bluuebunny/crossref_metadata_2025 huggingface dataset:
Garlic Extract Diallyl Sulfide (DAS) Activates Nuclear Receptor CAR to Induce the Sult1e1 Gene in Mouse Liver
Authors: Sueyoshi Tatsuya , Green William D. , Vinal Kellie , Woodrum Tyler S. , Moore Rick , Negishi Masahiko
Published: June 2021

Constituent chemicals in garlic extract are known to induce phase I and phase II enzymes in rodent livers. Here we have utilized Car+/+ and Car−/− mice to demonstrate that the nuclear xenobiotic receptor CAR regulated the induction of the estrogen sulfotransferase Sult1e1 gene by diallyl sulfide (DAS) treatment in mouse liver. DAS treatment caused CAR accumulation in the nucleus, resulting in a remarkable increase of SULT1E1 mRNA (3,000 fold) and protein in the livers of Car+/+ females but not of Car−/− female mice. DAS also induced other CAR‐regulated genes such as Cyp2b10, Cyp3a11 and Gadd45β. Compared with the rapid increase of these mRNA levels which began as early as 6 hrs after DAS treatment, the SULT1E1 mRNA began increasing its levels after 24 hrs. This slow response to DAS suggested that CAR required an additional factor to activate the Sult1e1 gene or that this activation was indirect. Despite the remarkable induction of SULT1E1, there was no decrease in the serum levels of endogenous E2 or increase of estron sulfate while the clearance of exogenously administrated E2 was accelerated in DAS treated mice.
Identification of Constitutive Androstane Receptor (CAR) Activators by Integrated Computational and Biological Approaches
Authors: Lynch Caitlin , Pan Yongmei , Swaan Peter , Wang Hongbing
Published: June 2021

The nuclear receptor CAR is a well established xenobiotic sensor governing the transcription of numerous hepatic genes associated with xenobiotic metabolism and clearance. Recent evidence suggests that mouse CAR also functions as a mediator of energy metabolism and cancer development. However, whether its human counterpart holds these novel functionsis largely unknown due to the lack of selective human (h) CAR modulators. Here, we have identified several novel hCAR activators by applying 1) virtual screening of chemical databases with ligand‐based structure‐activity models; and 2) cell‐based assays that proved to be efficient in the determination of hCAR activation. Initial screening of the CDD drug database using structure‐activity models and docking has resulted in 20 lead compounds with optimal pharmacophore parameters. In cell‐based reporter assays, two of the 20 leads exhibited robust activation of hCAR. In human primary hepatocyte cultures, these two compounds efficiently translocated hCAR from cytoplasm to nucleus, which has been recognized as the initial step of CAR activation. Moreover, these compounds also induced the expression of the prototypical hCAR target genes, CYP2B6 and CYP3A4. These results clearly indicate that our combined strategy is efficient in the identification of novel hCAR modulators. This work was partly supported by NIH Grant (DK061652).
Pulmonary effects of inhaled oil dispersant (COREXIT® EC9500A) in rats
Authors: Roberts Jenny R , Reynolds Jeffrey S , Thompson Janet A , Goldsmith William T , Jackson Mark , McKinney Walter , Frazer David G , Zaccone Eric J , Shimko Michael J , Kashon Michael L , Castranova Vincent , Fedan Jeffrey S
Published: June 2021

COREXIT® EC9500A (CE) was used to disperse crude oil during the 2010 Deepwater Horizon oil spill response. While the environmental impact of CE has been examined, the pulmonary effects of CE are unknown. Here we investigated whether inhaled CE elicits airway inflammation, alters pulmonary function, or affects airway reactivity to methacholine (MCh). Rats were exposed to CE (mean 27 mg/m3, 5 h) and examined 1 and 7 d post‐exposure. Under anesthesia, the lung lavage fluid was harvested. Lactate dehydrogenase (LDH) and albumin (A) were measured as indices of lung injury, recovered lung cells were differentiated (macrophages, neutrophils, lymphocytes, and eosinophils) to evaluate inflammation, and oxidant production by macrophages was measured. There were no significant differences in LDH, A, inflammatory cell levels or oxidant production between air‐ and CE‐exposed groups at either time point. Pulmonary function assessed in conscious animals indicated that neither breathing frequency nor specific airway resistance were altered at 1 and 7 d post‐exposure. Airway resistance responses to MCh aerosol in anesthetized animals were unaffected at 1 and 7 d post‐exposure, while dynamic compliance responses were decreased at 1 d but not at 7 d post‐exposure. Our findings suggest that CE inhalation does not initiate lung inflammation, but may transiently increase the difficulty of breathing.
Down‐regulation of Smurf1 and Smurf2 is involved in the Initiation and Progression of Hepatic Fibrotic rats
Authors: ZHANG GUO , PENG PENG , CHEN LI , CHEN XINZHOU , ZHANG FACAN
Published: June 2021

The detail mechanisms undergoing degradation of TGFbeta/Smad molecules are poorly understood. This study was designed to determine if novel Smurfs‐dependent degradation machinery is involved in the hepatic fibrosis. Rats with hepatic fibrosis were induced by 40% carbon tetrachloride. Immunohistochemistry staining and western blot were performed to quantify the expression levels of Smurf1 and Smurf 2 in normal and fibrotic liver tissues. In parallel, the hepatic stellate cells (HSCs) marker Desmin and alpha‐SMA were dynamically measured as well. Furthermore, in vitro study was performed to elucidate the relationship among Smad3, Smad7, alpha (2) I collagen and Smurf1&2 levels in cultured HSCs by using specific siRNA to block the Smurf1&2 mRNA. It was found that either Smurf1 or Smurf2 was enriched in normal hepatic Disse space. However, the expression levels of alpha‐SMA but not Desmin, Smurf1 or Smurf2 have significantly increased in fibrotic rats (P<0.01). In those rats with hepatic fibrosis, both Smurf1 and Smurf2, particular Smurf2, were found to markedly decrease (P<0.01). Both decrement of Smurf1 and Smurf2 are associated to the increasing levels in phosphorylation of Smad3 as well as alpha (2) I collagen mRNA in these cells. Our data indicate that down‐regulation of Smurf1&2 in liver may importantly initialize and participate in the process of hepatic fibrosis. (Supported by NSFC 30960145 and GHF 200958, zhangfacan@126.com)
Isoliquiritigenin, an antioxidant flavonoid from licorice, inhibits high‐fat diet‐induced hepatic steatosis and oxidative injury through JNK1 inhibition
Authors: Kim Tae Hyun , Kim Young Mi , Kim Young Woo , Yang Yoon Mee , Kim Sang Geon
Published: June 2021

