Published: June 2021

The aim of this study was to investigate the neuroprotective effect of bicyclol against global brain ischemia/reperfusion injury in male C57BL/6 mice. Before ischemia, the animals received intragastric administration once a day with bicyclol at different concentrations (10, 25, 50 and 100 mg/kg) for 3 days. Global ischemia were achieved 6 h after the last administration by bilateral carotid arteries occlusion for 15 min. 24 h after reperfusion, half animals in each group were sacrificed for measuring the serum lactate dehydrogenase (LDH) level and the activity of superoxide dismutase (SOD) in the brain tissue. The other animals were allowed reperfusion for another 6 days to determine the histological injury in CA1 area of hippocampus. We found that bicyclol at 25, 50 and 100 mg/kg decreased serum LDH release, increased the activity of brain SOD and survival rate of neurons in the hippocampal CA1 area in mice subjected to global brain ischemia and reperfusion. In another series of experiments for focal brain ischemia, bicyclol at 25, 50 and 100 mg/kg reduced the infarct size and neurological severity scores significantly. We concluded that bicyclol can protect the brain from ischemia/reperfusion injury in mice.
PPARs attenuate HIV‐1‐induced dysregulation of tight junction protein expression in human brain endothelial cells
Authors: Huang Wen , András Ibolya E , Eum Sung Yong , Zhong Yu , Couraud Pierre‐Olivier , Hennig Bernhard , Toborek Michal
Published: June 2021

Blood‐brain barrier (BBB) plays an important role in HIV‐1 trafficking into the brain and the development of CNS complications in the course of HIV‐1 infection. Exposure of human brain microvascular brain endothelial cells (HBMEC) to HIV‐1‐infected monocytic U937 cells resulted in decreased expression of tight junction proteins, such as JAM‐A, ZO‐1, and ZO‐2. In addition, exposure to HIV‐1‐infected U937 cells stimulated their transendothelial migration and decreased transendothelial resistance of an in vitro model of the BBB. Most importantly, these effects were markedly attenuated by overexpression of PPARalpha or PPARgamma in HBMEC. To determine the mechanisms involved in HIV‐1‐induced disruption of tight junctions, our studies focused on the role of matrix metalloproteinases (MMPs) and proteasome. HIV‐1 induced dysregulation of JAM‐A and occludin expression was attenuated by GM6001, a general inhibitor of MMPs. In addition, alteration of ZO‐1 and ZO‐2 expression was inhibited by blocking of proteasome activity by lactacystin. Overexpression of PPARs resulted in decreased MMP and proteasome activities. The present data indicate that PPARs play important roles in protecting against HIV‐1 induced dysfunction of brain endothelial cells by attenuating MMP and proteasome activities. Supported by NS39254, MH63022, and MH072567.
Caffeine blocks MPTP‐induced increases in blood‐brain barrier leakage in mouse striatum
Authors: Chen Xuesong , Lan Xun , Liu Rugao , Geiger Jonathan D.
Published: June 2021

The blood‐brain barrier (BBB) is important physiologically, and pathologically BBB disruption has been implicated in a wide spectrum of neurological disorders including Parkinson's disease. Recent studies indicate that caffeine is protective against Parkinson's disease, but by poorly understood mechanisms. Using a MPTP neurotoxin model of Parkinson's disease, we tested the hypothesis that caffeine protects against MPTP‐induced BBB leakage. FVB mice were pretreated with caffeine (10 mg/kg, i.p.) or saline 10 min before each dose of MPTP (20 mg/kg, i.p.) or saline for 7 days. Striatum (and for some studies hippocampus and cerebral cortex as well) were evaluated for BBB leakage, tight junction protein expression, presence of dopaminergic neurons, and neuroinflammation by immunostaining, immunoblotting or real‐time PCR techniques. We found that caffeine blocked MPTP‐induced decreases in TH‐positive dopaminergic neurons, increases in leakage of Evan's blue dye specifically in stratum but not in cerebral cortex or hippocampus, decreases in levels of the tight junction proteins occludin, claudin‐5 and ZO‐1, and increases in reactive astrocytosis. Our results suggest that caffeine might protect against Parkinson's disease, in part, by stabilizing the BBB. (Supported by P20 RR‐017699 from the NCRR, a component of the NIH)
Ischemia‐induced Stimulation of the BBB Na‐K‐Cl Cotransporter: Evidence for a Role of p38 MAP Kinase
Authors: Wallace Breanna K , Jelks Karen A , Foroutan Shahin , O’Donnell Martha E
Published: June 2021

Increased blood‐brain barrier (BBB) Na‐K‐Cl cotransporter activity appears to contribute to cerebral edema formation during ischemic stroke. The cotransporter is stimulated by hypoxia and aglycemia, although the mechanisms responsible are unknown. Inhibition of p38MAP kinase (p38MAPK), which is activated by ischemia, has been found to reduce brain infarct in ischemic stroke. In the present study we examined the possibility that p38MAPK participates in ischemia‐induced stimulation of the BBB Na‐K‐Cl cotransporter. Cerebral microvascular endothelial cells (CMEC) were exposed to hypoxia (7.5% O2) or to aglycemia, then subjected to Western blot analysis, using antibodies that recognize phosphorylated (activated) p38MAPK (p‐p38MAPK) or both phosphorylated and non‐phosphorylated (total) p38MAPK. CMEC were also evaluated for cotransporter activity as bumetanide‐sensitive K influx. Hypoxia (5 – 120 min) and aglycemia (30 min) caused 1.9‐ and 2.8‐fold increases in p‐p38MAPK levels without changing total p38MAPK. Exposing CMEC to the p38 MAPK inhibitor SB239063 (20 μM) abolished hypoxia‐ and aglycemia‐induced increases in Na‐K‐Cl cotransporter activity. Confocal immunofluorescence of perfusion‐fixed rat brain slices also revealed the presence of p38MAPK in BBB in situ. These findings suggest that p38 MAPK participates in ischemia stimulation of BBB Na‐K‐Cl cotransporter activity.

