scholarly journals Knockdown of tyrosine hydroxylase in the nucleus of the solitary tract reduces elevated blood pressure during chronic intermittent hypoxia

2013 ◽  
Vol 305 (9) ◽  
pp. R1031-R1039 ◽  
Author(s):  
Chandra Sekhar Bathina ◽  
Anuradha Rajulapati ◽  
Michelle Franzke ◽  
Kenta Yamamoto ◽  
J. Thomas Cunningham ◽  
...  
Keyword(s):  
Tyrosine Hydroxylase ◽  
Transcriptional Activation ◽  
Intermittent Hypoxia ◽  
Fluorescent Protein ◽  
Nucleus Tractus Solitarius ◽  
Cardiovascular Responses ◽  
Ventrolateral Medulla ◽  
Chronic Intermittent Hypoxia ◽  
Sprague Dawley ◽  
Green Fluorescent

Noradrenergic A2 neurons in nucleus tractus solitarius (NTS) respond to stressors such as hypoxia. We hypothesize that tyrosine hydroxylase (TH) knockdown in NTS reduces cardiovascular responses to chronic intermittent hypoxia (CIH), a model of the arterial hypoxemia observed during sleep apnea in humans. Adult male Sprague-Dawley rats were implanted with radiotelemetry transmitters and adeno-associated viral constructs with green fluorescent protein (GFP) reporter having either short hairpin RNA (shRNA) for TH or scrambled virus (scRNA) were injected into caudal NTS. Virus-injected rats were exposed to 7 days of CIH (alternating periods of 10% O2 and of 21% O2 from 8 AM to 4 PM; from 4 PM to 8 AM rats were exposed to 21% O2). CIH increased mean arterial pressure (MAP) and heart rate (HR) during the day in both the scRNA ( n = 14, P < 0.001 MAP and HR) and shRNA ( n = 13, P < 0.001 MAP and HR) groups. During the night, MAP and HR remained elevated in the scRNA rats ( P < 0.001 MAP and HR) but not in the shRNA group. TH immunoreactivity and protein were reduced in the shRNA group. FosB/ΔFosB immunoreactivity was decreased in paraventricular nucleus (PVN) of shRNA group ( P < 0.001). However, the shRNA group did not show any change in the FosB/ΔFosB immunoreactivity in the rostral ventrolateral medulla. Exposure to CIH increased MAP which persisted beyond the period of exposure to CIH. Knockdown of TH in the NTS reduced this CIH-induced persistent increase in MAP and reduced the transcriptional activation of PVN. This indicates that NTS A2 neurons play a role in the cardiovascular responses to CIH.

scholarly journals Chronic intermittent hypoxia increases blood pressure and expression of FosB/ΔFosB in central autonomic regions

2011 ◽  
Vol 301 (1) ◽  
pp. R131-R139 ◽  
Author(s):  
W. David Knight ◽  
Joel T. Little ◽  
Flavia R. Carreno ◽  
Glenn M. Toney ◽  
Steven W. Mifflin ◽  
...  
Keyword(s):  
Intermittent Hypoxia ◽  
Renin Angiotensin System ◽  
Brain Regions ◽  
Ventrolateral Medulla ◽  
Chronic Intermittent Hypoxia ◽  
Preoptic Nucleus ◽  
Sprague Dawley ◽  
Angiotensin System ◽  
Sprague Dawley Rats ◽  
Obstructive Sleep

Chronic intermittent hypoxia (CIH) models repetitive bouts of arterial hypoxemia that occur in humans suffering from obstructive sleep apnea. CIH has been linked to persistent activation of arterial chemoreceptors and the renin-angiotensin system, which have been linked to chronic elevations of sympathetic nerve activity (SNA) and mean arterial pressure (MAP). Because Fos and FosB are transcription factors involved in activator protein (AP)-1 driven central nervous system neuronal adaptations, this study determined if CIH causes increased Fos or FosB staining in brain regions that regulate SNA and autonomic function. Male Sprague Dawley rats were instrumented with telemetry transmitters for continuous recording of MAP and heart rate (HR). Rats were exposed to continuous normoxia (CON) or to CIH for 8 h/day for 7 days. CIH increased MAP by 7–10 mmHg without persistently affecting HR. A separate group of rats was killed 1 day after 7 days of CIH for immunohistochemistry. CIH did not increase Fos staining in any brain region examined. Staining for FosB/ΔFosB was increased in the organum vasculosum of the lamina terminalis (CON: 9 ± 1; CIH: 34 ± 3 cells/section), subfornical organ (CON: 7 ± 2; CIH: 31 ± 3), median preoptic nucleus (CON 15 ± 1; CIH: 38 ± 3), nucleus of the solitary tract (CON: 9 ± 2; CIH: 28 ± 4), A5 (CON: 3 ± 1; CIH: 10 ± 1), and rostral ventrolateral medulla (CON: 5 ± 1; CIH: 17 ± 2). In the paraventricular nucleus, FosB/ΔFosB staining was located mainly in the dorsal and medial parvocellular subnuclei. CIH did not increase FosB/ΔFosB staining in caudal ventrolateral medulla or supraoptic nucleus. These data indicate that CIH induces an increase in FosB/ΔFosB in autonomic nuclei and suggest that AP-1 transcriptional regulation may contribute to stable adaptive changes that support chronically elevated SNA.