Isoliquiritigenin (ILQ), a flavonoid originated from Glycyrrhizae species, has an anti‐oxidant effect. This study investigated the potential of ILQ for inhibiting liver X receptor‐alpha (LXRalpha)‐mediated lipogenesis and steatosis in hepatocytes, and the underlying molecular basis. Treatment of ILQ antagonized the ability of LXRalpha agonist (T0901317) to activate sterol regulatory element binding protein‐1c, thereby repressing downstream lipogenic genes induction. ILQ treatment inhibited activating phosphorylation of JNK1 elicited by palmitate or TNFalpha. JNK1, but not JNK2, increased LXRalpha phosphorylation at serine residues, promoting LXRalpha activation. The ability of ILQ to inhibit JNK1 led to the repression of T0901317‐inducible LXRalpha activation. In mice fed a high‐fat diet, ILQ treatment inhibited hepatic steatosis, as shown by a decrease in fat accumulation and repression of lipogenic genes. The results of blood chemistry and histopathology confirmed attenuation of high‐fat diet‐induced liver injury by ILQ. Moreover, ILQ inhibited oxidative stress, as shown by decreases in thiobarbituric acid reactive substance formation, iNOS and COX‐2 induction, and nitrotyrosinylation. Our results demonstrate that ILQ has the ability to repress LXRalpha‐dependent hepatic steatosis through JNK1 inhibition, and protect hepatocytes from oxidative injury induced by fat accumulation.
Therapeutic rescue of mice treated with a lethal dose of acetaminophen
Authors: Banerjee Sudip , Latendresse John R , Mehendale Harihara M
Published: June 2021

Earlier studies (Bhave et al., Toxicol. Appl. Pharmacol., 228: 225–238, 2008) revealed that secretory phospholipaseA2 (sPLA2) is a death protein that mediates progression of injury initiated by hepatotoxicants such as acetaminophen (APAP), CCl4, and thioacetamide. The objective of present work was to inhibit sPLA2 at 1 h after liver injury was initiated by a lethal dose (600 mg/kg, i.p. in 0.45N NaCl2) of APAP administered to male Swiss Webster mice. Survival and mortality were recorded every 12 hours for 14 days. Mortalities were 50%, and 80% at 12 and 24h, respectively, after APAP injection. No mortality occurred in mice receiving the sPLA2 inhibitor (sPLA2inh) until 72h. At 14 days, cumulative mortality was only 10 % in the sPLA2inh group. Serum ALT and sPLA2 activities revealed very high liver necrosis and continued progression of liver injury in mice treated with APAP alone, while decreasing in mice receiving the sPLA2inh. These findings confirm the destructive role of sPLA2 by mediating the progression of liver injury. Our findings also reveal that it is possible to develop lifesaving therapeutic intervention administered well after the exposure to lethal overdose of APAP. Supported by Kitty Degree Endowment.
A Novel Fast Skeletal Muscle Activator, CK‐2017357, Improves Muscle Function in a Rodent Model of Myasthenia Gravis
Authors: Hinken Aaron , Driscoll Lena , Godinez Guillermo , Lee Kenneth , Pannirselvam Malar , Hartman James J , Muci Alex R , Morgans David J , Morgan Bradley P , Russell Alan J , Malik Fady I
Published: June 2021

Myasthenia gravis is a chronic autoimmune disease wherein the body produces antibodies that inhibit transmission at the neuromuscular junction, causing skeletal muscle weakness and fatigue. CK‐2017357 (CK‐357) is a small molecule activator of fast skeletal muscle that sensitizes the sarcomere to calcium and increases sub‐maximal force production. The objective of this study was to determine if administration of CK‐357 could increase muscle strength and improve functional capacity in a rodent model of myasthenia gravis. Female, Sprague‐Dawley rats injected with a single intra‐peritoneal dose of inhibitory acetylcholine receptor antibody displayed decreased tension‐generating capacity of intact, fast twitch muscle in situ and decreased conscious functional capacity assessed by forelimb grip strength. Single doses of CK‐357 improved conscious grip function in a dose‐dependent manner. In addition, CK‐357 increased force‐generating capacity and decreased tension decline during repeated stimulation of intact muscle in situ. Together these data suggest CK‐357 may ameliorate muscle weakness and fatigue associated with myasthenia gravis to provide a novel therapeutic approach for treatment of these patients. Supported, in part, by NIH Award 1RC3NS070670. Content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.
Mechanisms of nerve growth factor release from a human urothelial cell line
Authors: Ochodnicky Peter , Michel Martina B. , Michel Martin C.
Published: June 2021

Nerve growth factor (NGF) has been recently implicated in urinary bladder dysfunction, but the cellular source and mechanisms controlling its release have not been characterized yet. We studied NGF release and underlying signal transduction pathways in a human urothelial cell line UROtsa.

UROtsa expressed NGF at mRNA and protein level, as determined by Real‐time PCR and immunocytochemistry, respectively. NGF was constitutively released to the medium (11.5 ± 1.4 pg NGF/mg protein/h), as measured by ELISA. Down‐regulation of proteinkinase C by pre‐incubation with phorbol ester PMA reduced NGF release down to 71±3 % of the baseline within 24 hours. In contrast, NGF release was enhanced by stimulation with bradykinin, thrombin or brain‐derived nerve factor by 121±9, 127±9, 135±8 %, respectively. In case of bradykinin, enhanced NGF release was associated with elevated mRNA expression.

These data suggest that urothelial cells are able to release NGF, potentially modifiable by several G‐protein coupled and tyrosine kinase receptor ligands. Targeting these modulatory urothelial receptors may provide a novel strategy in the treatment of urinary bladder overactivity.