Supported by NINDS
Characterization of a functional (pro)renin receptor (PRR) in brain neuron
Authors: Shan Zhiying , Cuadra Adolfo E. , Raizada Mohan K.
Published: June 2021

Discovery of PRR has provided a novel mechanism by which RAS can influence the cardiovascular system. Initial findings from the kidney suggest that (pro)renin binding to PRR initiates a signaling cascade that leads to an angiotensin‐independent physiological responses. While knowledge is limited for the periphery, even less is known of PRR function in the brain. Thus our objectives in this study were to determine if PRR exists in the brain and if it is functional. Primary cell cultures from WKY rat brain were used in this study. Levels of PRR mRNA were 2.4 fold higher in neuronal compared to astroglial cultures. Immunofluorescence in neurons showed that PRR were localized both throughout the soma and in neurites as immunoreactive punctae. Treatment of neurons with renin in the presence of 2 μM losartan, caused a time‐ and dose‐dependent, transient increase in phosphorylation of p42 and p44 isoforms of MAP kinase. Phosphorylation peaked at 4 fold by 2 min with 20 nM renin. Current patch clamp recordings showed that neurons treated with renin, in the presence of losartan, resulted in a steady and stable decrease in action potential frequency to 56 % of basal within 5 min. This renin‐mediated decrease was blocked by pretreatment with MAP kinase inhibitor, PD89059. These results demonstrate that neurons from rat brain express functional PRR that is coupled to MAPK and neuronal activity. (Supported by NIH HL336 10)
Role of Sodium Glucose Transporter in High Glucose Mediated Angiotensin Type 1 receptor Down‐regulation in Human Proximal Tubule Cells
Authors: Yesudas Rekha , Abbruscato Thomas , Thekkumkara Thomas
Published: June 2021

Previously, we have characterized the human angiotensin type 1 receptor (hAT1) promoter architecture for high glucose (HG) mediated transcriptional repression in human proximal tubule epithelial cells (hPTEC). In the present study, we investigated the role of glucose transporters in HG mediated hAT1 repression. Primary hPTEC were exposed to 5.5 mM normal glucose (NG) or 25 mM HG, and changes in receptor expression and glucose transporter activity were determined. Exposure of cells to HG resulted in down‐regulation of angiotensin II specific binding (4034 to 1360 dpm/well) and hAT1 mRNA expression (58.0 % of normal) at 48 h. Under similar conditions, we observed significant increases in glucose uptake (influx) in HG treated cells that were concentration and time dependent. In NG treated cells, inhibiting the sodium glucose transporters (SGLTs) with phloridzin and facilitative glucose transporters (GLUTs) with phloretin decreased glucose influx by 28.8 and 45.5 %, respectively. In contrast, in HG treated cells inhibiting the SGLTs decreased glucose influx by 52.0 %, however GLUT mediated glucose uptake remained the same. Pretreatment of cells with SGLT inhibitor reversed HG mediated down‐regulation of hAT1. Our study shows for the first time that in hPTEC HG mediated hAT1 down‐regulation is largely mediated through SGLT dependent glucose influx pathway.
Farnesoid X receptor regulation of adrenal cell steroidogenic enzyme expression
Authors: xing yewei , Saner‐Amigh Karla , Hinshelwood Margaret , Carr Bruce , Rainey William
Published: June 2021

Farnesoid X receptor (FXR, NR1H4), a member of the nuclear receptor superfamily, is expressed in a number of tissues including the liver, ovary and adrenal. In the liver, FXR regulates cholesterol and glucose homeostasis, but its function in the ovary and adrenal remains unknown. Herein, we tested the hypothesis that FXR regulates expression of the enzymes involved in steroid hormone biosynthesis. Human adrenocortical cells were transfected with FXR expression vectors and treated with the FXR ligand GW4064; followed by microarray analysis. Among the adrenal FXR target genes were the well established liver FXR target, OSTα as well as the steroid‐metabolizing enzyme, 3β‐hydroxysteroid dehydrogenase type II (HSD3B2). HSD3B2 promoter construct studies and quantitative real‐time RTPCR showed that FXR stimulates HSD3B2 transcriptional activity and mRNA levels. Sequential deletions and mutational analysis of the HSD3B2 promoter defined an inverted hormone response element repeat between −137 and −124 that was necessary for FXR‐induced reporter activity. Chromatin immunoprecipitation assay demonstrated the ability of FXR to bind this response element. Since HSD3B2 is critical for the production of both adrenal and ovarian steroid hormones, these results indicate a new and important role of FXR in the regulation of steroidogenesis in the ovary and adrenal.
Angiotensin II and Potassium Regulate Aldosterone Production: the Expression of Steroidogenic Enzymes and Type 1 Angiotensin II Receptors in a Newly Developed Adrenocortical Cell Line
Authors: Parmar Jeniel , Key Rebecca , Rainey William
Published: June 2021