scholarly journals Central losartan attenuates increases in arterial pressure and expression of FosB/ΔFosB along the autonomic axis associated with chronic intermittent hypoxia

2013 ◽  
Vol 305 (9) ◽  
pp. R1051-R1058 ◽  
Author(s):  
W. David Knight ◽  
Ashwini Saxena ◽  
Brent Shell ◽  
T. Prashant Nedungadi ◽  
Steven W. Mifflin ◽  
...  
Keyword(s):  
Arterial Pressure ◽  
Transcriptional Activation ◽  
Intermittent Hypoxia ◽  
Renin Angiotensin System ◽  
Ventrolateral Medulla ◽  
Chronic Intermittent Hypoxia ◽  
Dark Phase ◽  
Angiotensin System ◽  
The Brain

Chronic intermittent hypoxia (CIH) increases mean arterial pressure (MAP) and FosB/ΔFosB staining in central autonomic nuclei. To test the role of the brain renin-angiotensin system (RAS) in CIH hypertension, rats were implanted with intracerebroventricular (icv) cannulae delivering losartan (1 μg/h) or vehicle (VEH) via miniosmotic pumps and telemetry devices for arterial pressure recording. A third group was given the same dose of losartan subcutaneously (sc). Two groups of losartan-treated rats served as normoxic controls. Rats were exposed to CIH or normoxia for 7 days and then euthanized for immunohistochemistry. Intracerebroventricular losartan attenuated CIH-induced increases in arterial pressure during CIH exposure (0800-1600 during the light phase) on days 1, 6, and 7 and each day during the normoxic dark phase. FosB/ΔFosB staining in the organum vasculosum of the lamina terminalis (OVLT), median preoptic nucleus (MnPO), paraventricular nucleus of the hypothalamus (PVN), the rostral ventrolateral medulla (RVLM), and the nucleus of the solitary tract (NTS) was decreased in icv losartan-treated rats. Subcutaneous losartan also reduced CIH hypertension during the last 2 days of CIH and produced bradycardia prior to the effect on blood pressure. Following sc losartan, FosB/ΔFosB staining was reduced only in the OVLT, MnPO, PVN, and NTS. These data indicate that the central and peripheral RAS contribute to CIH-induced hypertension and transcriptional activation of autonomic nuclei and that the contribution of the central RAS is greater during the normoxic dark phase of CIH hypertension.

Assessments of the effects of nicotine and ketamine using tyrosine hydroxylase-green fluorescent protein transgenic zebrafish asbiosensors

2013 ◽  
Vol 42 ◽  
pp. 177-185 ◽  
Author(s):  
Miranda F.K. Suen ◽  
W.S. Chan ◽  
Karen W.Y. Hung ◽  
Y.F. Chen ◽  
Z.X. Mo ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Tyrosine Hydroxylase ◽  
Fluorescent Protein ◽  
Transgenic Zebrafish ◽  
Green Fluorescent

scholarly journals Microglia is a Key Player in the Reduction of Stroke Damage Promoted by the New Antithrombotic Agent Ticagrelor

2014 ◽  
Vol 34 (6) ◽  
pp. 979-988 ◽  
Author(s):  
Paolo Gelosa ◽  
Davide Lecca ◽  
Marta Fumagalli ◽  
Dorota Wypych ◽  
Alice Pignieri ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Interleukin 1 ◽  
Artery Occlusion ◽  
Cerebral Injury ◽  
Sprague Dawley ◽  
Antithrombotic Agent ◽  
Fluorescent Reporter ◽  
Monocyte Chemoattractant Protein 1 ◽  
Proinflammatory Mediators ◽  
Green Fluorescent