Supported by EU grant HEALTH‐F2‐2008‐223234.
In‐vitro antihistaminergic, anticholinergic, antioxidant and mast cell stabilizing effects of partially purified Labisia pumila leaf extract
Authors: okechukwu Patrick Nwabueze , Lee audrey jiajia , Akowuah Gabriel Akyirem
Published: June 2021

Labisia pumila is a popular medicinal plant in Malaysia and has been used in the treatments of various health complications. In this study, the in vitro anti‐histaminergic and anti‐cholinergic antioxidant and mast cell stabilizing effect of Labisia pumila crude leaf extract and partially purified fractions were investigated on guinea pig ileum. HPLC profiling of the bioactive extract were also examined. The crude extract was extracted using dichloromethane and partially purified into five fractions (A‐E) using column chromatography. The in vitro anti‐histaminergic and anti‐cholinergic studies were examined using organ bath containing Krebs‐Henseleit solution (pH 7.4) gassed with 95% O2 and 5% CO2 and maintained at 37°C. Two concentrations of the crude and partially purified fractions (1×10−6g/ml and 1×10−4g/ml) were challenged against a range of doses of histamine (10−6– 10−2M) and acetylcholine (10−8‐ 10−2M). Mast cell stabilizing effect was test using 0.5 ml subcutaneous injection horse serum along with 0.5ml of triple antigen containing 20000 million microorganisms. Determination of DPPH, TPC, hydroxyl, nitric oxide and superoxide radical‐scavenging activity was performed. HPLC analysis was performed using C18 column with 70% MEOH: 30%H2O as mobile phase and flow rate of 1 ml/min. The crude and partially purified fractions showed significant (p < 0.05) competitive dose‐dependent anti‐histaminergic and anti‐cholinergic, mast cell stabilizing and antioxidant activity. HPLC profiling revealed the presence of flavonoids (quercetin‐like, catechin‐like and kaempferol‐like compounds), alkaloids (berberine‐like compound), steroids, saponins, tannins and gallic acid‐like compound in the extract. Most of these phytochemicals have been reported to possess anti‐histaminergic, anti‐cholinergic, and antioxidant effects
5‐HT is unable to relax the isolated mesenteric artery: molecular and functional evidence
Authors: Pattison Jillian , Davis Robert Patrick , Thompson Janice , Watts Stephanie W.
Published: June 2021

Chronic serotonin (5‐hydroxytryptamine, 5‐HT) infusions resulted in a prolonged fall in blood pressure (BP) in the deoxycorticosterone acetate (DOCA)‐salt and sham rat. We hypothesized that 5‐HT lowers BP through direct receptor‐mediated vascular relaxation. The 5‐HT2B, 5‐HT1B, and 5‐HT7 receptors are linked with 5‐HT‐mediated vasorelaxation. mRNA (2‐deltaCt ×1000) for the 5‐HT2B (10.4 ± 2.32; 4.46 ± 0.442), 5‐HT1B (0.801 ± 0.108; 0.828 ± 0.239), and 5‐HT7 (0.208 ± 0.035; 0.779 ± 0.616) receptors was observed in DOCA and sham superior mesenteric arteries (SMA), respectively. Immunohistochemistry and western blotting validated the presence of the 5‐HT2B and 5‐HT7 receptors in the DOCA and sham SMA; the results for the 5‐HT1B receptor were unclear. Concentration response curves to BW‐723C86 (5‐HT2B agonist), 5‐CT (5‐HT1 agonist), and LP‐44 (5‐HT7 agonist) were performed in tissue baths in the presence of ketanserin (5‐HT2A antagonist). Maximum concentrations of BW‐723C86 and 5‐CT did not relax DOCA (118 ± 6.7%; 81; 180 ± 15% of half‐maximal U46619‐induced contraction) or sham (81 ± 7.1%; 88 ± 8.9%) SMA vs. H2O (60 ± 5.5% DOCA; 41 ± 8.3 % sham). Maximum concentrations of LP‐44 did not relax DOCA (75 ± 4.7%) or sham (55 ± 6.4%) SMA vs. DMSO (73 ±7.8% DOCA; 51 ± 7.8% sham). Thus, although these receptors are present in the SMA, their activation does not play a role in relaxation, and are not involved in a 5‐HT‐induced fall in BP.
Functional changes of vascular responses in familial hemiplegic migraine type 1
Authors: Soydan Guray , Ferrari Michel D. , van den Maagdenberg Arn M. J. M. , Ayata Cenk
Published: June 2021

The R192Q mutation of the CACNA1A gene, encoding for the α1 subunit of voltage‐gated P/Q‐type Ca2+ channels (Ca(v)2.1), is associated with familial hemiplegic migraine‐1. We aimed to investigate functional responses in different arteries of R192Q mutant mice. Since the phenotype is stronger in female R192Q mutants we isolated basilar, carotid artery and aorta of female mice. Using a wire myograph vasomotor responses to KCl, endothelin‐1 (ET‐1) and U46619 (a tromboxane A2 agonist) in basilar artery were evaluated. In aorta and carotid artery, vasoconstrictions to KCl, serotonin (5‐HT) and phenylephrine (PE) and vasodilatations to acetylcholine (Ach) and sodium nitroprusside (SNP) were measured. U46619‐induced constrictions slightly but not significantly increased in basilar artery of R192Q mice, but no changes were measured to KCl and ET‐1 induced constrictions. PE‐induced vasoconstriction decreased in both aorta and carotid artery of mutant mice. Vasomotor responses to 5‐HT decreased in carotid artery of R192Q knockin mice but it did not differ in aorta. No difference between groups to Ach and SNP vasodilations were observed. Our results indicate a change in vascular responsiveness to vasoconstrictors in carotid and aorta but not in basilar artery of R192Q mutant mice. Since basilar artery of mice does not have PE and 5‐HT constrictions, molecular studies are needed to test the changes in cerebral arteries.
Rosiglitazone prevents insulin deficiecy induced hypertension in rats
Authors: El‐Bassossy Hany , Fahmy Ahmed , Abo‐Warda Shaimaa
Published: June 2021

The macrovascular complications of diabetes represent the leading cause of morbidity and mortality in affected subjects. Rosiglitazone has been widely used in treatment of insulin resistance. Here we investigated the effect of rosiglitazone on insulin deficiency induced hypertension.