The molecular mechanisms regulating adrenal glomerulosa production of aldosterone remain poorly defined in part due to limitations in obtaining sufficient adrenal glomerulosa tissue and cells. The only current adrenocortical cell model for aldosterone studies is the NCI‐H295 line, which was developed from a human adrenocortical carcinoma (HAC). Herein, we attempted a similar approach with the goal of isolation and characterization of clonal cell lines from a HAC. To determine if the HAC cells represent a model for adrenal glomerulosa, the cells were treated with angiotensin II (Ang II) and potassium (K+), followed by examination of aldosterone production and steroidogenic enzyme mRNA expression. Treatment with Ang II (10 nM) and K+ (18 mM) increased aldosterone production in a time‐dependent manner. Ang II and K+ increased expression of mRNA encoding all enzymes needed for aldosterone synthesis; namely StAR, CYP11A, HSD3B2, CYP21, and CYP11B2. In addition, the cells expressed mRNA for the Ang II type 1 receptor and its expression was increased upon treatment with Ang II and K+. The current study describes the development of a novel human adrenal cell line with responses seen in normal glomerulosa cells. The HAC cell line should provide an important alternative model system for defining the molecular mechanisms regulating aldosterone production.
Integrative mechanisms of blood pressure regulation in humans and rats: cross‐species similarities
Authors: Charkoudian Nisha , Gusman Elle , Joyner Michael J. , Wallin Gunnar , Osborn John
Published: June 2021

Inter‐individual variability in hemodynamic variables appears to be important in human blood pressure regulation. It is not known whether the rat, often used for studies of cardiovascular regulation, exhibits similar inter‐individual variability. In the present study, we evaluated whether inter‐individual variability in cardiac output (CO) and total peripheral resistance (TPR) is similar in humans and in rats, and whether a balance between these variables is similar in the two species. We assessed the relationship between CO and TPR during control conditions in humans (N = 40) and in Sprague‐Dawley rats (N = 16). We further assessed whether the relationship between CO and TPR was altered during acute increases in blood pressure, which were accomplished in humans with acute systemic L‐NMMA (N = 17), and in rats with a DOCA‐salt model (N = 16). Both humans and rats showed marked inter‐individual variability in CO and TPR but low variability in blood pressure. In both species, CO and TPR showed strong inverse correlations during normotension (r2 = 0.88 (humans) and 0.89 (rats)) and during blood pressure increases (r2 = 0.91 (humans) and 0.58 (rats)). We conclude that inter‐individual variability CO and TPR, and the balance between these variables, are important aspects of blood pressure regulation in both humans and rats.

Supported by NIH HL32352, HL83947, HL64176, UL1‐RR24150 and Swedish Research Council 12170.
Intermediate‐ and long‐term cardiovascular responses to gravitational stress – role of the vestibular system
Authors: Raffai Gábor , Csekõ Csongor , Nádasy György L , Monos Emil
Published: June 2021

This study investigated the role of vestibular apparatus on cardiovascular responses under various gravitational stresses in adult male telemetered Wistar rats. Arterial blood pressure (BP) and heart rate (HR) responses to 45º head‐up (HUT) and head‐down (HDT) tilt performed in tubular cages for 2 or 24 hour duration were compared using intact rats and rats with bilateral vestibular lesions. The sustained 2 hour hypertensive responses to HUT (8.4 mmHg) and HDT (10.6 mmHg) were abolished by vestibular lesions. HR was not influenced in either of the cases. HUT for 24 hours also resulted in sustained rise of BP (5.5 mmHg) which was accompanied with tachycardia (27.3 BMP). Vestibular lesion fully eliminated this BP response too, while that of the HR remained unaltered (35 BPM). Long‐term HDT also caused arterial hypertension (9.4 mmHg), while tachycardia (32 BPM) was seen in the early period (<10 h) of tilting only. Both responses were suppressed by vestibular lesions. In conclusion, blockade of labyrinthine inputs attenuates cardiovascular responses elicited by intermediate‐ and long‐term gravitational stresses. However, other sensory inputs derived from non‐vestibular cues (e.g. proprioceptive, visual, visceral, cutaneous etc.) seem to be effective enough to maintain arterial blood pressure. (Hungarian OTKA T‐042670/2003 and ETT 128/2006).
Acute blood pressure lowering and vasoprotective effects of dietary nitrate
Authors: Webb Andrew , Patel N , Loukogeorgakis S , Okorie M , Aboud Z , Misra S , Rashid R , Miall P , Deanfield J , Benjamin B , Macallister R , Hobbs A , Ahluwalia A
Published: June 2021

Diets rich in fruits and vegetables reduce blood pressure (BP) and the risk of adverse cardiovascular events. However, the mechanisms of this effect have not been elucidated. Many vegetables possess a high nitrate content and we hypothesized that this might represent a source of vasoprotective NO via bioactivation to nitrite. In 14 healthy volunteers, approximately 3h following ingestion of a dietary nitrate load (500ml beetroot juice), BP was substantially reduced (Δmax‐10.4/8 mmHg); this effect correlated with peak increases in plasma nitrite, but not nitrate, concentration. The dietary nitrate load also prevented endothelial dysfunction induced by an acute ischemic insult in the human forearm and significantly attenuated ex vivo platelet aggregation in response to collagen and ADP. Interruption of the enterosalivary conversion of nitrate to nitrite (facilitated by bacterial anaerobes situated on the surface of the tongue), prevented the rise in plasma nitrite, blocked the decrease in BP and abolished the inhibitory effects on platelet aggregation, confirming that these vasoprotective effects were due to the activity of nitrite converted from the ingested nitrate. These findings suggest that dietary nitrate underlies the beneficial effects of a vegetable‐rich diet and highlights the potential of a ‘natural’, low cost approach for the treatment of cardiovascular disease.
Sympathoexcitatory outflow from the Rostral Ventrolateral Medulla is attenuated in aged Wistar‐Kyoto Rats
Authors: Yajima Yoshiharu , Komatsu Kazutoshi , Suzuki Toshirou , Kanai Takashi , Hiratsuka Makoto , Ito Satoru , Tsukamoto Kazuyoshi , Hirayama Atushi
Published: June 2021

The rostral ventrolateral medulla (RVLM) vasomotor neurons are responsible for maintaining the tonic excitation of sympathetic preganglionic neurons. It is well known that the aging causes the physiological alternation in cardiovascular regulation. However, the cardiovascular influence of central vasomotor neurons in the advanced aged animals are not fully examined. In this study, using more than 80 weeks old Wistar‐Kyoto rats (WKY) as high aged model, we sought to determine the cardiovascular response of RLVM in glutamatergic stimulation. In alpha‐chloralose‐anesthetized animals, microinjection of glutamate into unilateral RVLM were performed. In elderly WKY rats, although the intravenous injection of sodium nitroprusside or phenylephrine evoked similar response of the groups of animals in 16–20 weeks old, the response evoked by microinjection of glutamate into RVLM were smaller than that of control. These results suggest the sympathoexcitatory influence from RVLM is attenuated in advanced aged animals.
Central but not peripheral G‐alpha i/o protein inhibition potentiates hypertension in hypertensive Dahl salt‐sensitive rats: a protective role for central G‐alpha i/o proteins
Authors: Wainford Richard David , Kapusta Daniel R
Published: June 2021

We have shown that intracerebroventricular (i.c.v.) pre‐treatment of rats with pertussis toxin (PTX) prevents the hypotensive and bradycardic responses to the Gαi/o‐protein coupled receptor ligands, nociceptin/orphanin FQ and clonidine. These studies examined whether central Gαi/o protein inhibition alters blood pressure in hypertensive Dahl salt sensitive (DSS) rats.

Methods DSS rats were implanted with radio‐telemetry probes and MAP and HR were continuously monitored. After 1‐week, rats were maintained on 8% NaCl chow for 3 weeks then administered either i.c.v. iso‐saline vehicle (N=6), i.c.v. PTX (1 μg) (N=6), or intraperitoneal (i.p.) PTX (15 μg/kg) (N=6).

Results DSS rats on 8% NaCl chow showed a significant increase in MAP (normal chow MAP, 130 ± 2 mmHg; 8% NaCl chow, 164 ± 8 mmHg, P<0.05). In hypertensive DSS rats, central PTX significantly increased MAP (day 4 post i.c.v. PTX; MAP Δ +25 ± 2 mmHg, P<0.05). In contrast, peripheral PTX administration produced a significant hypotension (day 4 post i.p. PTX; MAP Δ −58 ± 9 mmHg, P<0.05).

Conclusion In hypertensive DSS rats, endogenous central Gαi/o protein pathways participate in reducing the severity of high NaCl‐induced hypertension. This is in contrast to the role of peripheral Gαi/o proteins, which in these (DSS) and previous studies (spontaneously hypertensive rats) play a pathological role in the maintenance of hypertension. Thus, endogenous central vs. peripheral Gαi/o proteins have differential affects on blood pressure control and potentially in the etiology of hypertension. DK43337, HL 071212, P20 RR018766
Norepinephrine release during orthostasis is similar in healthy individuals who are and are not susceptible to syncope
Authors: Fu Qi , Shibata Shigeki , VanGundy Tiffany B , Okazaki Kazunobu , Levine Benjamin D
Published: June 2021

Previous studies showed that individuals susceptible to syncope had low norepinephrine release during orthostasis. However under most circumstances, presyncopal subjects were returned to supine first, then blood samples were drawn, while blood was collected upright in those who didn't have presyncope. We tested the hypothesis that plasma norepinephrine is similar between individuals who are and aren't susceptible to syncope after returning to supine from upright. Muscle sympathetic nerve activity, blood pressure, and heart rate were recorded in 35 healthy young subjects who had no previous history of fainting, both supine and during 45‐min of 60° upright tilt. Blood samples were drawn supine, during tilt, and immediately after the subject was returned to supine either because they completed the prolonged tilt or developed presyncope. Fourteen had presyncope (fainters), while 21 completed the tilt (non‐fainters). There were no differences in any variable between fainters and non‐fainter supine and during acute tilt, though sympathetic withdrawal and a rapid decrease in blood pressure occurred at presyncope in all fainters. Plasma norepinephrine in the supine position (P=0.222), after 5 (P=0.234) and 20 min of tilt (P=0.669), and after returning to supine (P=0.305) were comparable between fainters and non‐fainters. These results suggest that norepinephrine release during orthostasis is similar between individuals who are and are not susceptible to syncope. The specific trigger precipitating syncope (sympathetic withdrawal in all fainters) remains unknown.
Effect of severe, nonhypotensive hypovolemia on heart rate in rats
Authors: Stedenfeld Kristen A , Hoffmann Myriam L , Sved Alan F , Stricker Edward M
Published: June 2021

The present experiments evaluated the effects of progressive, severe hypovolemia on heart rate (HR) and mean arterial blood pressure (MAP) in adult rats. HR and MAP were continuously monitored with telemetry. Subcutaneous injection of a 30% polyethylene glycol (PEG) solution was used to produce a volume loss of 4–5% per hour for 5–6 h, a rate that subsequently slowed with volume deficits asymptoting at ~40% during hours12–24 post‐injection. In all animals, MAP was stable at ~110 mm Hg throughout the 24‐h test period. In contrast, HR increased gradually by ~50 bpm during the first 4 h from basal values of 370–400 bpm, and then it fell by 150–200 bpm during the next 4 h before gradually rebounding during the next 12 h. These results allow the hypotheses that the initial tachycardia reflected excitation of the sympathoadrenal system, the subsequent fall in HR reflected an inhibition of the sympathetic nervous system associated with a large and sustained drop in plasma volume unrelated to changes in MAP, and the later restoration of HR reflected a return of sympathetic activity when the rate of volume loss slowed. Similar to these findings, previous studies have shown that bradycardia follows tachycardia after severe hemorrhage; consequently, the present results suggest that this pattern results from the induced change in blood volume. Ongoing experiments are evaluating these hypotheses.
Peripheral chemoreflex activation in conscious mice
Authors: Burmeister Melissa A. , Braga Valdir A. , Sharma Ram V. , Davisson Robin L.
Published: June 2021