Insulin deficiency was induced by streptozotocin (STZ, 50 mg/kg). Rosiglitazone (10 mg/kg) was daily administered two weeks after STZ injection and for 6 weeks. Insulin deficient group were left untreated for 8 weeks after STZ injection. At the end of study, non invasive blood pressure (BP) was recorded. Rings of isolated thoracic aorta were used to measure the responses to receptor (phenylephrine, PE) and depolarization (KCl) vasoconstrictors, and the endothelial dependent (ACh) and independent (sodium nitroprusside, SNP) relaxants.

Insulin deficiency was associated with elevated systolic and diastolic BP while, rosiglitazone treatment prevented this elevation in BP. Insulin deficiency increased vasocontractive response of aorta to PE and KCl, decreased vasorelaxatant response to ACh without affecting response to SNP, while rosiglitazone treatment prevented the impairment in response to KCl, ACh but not to PE. In conclusion, rosiglitazone protects from the insulin deficiency induced elevation in BP through a mechanism involving prevention of the impairment in aortic vascular reactivity.
Characterization of the Cardiovascular Profile of Two Novel Ca2+ Channels Blockers
Authors: Milicic Ivan , Vortherms Timothy A. , Shieh Char‐Chang , Simler Gricelda , Banfor Patricia , King Andrew , Stewart Andrew O. , Mack Helmut , Bhatia Pramila , Doherty George , Jarvis Michael F. , Scott Victoria E.
Published: June 2021

Voltage‐gated Ca2+ channels modulate neurotransmitter release and cellular excitability in neurons. While the N‐Type selective Ca2+ channel pore blocker Prialt® attenuates chronic pain in patients, its clinical utility is limited by intrathecal delivery. Identification of state‐dependent small molecule Ca2+ channels blockers has the potential to have similar analgesic efficacy to Prialt® with a superior cardiovascular safety profile. We have identified two novel compounds, A‐686085 and A‐1048400, that potently block N‐ (IC50 = 0.84 and 0.8 μM, respectively) and T‐Type (IC50 = 4.6 and 0.3 μM, respectively) Ca2+ channels. Functional selectivity versus L‐Type Ca2+ channels was determined by measuring relaxation of rat aorta rings (EC50 of 0.37μM for A‐686085 and 14.4 μM for A‐1048400). Both blockers reversed mechanical nociception in the capsaicin model of secondary hyperalgesia with similar potency (EC50 = 0.5 – 0.8 μg/ml). In an anesthetized rat cardiovascular assay, A‐686085 produced large dose‐dependent decreases in mean arterial pressure (13%, 37%, 42%) with no effect on heart rate at plasma levels of 0.1, 0.3 and 0.7 μg/ml, respectively. In contrast, A‐1048400 produced only modest effects on mean arterial pressure (4%) and no effects on heart rate up to 4.9 μg/ml. These data show that A‐1048400 has a superior therapeutic index relative to A‐686085 and demonstrates that analgesic calcium channel blockers can be identified that have reduced hemodynamic liabilities. Research supported by Abbott Laboratories.
PTH (1–84)‐induced augmented bone metabolism and endothelium‐dependent vasodilation of the bone vasculature are coupled via VEGF‐related mechanisms in rats
Authors: Campbell Jeremiah , Menezes Thomas , Prisby Rhonda
Published: June 2021

Circulation is requisite for bone growth and repair. Parathyroid hormone (PTH) augmented bone but not angiogenesis. Since PTH enhances bone blood flow, perhaps bone accrual is related to enhanced vasodilation. The principal nutrient artery (PNA) is the primary conduit for flow to long bones. This study assessed 1) whether PTH improves bone vascular function 2) via mechanisms related to VEGF. Male Wistar rats (4 mon) were injected (5 d/wk) for 2 wk accordingly: 1) CON (vehicle), 2) PTH (100 μg/kg/d), 3) PTH + anti‐VEGF antibody (0.7 mg/kg/d) (PAV), and 4) anti‐VEGF Ab (AV). Endothelium‐dependent (ACh: 10−9 – 10−4 M) and ‐independent (SNP: 10−10 – 10−4 M) vasodilation was assessed in femoral PNAs (n=6/group). Femora were analyzed by bone histomorphometry. PTH increased bone volume (CON; 12 ± 2%, *PTH; 23 ± 2%, PAV; 10 ± 2%, and AV; 11 ± 1%) and OS/BS by 88%. Dynamic properties revealed enhanced double labeled surfaces and mineralizing surfaces and therefore, overall bone formation rate (μm3/μm2/day); CON; 0.08 ± 0.05, *PTH; 1.02 ± 0.5, PAV; 0.08 ± 0.5, and AV; 0.08 ± 0.04). PTH enhanced maximal endothelium‐dependent vasodilation which was blunted by anti‐VEGF (CON; 67 ± 5%, *PTH; 86 ± 5%, PAV; 59 ± 8%, and AV; 67 ± 7%). No change in smooth muscle cell responsiveness was observed. These results suggest that PTH‐induced bone metabolism and enhanced vascular function are coupled to VEGF and presumably related to enhanced bone perfusion.
Inter‐individual differences in coupling oxygen delivery to exercising muscle metabolic demand
Authors: Walsh Jeremy James , Moynes Jackie , Kellawan Jeremy Mikhail , Bentley Robert , Bravo Michael , Shantz Rebecca , Tschakovsky Michael E
Published: June 2021

That oxygen delivery (O2del) is “tightly matched” to metabolic demand is widely accepted based on traditional data analysis using group means. Little consideration has been given to the investigation of inter‐individual differences in this coupling.

PURPOSE

To determine the degree of inter‐individual differences in O2del:metabolic demand coupling in healthy young males.

METHODS

Eleven recreationally active (271.38 ± 6.74 METS) healthy males (24.18 ± 2.08 yrs) performed rhythmic isometric handgrip contractions at a fixed metabolic demand (1 s: 2 s contraction: relaxation duty cycle, 30 kg force) until failure. Forearm O2del was calculated as arterial oxygen content (O2 saturation via pulse oximeter; [hemoglobin] via venous blood sample) x brachial artery blood flow (BABF; Doppler and Echo ultrasound). Data are mean ±SD.