Peripheral chemoreceptor activation with potassium cyanide (KCN) in conscious rats produces a sympathoexcitatory/pressor response, bradycardia and tachypnoea, which are potentiated during hypertension (HTN). However, this reflex has not been studied in conscious mice. We hypothesized that stimulation of peripheral chemoreceptors would evoke similar cardiovascular (CV) responses in mice, which would be altered during HTN. Mice were instrumented with intravenous (i.v.) catheters for drug delivery, and heart rate and blood pressure were measured by telemetry. Compared to saline (n=6), i.v. KCN (0.03%, n=6) elicited a pressor response (KCN: 25±3; sal: 5±1 mmHg, p<0.05) followed by hypotension (KCN: −31±5; sal: −4±2 mmHg, p<0.05). KCN also produced marked bradycardia (KCN: −218±30; sal: −9±8 bpm, p<0.05). The depressor and bradycardic responses were abolished by pre‐treatment (pt) with atropine (i.p.) (4±1 mmHg & 20±35 bpm, p<0.05), and the pressor response was abolished by pt with prazosin (i.p.) (3±1 mmHg, p<0.05). Similar studies were performed in mice receiving angiotensin‐II (Ang‐II, 600 ng.kg−1.min−1, s.c., n=6). The CV responses to KCN were unaltered after 2‐wks of Ang‐II infusion when mice were maximally hypertensive. These studies are the first to characterize the chemoreflex in mice and propose a new animal model for studying hypoxia‐related diseases such as obstructive sleep apnea.
Human muscle sympathetic action potential patterns during graded lower body negative pressure
Authors: Liu Yinchun , Shoemaker J. Kevin
Published: June 2021

This study assessed baroreflex modulation of within‐burst neural action potential (spike) discharge in muscle sympathetic nerve activity (MSNA). Spike rate and inter‐spike interval were exposed from raw MSNA recorded from 6 healthy individuals during graded lower body negative pressure (LBNP) (baseline, −10, −40, −60 mmHg). The first 30s raw MSNA of the 4th min for each LBNP level was analyzed after applying a spike detection method (Autonomic neuroscience: Basic & Clinical 143(2007) 92–105). Spikes/30 sec increased progressively with LBNP beginning at −10 mmHg (P<0.05), with a doubling of spike rate (range = 40 – 290%) at −60 mmHg. Mean inter‐spike interval decreased progressively from 0.175±0.045 s at Baseline to 0.087±0.028 s at −60 LBNP (P<0.05) reflecting increases in average discharge frequencies from 5 to 10 Hz. In probability distributions, progressive increases in the proportion of spikes occurred only within the shortest inter‐spike interval bin (ranging 0.04 (25 Hz) – 0.1 (10 Hz) s across participants). Thus, the increment in MSNA with baroreceptor unloading is focused at high‐frequency discharge patterns and reflects progressive increments in action potential content with evidence for a doubling of neurons recruited between baseline and high levels of orthostatic stress.

Supported by NSERC‐CHRP
Reactive Oxygen Species Mediate Angiotensin‐II (Ang‐II)‐Dependent Activation of Calcium/Calmodulin Dependent Kinase II (CaMKII) in Forebrain Circumventricular Organs (CVOs)
Authors: Peterson Jeffrey , Stupinski John A. , Sharma Ram V. , Davisson Robin L.
Published: June 2021

Reactive oxygen species (ROS) play a crucial role in central Ang‐II signaling. However, the downstream effectors of Ang‐II‐induced ROS in neurons are not known. Here, we tested the hypothesis that Ang‐II‐dependent activation of CaMKII in CVOs is redox‐sensitive. First, Neuro‐2A cells were treated with adenovirus expressing CuZnSOD (AdCuZnSOD), a control vector (AdLacZ), or vehicle for 24 hrs and stimulated with Ang‐II (1μM, 30 min). CaMKII activation was measured by immunofluorescence using an antibody against the autophosphorylated form (pCamKII). Ang‐II induced a marked increase in pCaMKII levels in control cells which was significantly inhibited by treatment with AdCuZnSOD (saline 1.5±.02, AdLacZ 1.7±.06 vs AdCuZnSOD 1.2±.04 fold vs baseline, p<.05, n=27–61 cells). We next examined the role of ROS in CaMKII activation in forebrain CVOs during slow‐pressor Ang‐II infusion. Following forebrain CVO‐targeted delivery of AdCuZnSOD or AdLacZ, mice underwent either saline or slow‐pressor Ang‐II infusion. CaMKII activation was assessed by western analysis for pCaMKII in pooled (n=3) micropunches from the lamina terminalis taken at day 16 during the infusions. In AdLacZ‐treated mice, Ang‐II induced a significant (1.7±.02 fold) increase in CaMKII activation in forebrain CVOs as compared to saline. Importantly, treatment with AdCuZnSOD blunted this rise in activated CaMKII (1.1±.04 fold increase vs. saline, p<.05, n=3). These studies demonstrate a key role for ROS in Ang‐II‐dependent CaMKII activation in neurons, and implicate CaMKII as a potential downstream effector of Ang‐II‐induced ROS in the development of neurogenic hypertension.
Endoplasmic reticulum stress signaling modulates smooth muscle phenotypes
Authors: Shen Hua , Manabe Ichiro , Eguchi Kosei , Fujiu Katsuhito , Nagai Ryozo
Published: June 2021

Metabolic syndrome is a major risk factor for atherosclerotic cardiovascular disease. Increased circulating free fatty acid (FFA) levels have been suggested to directly contribute to insulin resistance. Moreover, FFAs and changes in their contents have been shown to be associated with higher risks for cardiovascular disease. However, little is known regarding effects of FFA on vascular cells. Endoplasmic reticulum (ER), in which proteins are synthesized and folded, is increasingly regarded as a major site that responds to various stress, such as cellular energy levels, redox state, and hypoxia. We found that palmitate, a major free fatty acid in plasma, elicited ER stress markers including Bip, spliced XBP, and CHOP in cultured vascular smooth muscle cells (SMCs). Moreover, infusion of palmitate induced ER stress response in aorta in mice. Palmitate treatment resulted in upregulation of KLF5, PDGF, MMP3, and MMP9, whose expression are characteristics to phenotypic modulated SMCs. The ER stress attenuator, tauroursodeoxycholate (TUDCA), suppressed ER stress response as well as the upregulation of the genes involved in phenotypic modulation. Taken together, palmitate induced phenotypic modulation and proinflammatory gene programs in SMCs. ER stress response signaling appears to play an important role in mediating proinflammatory signals in SMCs in metabolic syndrome.
Glucosamine Inhibits TNF‐α‐Induced Expression of Inflammatory Mediators In Rat Aortic Smooth Muscle Cells through Inhibition of NFκB Activation
Authors: XING DONGQI , Feng Wenguang , Chen Yiu‐Fai , Chatham John C , Oparil Suzanne
Published: June 2021

Objective:

This study tested whether glucosamine (GlcN), an amino suger that increases O‐GlcNAc modification of proteins by increasing flux through the hexosamine biosynthesis pathway, can protect vascular smooth muscle cells (SMCs) against inflammatory stress.

Methods and Results:

Quiescent cultured rat aortic SMCs (RASMCs) were pretreated with GlcN (5 mM) or vehicle for 1 hr and then stimulated with tumor necrosis factor (TNF)‐α (10 ng/ml) for another 15 min or 6 hrs. Total and phosphorylated IκBα were measured by western blot analysis. NFκB DNA binding activity were determined by ELISA. RNA was extracted for real‐time RT‐PCR analysis of chemokine and adhesion molecule expression. Nucleocytoplasmic protein subjected to western blot analysis of global O‐GlcNAcylated protein using the selective CTD110.6 antibody. GlcN treatment enhanced overall O‐GlcNAcylated protein levels and inhibited chemokine and adhesion molecule expression, IκBα phosphorylation, prevented the degradation of IκBα and inhibited NFκB activation in TNF‐α‐stimulated RASMCs.

Conclusions:

O‐GlcNAc modification may protect the vasculature from inflammatory stresses.

This work was supported, in part, by American Heart Association Greater Southeast Affiliate grant 0425455B, 0765398B.
Inhibition of Poly (ADP‐ribose) Polymerase (PARP) by PJ‐34 regulates angiogenesis and VEGF‐induced MAPK‐signalling
Authors: Olah Gabor , Pyriochou Anastasia , Papapetropoulos Andreas , Szabo Csaba
Published: June 2021

Angiogenesis is a tightly regulated process known to be essential for normal embryonic development but also in physiological and pathological phenomena in the adult. Excessive and deregulated angiogenic response is thought to contribute to cancer, diabetic retinopathy, arthritis and psoriasis. The mitogen‐activated protein kinase (MAPK) pathway plays an essential role in the regulation of proliferation and survival. PARP is a multifunctional nuclear enzyme that also regulates transcription by affecting histones, DNA methylation and enhancer/promoter regions. Here we investigate the effects of poly (ADP‐ribose) polymerase (PARP) inhibition on angiogenesis and MAPK‐signalling. The results show that PARP inhibition with PJ34 reduces endothelial cell proliferation, migration and organization leading to decreased angiogenesis. PARP inhibition also decreased VEGF‐induced Akt, ERK1/2 and p38 phosphorylation. PARP‐1 inhibitors might be useful in disease states when suppression of angiogenesis and phosphorylation of MAPK‐pathway member is required.
Hemodynamic patterns and vascular differentiation in embryonal development.
Authors: Styp‐Rekowska Beata , Tabuchi Arata , Le Noble Ferdinand , Pries Axel Radlach
Published: June 2021

Differentiation of blood vessels into either arteries or veins is not sufficiently understood. Whereas the earliest vascular development appears to be controlled exclusively on the gene level, in later embryonic stages vessels are influenced by hemodynamic and metabolic stimuli. To investigate the role of hemodynamic patterns on vascular development in different vascular beds, we aquired flow patterns in zebrafish embryos and chick chorion allantois membranes (2–4 days post fertilization, dpf). In allantois membrane we observed high pulsatile changes of shear combined with low averaged shear stress in arteries and not in veins. The main vessels of zebrafish embryos undergo a “hemodynamic switch” at 3 dpf. While at 2 dpf arteries and veins exhibit similar hemodynamic profiles with high pulsatility and accelaration, at day 4 vein profiles change and show low amplitude and accelaration. The data suggest the differences of hemodynamic profiles during embryonal development could trigger expression of mechano‐sensitive genes and support arterial‐venous differentiation.
Qualitative estimation of the waters of the river Llap according to algae
Authors: Kurteshi Kemajl , Vehapi Idriz , Ismaili Muharrem , Amiti Sulltan , Sahiti Naim , Ibrahimi Arsim
Published: June 2021

During the summer 2003 we investigate the algae of the upstream part of the river Llap. We taken the samples from 4 locality (Rep,Kërpimeh, Bajqin and Podujevë). We determinate the composition of the algae: 3 species belong divission Cyanophyta , 45 species belongs Bacillariophyta, 3 species Euglenophyta and 7 species Chlorophyta, altogether 58 species. According the bioindicator species we are done the qualitative estimation of waters of river Llap.Based on the presence of species as saprobiologic bioindicator and saprobic level , the waters of river Llap belongs the second level of bonity respectively beta‐mesosaprob
A computational model for biochemical Interaction of NO and reactive oxygen species in paired arteriole‐venule
Authors: Kavdia Mahendra
Published: June 2021