RESULTS

BABF in last 30 s of exercise: 478 ±150 ml/min, Coefficient of variation (CV) 31%, individual subject values 249, 589, 465, 317, 475, 412, 391, 420, 700, 739, 499 ml/min. O2del last 30 s of exercise: 98 ±30 ml/min, CV 31%, individual subject values 55, 117, 92, 69, 88, 85, 75, 88, 141, 152, 115 ml/min.

CONCLUSIONS

“Tightness” of O2del:metabolic demand coupling varies substantially between individuals. Investigating inter‐individual variation may provide an important new approach to advancing understanding of integrated cardiovascular response support of exercising muscle. NSERC
Hemodynamic changes are encoded into distinct astrocyte and neuronal responses highlighting the importance of bidirectional communication at the neurovascular unit
Authors: Kim Ki Jung , Filosa Jessica A
Published: June 2021

To gain insights into the dynamic signaling events at the neurovascular unit, we evaluated if hemodynamic changes within parenchymal arterioles altered glial and/or neuronal activity in cortical brain slices. We hypothesize that astrocytes monitor brain hemodynamics and that changes in pressure and/or flow are rapidly sensed by these cells and tranduced to neighboring neurons adjusting, in this way, activity to blood flow supply. We used a novel in vitro approach which included pressure/flow changes in parenchymal arterioles while simultaneously measuring Ca2+ activity in perivascular astrocytes and firing activity from pyramidal or cortical interneurons. Our findings show that astrocytes rapidly sense increases in pressure (from 40 to 80 mmHg) resulting in a 124% increased in Ca2+ oscillation frequency. Moreover, an 80% reduction in neuronal firing activity was observed in the presence of the hymodynamic stimulus. A detailed characterization of astrocytic responses revealed the presence of multiple response patterns to the hemodynamic stimulus including: transient activation (21%), tonic/oscillatory activity (26%) and inhibition (52%). The present study support a complex form of vascular‐to‐glial‐to‐neuronal communication in which hemodynamic changes are encoded into distinct astrocyte/neuronal responses.
Effect of combined inhibition of neuronal nitric oxide synthase, cyclooxygenase‐2, and metabotropic glutamate receptors on the cerebral blood flow response to neuronal activation of whisker barrel cortex
Authors: Liu Xiaoguang , Koehler Raymond C
Published: June 2021

The increase in cerebral blood flow (CBF) during neuronal activation can be only partially attenuated by individual administration of inhibitors of neuronal NO synthase with 7‐nitroindazole, cyclooxgenase‐2 with NS‐398, or metabotropic glutamate receptor subtype 1 with LY‐367385 and subtype 5 with MPEP (which block the increase in astrocyte calcium). Here, we determined if combining these inhibitors completely blocks the CBF response to whisker stimulation in anesthetized rats. With combined inhibition, the CBF response was reduced from 20±2% to 8±1% (±SD). We postulated that the inability to completely block the CBF response was due to tissue acidosis resulting from impaired clearance of metabolically produced CO2. To test this idea, the bicarbonate of the CSF superfusate was increased from 24 to 60 mM for 1 h, which was found to markedly reduce the CBF response to hypercapnia. However, increasing the CSF bicarbonate had no effect on the CBF response to whisker stimulation after combined inhibition (8±1% to 7±2%). We conclude that the residual response after inhibition of known astrocyte and neuronal‐based vasodilatory mechanisms is not due to acidosis arising from impaired CO2 clearance when the CBF response is reduced. Perhaps an unidentified redox sensing mechanism or neurogenic mechanism is responsible for the rapid (2–5 s) residual vasodilation during neuronal activation. (Supported by NIH HL59996.)
Inter‐individual oxygen delivery differences strongly influence estimated critical power in an all out exercise test
Authors: Kellawan Jeremy Mikhail , Bravo Michael F , Moynes Jackie S , Bentley Robert , Shantz Rebecca , Tschakovsky Michael E
Published: June 2021

PURPOSE

To determine if differences in O2 delivery (O2D) between healthy subjects predicts performance in a critical power (CP) test.

METHODS

10 healthy male subjects lay supine with the arm at heart level and completed 10 min of rhythmic maximal voluntary contractions (1 s contraction: 2 s relaxation duty cycle). CP was calculated as the average peak contraction force during the last 30 s of the trial where force plateaued. Forearm brachial artery blood flow (FBF) during exercise (echo and Doppler ultrasound) and O2D (FBF x arterial oxygen content ([haemoglobin] from venous blood sample and oxygen saturation from pulse oximetry)) were measured.

RESULTS

means ± SD. A wide range of O2D and CP was observed. O2D 101.5 ± 25.1 ml O2/min (range 70.92–154.1 ml O2/min) & CP 25.4 ± 6.0 kg (range 17.5–34.7 kg). O2D & FBF area under the curve (AUC, ml O2/forearm volume & ml/forearm volume) for the first 200 s of exercise were predictors of CP (r2 = 0.47, 0.48 p ≤ 0.03). However, O2D & FBF total AUC during exercise was the strongest predictors rather than steady state levels achieved (r2 = 0.63, 0.66 p ≤ 0.01).

CONCLUSIONS

Considerable inter‐individual differences in oxygen delivery exist during an all out CP test. These differences account for much of the inter‐individual differences in CP, supporting the contention that vasodilatory responsiveness and capacity are important determinants of small muscle mass CP. NSERC
Integrated control of coronary blood flow in exercising swine by adenosine, nitric oxide and KATP channels
Authors: Duncker Dirk J , de Beer Vincent J , Taveerne Yannick , de Graaff Han , Merkus Daphne
Published: June 2021

The mechanism of exercise‐induced coronary vasodilation remains incompletely understood. KATP channels in conjunction with nitric oxide (NO) and adenosine have been proposed to mediate exercise hyperemia in dogs but not in swine. Different results may have been due, in part, to different routes of administration of drugs to block these pathways. Hence, we studied the role of KATP channels together with NO and adenosine in exercise hyperemia in swine, using an identical drug dosing protocol and administration route that was previously shown to virtually abolish the exercise hyperemia in dogs. Swine exercised on a treadmill at 0–5 km/h, before and after blockade of KATP channels (glibenclamide, 50 μg/kg/min intracoronary (ic)), in the absence or presence of blockade of adenosine receptors (8‐phenyltheophylline, 8PT, 5 mg/kg iv) and NO‐synthase (nitro‐L‐arginine, NLA, 1.5 mg/kg, ic). Glibenclamide reduced myocardial O2 delivery and coronary venous O2 tension at rest and during exercise. Administration of 8PT + NLA reduced coronary venous O2 tension but did not enhance vasoconstriction to glibenclamide. Despite an increase in O2‐extraction to ~90% during combined NLA + 8PT + glibenclamide, exercise‐induced hyperemia was maintained. Thus, KATP channels, adenosine and NO do not appear to be critical for exercise coronary hyperemia, and act in a linear additive fashion, in swine.