All cells of vascular tissue including endothelial cells, and smooth muscle cells have enzyme systems that produce reactive oxygen species (ROS). Important ROS generated in the vascular tissue are superoxide or hydrogen peroxide. Superoxide can react with NO to form peroxynitrite. Antioxidant systems protect against damage from ROS in normal physiological conditions. When ROS generation overwhelms the antioxidant defense (known as oxidative stress), these radicals can alter cellular function by interacting with DNA, RNA, and fatty acids, contribute to impairment of endothelium‐dependent vasodilation and lead to apoptosis. In this study, a computational model to describe the interactions of NO, superoxide and peroxynitrite in a tissue containing an arteriole/venule pair is presented. We simulated two sources of oxidative stress 1) NADPH oxidases and 2) adhered/migrated leukocytes. Model results predicted that 1) an increase in oxidative stress in the venule decreased NO levels in regions of both venule and neighboring arteriole, and 2) the amount of NO reduction varied depending on the location of oxidative. The results provide a comprehensive quantitative knowledge of NO and ROS interactions in vascular tissue, which is an important step towards understanding vascular complications in many pathological conditions.
NMDA Receptors Mediates Homocysteine‐Induced Sclerotic Action on Rat Mesangial Cells
Authors: Yi Fan , Abais Justine , Jin Si , Zhang Fan , Li Ningjun , Li Pin‐Lan
Published: June 2021

The N‐methyl D‐aspartate receptors (NMDAr) are a glutamate receptor that has been extensively studied in the central nervous system and is suggested as a homocysteine (Hcys) binding receptor. However, the role of NMDAr in the Hcys‐induced renal glomerular dysfunction or injury is still unknown. The present study determined whether the NMDAr is present in rat mesangial cells (RMCs) and whether these receptors are involved in Hcys‐induced glomerular injury. Real‐time RT‐PCR and Western blot analysis confirmed that NMDAr subunits are present both in RMCs and in rat kidneys. Dizocilpine (MK801), a noncompetitive antagonist of the NMDA receptors, blocked the Hcys‐induced Rac GTPase activation by 46%, which was accompanied by a 66% reduction of Hcys‐induced NADPH oxidase dependent superoxide (O2‐.) production as measured by Electron Spin Resonance (ESR) spectrometric assay. Correspondingly, MK801 also attenuated Hcys‐increased expression of tissue inhibitor of metalloproteinase‐1 (TIMP‐1) and consequently led to increase in matrix metalloproteinase‐1 (MMP‐1) activity. These results indicate that NMDAr exist in rat mesangial cells and mediate, at least in part, Hcys‐induced enhancement of local oxidative stress and abnormal metabolism of extracellular matrix components in rat mesangial cells (Supported by NIH Grants HL57244, HL75316, DK‐54927 and HL89563).
Efficacy and potency of alpha adrenergic receptor agonists at inhibiting rat hippocampal CA3 network activity
Authors: O'shea Lorraine , Boese Sarah , Xu Ke , Nelson Brian , Goldenstein Brianna , Pribula Jacquline , O’Brien Jasmine , Davis Kylie , Laverdure Floyd , Schulte Christina , Schlosser Danielle , Luger Elizabeth , Jurgens Chris , Porter James , Van Doze
Published: June 2021

α2 adrenergic receptors (ARs) are involved in many physiological processes, including mediating the antiepileptic effects of the catecholamine norepinephrine (NE). We generated data from agonists of α2 ARs. We hypothesized the performance of catecholamines to be higher than synthetic compounds of imidazolines and guanidines. Using extracellular recordings in the hippocampal CA3 region of rat brain slices, antiepileptiform actions of agonists of these chemical classes were recorded. Variations of the potency and efficacy of the agonists were considered with the parameters of their structural differences. Agonist classifications (full vs. partial) were based on relative efficacy to the endogenous neurotransmitter NE. Results indicated that catecholamines were full agonists. Among imidazolines and guanidines, UK‐14303 was the most efficacious. These results confirm our hypothesis that catecholamines were more efficacious. However, imidazolines and guanidines had higher potencies. Since differences in structures between catecholamines were trivial while structural differences among classes of agonist were significant, conclusions can be drawn that structural attributes of catecholamines contribute to their efficacy and potency at inhibiting hippocampal epileptiform activity. Supported by NSF ND EPSCoR, NSF CAREER, NSF REU Site, NIH, Epilepsy Foundation, American Physiological Society.
Mechanism of vasodilatation with N‐acetylcysteine and NaHS: Contribution of ATP‐sensitive potassium channels
Authors: Yamamoto Michiko , Adachi Takeshi , Ishiwata Kyoko , Ikeda Satsuki , Hirayama Yoshiaki , Soga Tomoyoshi , Suematsu Makoto
Published: June 2021

Objective: Hydrogen sulfide (H2S) is known as a toxic gas, however, recent studies reveal H2S as the third gaseous vasodilator, following nitric oxide and carbon monoxide. N‐acetylcysteine (NAC) is often used as an anti‐oxidant and it can increase intracellular cysteine (Cys) levels to produce H2S. In this study we tested the contribution of ATP‐sensitive potassium channels (IKATP) to NAC‐ and NaHS‐induced relaxation and the metabolic changes induced by H2S.

Methods: Vascular function was assessed with isometric tension measurement of aortic rings equipped in the organ bath. After aortic constriction with phenyrephrin, a dose‐dependent relaxations to NAC/NaHS (0.3–10 mM) were measured. Some rings were pre‐treated with L‐NAME (100 μM), ODQ (10 μM), or glybenclamide (GLB: 10 μM), an IKATP blocker. Aortic homogenates were prepared and vascular metabolome analysis was performed with capillary electrophoresis/mass spectrometry. Results: Neither L‐NAME nor ODQ affected the aortic relaxations to NAC/NaHS. GLB decreased the maximum relaxation to NaHS. Notably, GLB decreased the speed of relaxation to NAC/NaHS in the early phase. NaHS increased the vascular AMP levels about five folds (31.4±5.8 vs 158.5±31.6 nM).

Conclusion: Aortic relaxations to NAC/NaHS are involved with hyperpolarization via the activation of IKATP. An increase in intracellular AMP levels by H2S, activates IKATP to modulate vascular tone.
Loss of O‐GlcNAc transferase activity sensitizes cardiac myocytes to post‐hypoxic death
Authors: Ngoh Gladys Afor , Watson Lewis J. , Jones Steven P.
Published: June 2021

We recently reported that diabetic hearts have a paradoxical reduction in β‐O‐linked‐N‐acetylglucosamine (O‐GlcNAc) signaling. Because enhancement of O‐GlcNAc signaling is cardioprotective, we hypothesized that loss of OGT activity is sufficient to sensitize nondiabetic cardiac myocytes to post‐hypoxic injury. Here, loxP‐ flanked OGT neonatal mouse cardiac myocytes (n=3/group) were infected with an adenovirus carrying cre recombinase gene (AdCre, 72hours; 50 moi). Expression of cre recombinase reduced OGT expression (44+/−16%, p<0.05) and O‐GlcNAc levels (28+/−5%, p< 0.05) according to western blotting. Furthermore, such transcriptional silencing of OGT significantly (122+/−7%, p< 0.05) exacerbated post‐hypoxic injury according to LDH release. To further confirm these findings, neonatal rat cardiac myocytes were transfected with 30nM OGT siRNA, to knockdown OGT. OGT siRNA‐treatment significantly reduced OGT (49+/−8%, p<0.05) and O‐GlcNAc levels (75+/−13%, p<0.05) compared with control siRNA. Such decrement in O‐GlcNAc signaling corresponded with exacerbated post‐hypoxic LDH release (120+/−2%, p<0.05). We conclude that reduced OGT activity diminishes O‐GlcNAc levels and sensitizes cardiac myocytes to lethal cell injury. Such insights support the hypothesis that defects in O‐GlcNAc signaling are at least partially responsible for the sensitivity of the diabetic myocardium to ischemia‐reperfusion injury. Moreover, basal OGT expression is elemental for the ability of cardiac myocytes to withstand hypoxic injury. NIH (HL0833220), AHA (0535270N) and AHA (0715493B).
HIF‐1 specific prolyl hydroxylases in elite athletes
Authors: Rundqvist Helene , Klintberg Malene , Norrbom Jessica , Gustafsson Thomas , Sundberg Carl Johan
Published: June 2021

The hypoxia sensitive transcription factor HIF‐1 has been suggested to be involved in skeletal muscle adaptation to endurance‐type exercise training. HIF‐1 protein levels are regulated by hydroxylation of critical proline residues and subsequent protein degradation of the alpha subunit. Three prolyl hydroxylation domain‐containing proteins (PHD1‐3) have been identified in mammalian cells, PHD2 appears to be the predominant form that regulates HIF‐1alpha protein stability in vivo1. HIF‐1 is stabilised in response to an acute bout of exercise in sedentary individuals2 but there is debate regarding the activity of the HIF‐1 system in response to long term training. Skeletal muscle biopsies were obtained from the vastus lateralis muscle at rest from 12 males with high oxidative capacity and 9 normally trained male controls. PHD2 mRNA displayed the highest expression levels of the three forms in human skeletal muscle, regardless of training status. Furthermore, PHD2 and 3 mRNA levels were both higher in elite athletes compared to the control individuals and PHD2 protein levels were significantly higher in the elite group. Conclusion: The HIF‐1 regulating prolyl hydroxylase PHD2 was significantly higher in elite athletes, which may explain an attenuated HIF‐1 activity in response to long term endurance training. This work was supported by the Swedish Research Council, the Swedish National Centre for Research in Sports
Resistance and endurance training differentially affect myofibrillar and mitochondrial protein synthesis at rest and following exercise in human skeletal muscle
Authors: Wilkinson Sarah Brianne , Phillips Stuart M. , Atherton Philip J. , Patel Rekha , Yarasheski Kevin E. , Chen Jennifer , Rennie Michael J.
Published: June 2021

We aimed to determine the acute and chronic changes in myofibrillar (MYOFIB) and mitochondrial protein synthesis rates (MITO) with aerobic exercise (AE) and resistance exercise (RE). We infused d3‐α‐KIC at REST and for 4 hours after an acute bout of RE or AE prior to (UT) and following 10 weeks (TR) of training in healthy 20 yr old men. In the UT state, both MYOFIB (REST: 0.06±0.01, RE: 0.10±0.01 %/h, p=0.01) and MITO (REST: 0.08±0.02, RE: 0.14±0.04 %/h, p=0.02) increased following RE. Following TR only MYOFIB increased with RE (REST: 0.08±0.01, RE: 0.11±0.01 %/h, p=0.05), while MITO did not change from resting values (p=0.43). AE stimulated MITO (UT REST: 0.07±0.02, AE: 0.18±0.03, TR REST 0.07±0.01, AE: 0.15±0.03 %/h, p<0.05) but not MYOFIB regardless of training. In conclusion, RE acutely increases both MYOFIB and MITO, while AE increases MITO only. Acute responses to RE appear to be redundant in that synthesis of both MYOFIB and MITO were both stimulated. Following TR, the rate of MYOFIB was increased at rest with RE, and also following an acute bout of RE. By contrast AE activated only MITO in both the UT and TR states. These protein fraction responses are exercise specific and indicate the responses of mixed muscle protein synthesis are likely heavily influenced by exercise mode and training status. Supported by NSERC, CIHR, UK BBSRC, NIH‐NCRR.