Supported by NHF Grant 2000T042
Role of ryanodine receptors in acidic pH‐induced dilation of brain parenchymal arterioles
Authors: Dabertrand Fabrice , Nelson Mark T. , Brayden Joseph E.
Published: June 2021

Ryanodine receptors (RyRs) are Ca2+ permeable channels in the sarcoplasmic reticulum. RyRs regulate vascular smooth muscle cell (VSMC) membrane potential through their localized Ca2+‐release events, termed Ca2+ sparks, which activate hyperpolarizing BK currents leading to vasodilation. This mechanism is clearly demonstrated by the non‐additive constrictions induced by RyR or BK channel blockers in pressurized pial arteries. However these blockers have little effect on the diameter of pressurized parenchymal arterioles (PAs) from the brain, even though functional BK channels and RyRs are present. At normal pH (7.4), VSMCs in pressurized PAs exhibit largely Ca2+ waves but not Ca2+ sparks. Alkaline pH has been shown to shift Ca2+ sparks to Ca2+ waves in pial arteries. Therefore, we hypothesized that acidic pH might dilate PAs by reshaping the intracellular Ca2+ dynamic from Ca2+ waves to Ca2+ sparks, thereby activating BK channels. As predicted, reducing extracellular pH from 7.4 to 7.0 dramatically decreased Ca2+ wave activity, and increased spark activity. Acidic pH caused a dilation of up to 70% of the maximal diameter, and this dilation was inhibited by about 60% by BK or RyR blockers. These findings suggest that RyRs play a major role in the acidic pH‐induced dilation of PAs through Ca2+ spark‐mediated activation of BK channels. Supported by AHA 09POST2290090 (FD) and the NIH HL095488, HL44455 and HL58231.
Pathways mediating Ca2+ senzitization in basilar artery of the rat: feature and mechanisms
Authors: Rutkai Ibolya , Czikora Agnes , Darago Andrea , Megyesi Zita , Edes Istvan , Papp Zoltan , Toth Attila
Published: June 2021

We characterized the Ca2+ sensitivity of isometric force production in arteria basilaris of the rat. Contractile force (in mN) and intracellular Ca2+ concentrations ([Ca2+]i, Fura‐2 340/380 ratio) were measured simultaneously in the presence of agonists and inhibitors of PKC and Rho kinase. KCl evoked 2.5±0.2 mN contraction with 0.17±0.01 increase in [Ca2+]i (n=97). The Ca2+ sensitivity of force production was 18.5 mN/340/380 ratio. Serotonin evoked 6.7±0.5 mN contraction with similar changes 0.15±0.02 in [Ca2+]i, resulting in dose‐dependent increase in Ca2+ sensitivity of force production (from 19 to 38.7 mN/340/380, EC50=32nM) (n=19). U‐46619, a thromboxane A2 receptor (TxA2) agonist evoked 4.2±0.5 mN contraction and 0.04±0.01 increase in [Ca2+]i. The Ca2+ sensitivity of force production increased from 20 to 97.9 mN/340/380 with an EC50 of 7,9 nM (n=23). The Ca2+ sensitivity of force production was not affected by PKC but decreased upon Rho‐kinase inhibition.

Rho kinase mediated increase in Ca2+ sensitivity of force production significantly contributed to vascular contraction in both serotonin and TxA2 receptor stimulation. Funding: TÁMOP‐4.2.1.B‐09/1/KONV‐2010‐0007
Arachidonic acid (AA) and Prostaglandin E2‐induced cerebral vasodilation is mediated by Carbon Monoxide independent of Reactive Oxygen Species in the newborn pig
Authors: Kanu Alie Joseph , Leffler Charles W.
Published: June 2021

Arachidonic acid (AA) and prostaglandin (PG) E2 stimulate carbon monoxide (CO) production, and AA metabolism is known to be associated with generation of reactive oxygen species (ROS). Therefore, AA and PGE2 generated in cerebral microvessels might increase cerebrovascular dilation through generation of CO or ROS. In this study we tested this hypothesis by examining the effect of inhibitors of CO and ROS production on the cerebral vasodilation produced by AA and PGE2 in newborn pigs. Experiments were performed in anesthetized newborn pigs with closed cranial windows. In experiments with PGE2, indomethacin (5mg/kg i.v.) was administered to inhibit endogenous prostanoid production. AA (10‐7 M, 10‐5 M) and PGE2 (10‐7M, 10‐5M) produced cerebral vasodilation that was inhibited by heme oxygenase (HO) inhibitor chromium mesoporphyrin (CrMP) (2×10‐5 M) (P<0.05), but not by ROS inhibitor tempol (SOD mimetic)(1×10‐5 M) or the H2O2 scavenger, catalase (100U/ml). Both tempol and catalase minimized the cerebral vasodilation produced by 0.2mM U/ml xanthine and 0.2mM xanthine oxidase. These data suggest that CO independent of ROS is involved in the cerebral vasodilation produced by AA and PGE2.
Effect of circulating factors on cerebral artery function during hyperglycemic stroke
Authors: Cipolla Marilyn J. , Sweet Julie G. , Gardner‐Morse Ira
Published: June 2021

We investigated the influence of circulating factors on vascular function of middle cerebral arteries (MCA) as a potential mechanism controlling perfusion and stroke outcome during hyperglycemic stroke. Nonischemic MCA were isolated from male Wistar rats, cannulated and perfused with plasma (20% in buffer) from rats that were hyperglycemic by STZ (50mg/kg) for 5 days and that underwent either MCA occlusion (MCAO) for 2 hrs with 2 hrs reperfusion (MCAO, n=8) or a 4 hour sham procedure (Sham, n=8). Control vessels were perfused with buffer without plasma (CTL, n=8). Myogenic responses and constriction to NOS inhibition by a single concentration (0.1mM) of L‐NNA were compared. Arteries perfused with Sham and MCAO plasma had significantly increased myogenic tone vs. CTL (at 75mmHg: 30±3 and 22±4% vs. 10 ±3%;p<0.01 vs. CTL). Sham and MCAO plasma vessels also had smaller lumen diameters vs. CTL (at 75mmHg: 210±14μm and 193±9μm vs. 243±9μm; p<0.05). Constriction to L‐NNA was decreased only in the MCAO plasma group. The percent constriction to L‐NNA for CTL, Sham and MCAO plasma was: 41±5%, 31±5% (ns) and 4±1% (p<0.01 vs. both). These results suggest that circulating factors in plasma during postischemic reperfusion increase tone in MCA possibly through an effect on NO. Further experiments are needed to determine what circulating factors are in both types of plasma that affect tone. Supported by RO1NS043316 and PO1HL9548801.
Effect of Warm‐Up Exercise on Myocardial Perfusion Studied with Wave Intensity Analysis
Authors: Rolandi Cristina , Lockie Tim , Guilcher Antoine , Perera Divaka , Redwood Simon , Marber Michael , Siebes Maria
Published: June 2021

Background

The effect of warm‐up on myocardial perfusion in patients with coronary artery disease (CAD) is not well understood. We compared outcomes of repeat exercise periods using wave intensity (WI) analysis. Coronary WI reflects the effects of cardiac contraction and relaxation on coronary blood flow dynamics and distinguishes between left ventricular (forward, FW) and microcirculatory (backward, BW) originating contributions.

Methods

During cardiac catheterization in 10 patients with CAD, we simultaneously recorded aortic pressure, distal coronary pressure (Pd) and flow velocity (U), while the patients performed 2 serial periods of supine bicycle exercise (Ex1, Ex2). WI was calculated as (dPd/dt)(dU/dt) and the separate BW and FW contributions were derived.

Results

Patients reached similar exercise levels in Ex1 and Ex2, but time to ischemia was longer during Ex2. At the same cardiac workload, Pd was unchanged in Ex2 indicating absence of vasoconstriction. Concomitantly, U increased by 10% (p<0.03) and microvascular resistance (Pd/U) decreased by 20% during Ex2 (p<0.01). In contrast to FW, BW wave energy was more than 30% higher in Ex2 (p<0.05). The energy of flow accelerating waves was 20% higher in Ex2, with no difference in the flow decelerating waves.

Conclusion

Our results suggest a warm‐up effect as evidenced by indices of improved myocardial perfusion at repeat exercise.
Function of the cAMP dependent transcription factors CREM and ATF1 in the vasculature
Authors: Seidl Matthias Dodo , Steingräber Annika Kathrin , Sur Huyen , Schmitz Wilhelm , Müller Frank Ulrich
Published: June 2021

Transcription factors of the CREB/CREM/ATF1‐family (cAMP responsive‐element (CRE) binding protein and modulator and activating transcription factor 1) play an important role in the cAMP mediated gene control. To address the vascular role of CREM and ATF1 we studied mice with a global inactivation of CREM (CKO) or ATF1 (AKO).

CKO mice showed an increase in neointima formation after ligation of the carotid artery associated with an elevated proliferation rate of VSMCs in carotid media. Atherogenic high‐fat‐diet revealed a 20% increase of plaque area in CKOxApoE‐KO vs. control mice. In primary VSMC proliferation rate was increased 1.3 fold in CKO vs. controls after platelet derived growth factor (PDGF) stimulation associated with a 3 fold increased CRE‐mediated transcriptional activity.

In AKO VSMCs PDGF induced proliferation was blunted compared to controls. AKO VSMCs also revealed a 1.9‐fold higher proportion of apoptotic cells after H2O2 treatment. These results were not associated with differences in neointima‐ or plaque formation.

We conclude that ATF1 is in principle involved in the regulation of vascular proliferation and apoptosis but may play an inferior role under physiological conditions. Anti‐proliferative effects suggest CREM as an important factor for the pathogenesis of vascular proliferative disorders. (Supported by the IZKF Münster).
Truncated β2 subunits increase membrane expression of LTCC subunits in A7r5 cells
Authors: Telemaque Sabine , Flynn Shawn E , Liu Meeiyueh , Grain Terrie , Marsh James D
Published: June 2021

Cytosolic Ca2+ is regulated by Ca2+ entry through L‐type calcium channels (LTCC) and release from intracellular Ca2+ stores via the IP3 receptors (IP3R) in vascular smooth muscle cells (VSMC). The β2 subunit of the LTCC is an intracellular chaperone that interacts with the α1C subunit via its beta interaction domain (BID) to regulate membrane localization and channel function. In order to alter assembly of the LTCC at the cell surface of VSMC, we used adenovirus constructs to overexpress either the β2 subunit (Full‐β2) or truncated β2 subunits lacking either the C‐, N‐terminus or both (N‐BID, C‐BID or BID, respectively) fused to GFP. Immunoblot analysis confirmed that Full‐β2 was primarily expressed in the membrane fraction while C‐BID and N‐BID expression at the membrane was reduced to 70% and 52%, respectively. BID was in the cytosol, like GFP. Full‐β2 significantly increased α1C subunit expression at the membrane, as did C‐BID and N‐BID. Membrane levels of α2 subunit were also increased by the 3 constructs. BID did not modify localization of the LTCC subunits. Membrane levels of IP3R were significantly increased only by Full‐β2. Our study showed that truncated β2 subunits can alter expression of the LTCC subunits at the cell surface and suggest that LTCC function could be regulated by these decoys. Development of VSMC‐specific constructs may represent a viable gene‐based treatment to regulate vascular reactivity.
Pregnancy increases mesenteric but not uterine artery response to thromboxane via activation of ERK pathway
Authors: Goulopoulou Styliani , Hannan Johanna L , Webb R Clinton
Published: June 2021

Pregnancy is associated with a reduction in systemic vascular resistance, resulting in maintenance, or a small decrease, in arterial pressure and a vast increase in uteroplacental blood flow. We examined the effect of pregnancy on arterial contractile response to thromboxane (TXA2). We hypothesized that UA and mesenteric (ME) arteries of pregnant rats will have reduced contractile response to TXA2. Segments of UA and 2nd order ME of late pregnant (20 d, LP) and virgin (V) Sprague Dawley rats (16 wks old) were mounted and concentration response curves to a TXA2 analogue (U46619, +/− ERK inhibitor, PD98059; RhoK inhibitor, Y27632; PKC inhibitor, BIM) were performed. ME but not UA from LP exhibited reduced contractile response to TXA2. In the presence of ERK inhibitor, UA response to TXA2 was reduced only in V, whereas in ME, TXA2‐induced contraction was reduced in both groups and this reduction was greater in LP (24% vs. 58%). Incubation with RhoK and PKC inhibitors had no effect on TXA2‐induced UA contraction in either group but diminished ME response to TXA2 only in V. TXA2‐induced UA contraction is primarily mediated via an ERK signaling pathway, whereas ME response to TXA2 is regulated by ERK, RhoK, and PKC signaling pathways in the non‐pregnant state. Pregnancy increases ME but not UA response to TXA2 via activation of ERK pathway. (Funded by NIH and Heart & Stroke Foundation Canada)
The effects of chronic estrogen supplementation on neuropeptide Y neurotransmission in skeletal muscle arterioles
Authors: Evanson Kirk W , Stone Audrey J , Kluess Heidi A
Published: June 2021

The purpose of this study was to determine the effects of chronic estrogen supplementation on NPY neurotransmission in gastrocnemius first‐order arterioles (G1A) of adult female rats. Female rats (4 mo; n = 30) were ovariectomized (OVX) with a subset (n = 15) receiving an estrogen pellet (OVE; 17β‐estradiol (4μg/day)). Red G1A were excised and cannulated with micropipettes connected to albumin reservoirs. NPY‐mediated vasoconstriction via a Y1‐agonist decreased vessel diameter 59.74 ± 3.80% as compared to baseline; however, there were no group differences in EC50 (OVE: −8.97 ± 0.36; OVX: −8.72 ± 0.20 log M [Leu31Pro34]NPY) or slope (OVE: −1.37 ± 0.38; OVX: −1.64 ± 0.31 % baseline/log M [Leu31Pro34]NPY). NPY overflow experienced a slight increase following field stimulation, and significantly increased (p < 0.05) over control conditions in the presence of a DPPIV inhibitor (diprotin A). These data suggest that NPY can induce an estrogen‐independent decrease in vessel diameter in G1A, and DPPIV is active in mitigating NPY overflow in adult female rats. This research was supported by the Arkansas Biosciences Institute and the NIA (grant no. 5R03AG033245).
Blockade of Angiotensin Receptor Ameliorates Insulin Resistance and Suppresses Plasma Leptin in OLETF Rats Supplemented with High Glucose
Authors: Aguil Jennifer Denyse , Rodriguez Ruben , Viscarra Jose , Minas Jacqueline , Nakano Daisuke , Nishiyama Akira , Ortiz Rudy
Published: June 2021

Renin angiotensin system blockade improves insulin resistance (IR); however, the effects of angiotensin receptor blockers (ARB) on plasma leptin in a model of IR are not well defined. We examined the benefits of ARB on IR and leptin in 5 groups of rats: 1) LETO (lean strain control), 2) OLETF, 3) OLETF + 5% glucose water (OLETF HG), 4) OLETF + ARB (10mg/kg/day) and 5) OLETF HG + ARB. Homeostasis Model Assessment of Insulin Resistance (HOMA‐IR) was calculated as an indicator of IR after 6 weeks of treatment. HOMA‐IR increased 3‐fold in OLETF compared to LETO. High glucose increased mean HOMA‐IR 50%, and ARB treatment alleviated IR in OLETF ARB and OLETF HG + ARB. Plasma leptin levels increased over 2‐fold in OLETF (10.7 ± 0.9 ng/mL) compared to LETO (4.3 ± 0.6 ng/mL). ARB (9.7 ± 0.7 ng/mL) was effective in normalizing the increase in plasma leptin associated with HG (21.1±2.5 ng/mL). Plasma leptin positively correlated with IR (HOMA‐IR = 0.71 + 0.94*plasma leptin; R = 0.96; p< 0.01) suggesting that increased plasma leptin contributes to the manifestation of IR in obesity. Preliminary data suggest that increased plasma leptin is not associated with increased phosphorylation (and thus activation) of the leptin receptor in the liver. Furthermore, the data suggest that AT1 activation in conjunction with elevated leptin contributes to the development of IR in a model of metabolic syndrome. Funded by NIH NCMHD 9T37MD001480.
Anoxia‐mediated elevation in phasic GABAA receptor currents is essential for anoxia‐tolerance in turtle cortex
Authors: Hogg David William , Buck Leslie Thomas
Published: June 2021

In mammalian brain, anoxia induces hyper‐excitability and cell death; however, the western painted turtle Chrysemys picta bellii is anoxia tolerant, neuronal activity is depressed by anoxia and cell death does not occur. In anoxic turtle brain [γ‐aminobutyric acid] (GABA) is elevated suggesting that the mechanism(s) responsible for anoxia‐tolerance involve GABAergic synaptic transmission. The objective of this study was to investigate the neuroprotective role of endogenous phasic GABAA receptor currents during anoxia in turtle cortex. Using whole‐cell patch clamp techniques we identified phasic GABA‐mediated IPSC's that double in amplitude with anoxia (41.2 ± 1.6 to 82.1 ± 2.1 pA) resulting in a 60–70% decrease in spontaneous action potential frequency. Inhibition of phasic GABAA receptor currents with gabazine resulted in hyper‐excitability. We conclude that anoxia increases phasic GABAA receptor peak currents resulting in a “shunting current” that prevents further depolarization and hyper‐excitability. Research supported by OGSST grant to DH and NSERC grant to LB.