The ADP-responsive P2Y12 receptor is expressed on both platelets and microglia. Clinical data show that ticagrelor, a direct-acting, reversibly binding P2Y12-receptor antagonist, reduces total cardiovascular events, including stroke. In our present study, we investigated the expression of P2Y12 receptors and the effects of ticagrelor on brain injury in Sprague-Dawley rats subjected to a permanent middle cerebral artery occlusion (MCAo). Rats were treated per os with ticagrelor 3 mg/kg or vehicle at 10 minutes, 22, and 36 hours after MCAo and killed after 48 hours. Immunofluorescence analysis showed an ischemia-related modulation of the P2Y12 receptor, which is constitutively expressed in Iba1+ resting microglia. After MCAo, activated microglia was mainly concentrated around the lesion, with fewer cells present inside the ischemic core. Ticagrelor significantly attenuated the evolution of ischemic damage—evaluated by magnetic resonance imaging (MRI) at 2, 24, and 48 hours after MCAo—, the number of infiltrating cells expressing the microglia/monocyte marker ED-1, the cerebral expression of proinflammatory mediators (interleukin 1 (IL-1), monocyte chemoattractant protein 1 (MCP-1), nitric oxide synthase (iNOS)) and the associated neurologic impairment. In transgenic fluorescent reporter CX3CR1-green fluorescent protein (GFP) mice, 72 hours after MCAo, ticagrelor markedly reduced GFP+ microglia and both early and late infiltrating blood-borne cells. Finally, in primary cultured microglia, ticagrelor fully inhibited ADP-induced Chemotaxis ( P<0.01). Our results show that ticagrelor is protective against ischemia-induced cerebral injury and this effect is mediated, at least partly, by inhibition of P2Y12-mediated microglia activation and Chemotaxis.

Abstract 3880: GRK2 Inhibition Prevents Development Of Cardiac Insulin Resistance And Impaired Glucose Uptake In Post Ischemic Heart Failure

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Michele Ciccarelli ◽  
Giuseppe Rengo ◽  
Kurt Chuprun ◽  
Gaetano Santulli ◽  
Bruno Trimarco ◽  
...  
Keyword(s):  
Heart Failure ◽  
Glucose Uptake ◽  
Fluorescent Protein ◽  
Sprague Dawley ◽  
Myocardial Glucose Uptake ◽  
Akt Phosphorylation ◽  
Neonatal Cardiomyocytes ◽  
Sprague Dawley Rats ◽  
Green Fluorescent ◽  
And Function

The beta adrenergic receptor (βAR) kinase, GRK2, is upregulated and participates to the evolution of heart failure (HF) through downregulation and desensitization of βARs. Recent studies showed that this molecule affects insulin signaling and reduce glucose uptake in hepatocytes and adipocytes. We hypothesized that in HF, GRK2 reduces cardiac performance also through inhibition of cardiac glucose metabolism. In 12 week old Sprague/Dawley rats, we measured cardiac glucose uptake by PET 3 days, 3 and 6 weeks after myocardial infarction (MI). Function and cardiac dimensions were measured by echocardiography. We observed that glucose uptake was reduced in animal post-MI at 3 and 6 weeks respect to healthy animals (3 rd week: 1.3±0.22 vs 2.1±0.3; 6 th week: 1±0.1 vs 2.4±0.2, ml/min/g, p<0.05). No difference was observed in glucose uptake acutely after surgery. Echo showed cardiac dilation and reduced function at 6 weeks (LVD: 9.2± 0.3 vs 7.2± 0.4 mm; EF: 42%±1.1 vs 66%±2.2, p<0.05, Sham vs MI). To inhibit GRK2 in the heart during post-ischemic HF, we delivered the GRK2 inhibitor βARKct by adeno-associated type 6 virus (AAV6) to the left ventricle before induction of the MI. As a control we treated rats with AAV6 encoding for the green fluorescent protein (GFP). Cardiac dilation and function were preserved after 6 weeks post MI in AAV6 βARKct respect to AAV6GFP rats (LVD: 7.73 ±0.25 vs 9.9 ±0.8 mm; EF: 55%±2.25 vs 44%±2, p<0.05). Glucose uptake was better preserved in AAV6βARKct rats after 3 and 6 weeks post MI respect to AAV6GFP group (3rd week: 2.3±0.3 vs 1.2±0.2; 6th week: 1.8±0.2 vs 1.1±0.05, ml/min/g, p<0.05). Since Akt mediates most of the anabolic effects of insulin in cells, we evaluated the effects of GRK2 overexpression by adenovirus (ADGRK2) in neonatal cardiomyocytes (NRVMs) on Akt phosphorylation later on insulin stimulation (ins, 10 – 6 M). As control we induced overexpression of GFP by adenovirus (ADGFP). We observed reduced activation of Akt in presence of GRK2 overexpression as compared to the ADGFP treated cells (1.2±0.2- vs. 3.5±0.4- fold activation over basal, p<0.05). Our data show that post MI, impaired glucose extraction precedes development of HF, and that early GRK2 inhibition prevents impaired myocardial glucose uptake and HF development.

Chronic intermittent hypoxia alters NE reactivity and mechanics of skeletal muscle resistance arteries

2006 ◽  
Vol 100 (4) ◽  
pp. 1117-1123 ◽  
Author(s):  
Shane A. Phillips ◽  
E. B. Olson ◽  
Julian H. Lombard ◽  
Barbara J. Morgan
Keyword(s):  
Skeletal Muscle ◽  
Intermittent Hypoxia ◽  
Chronic Intermittent Hypoxia ◽  
Physiological Salt Solution ◽  
Ang Ii ◽  
Sprague Dawley ◽  
Resistance Arteries ◽  
Sprague Dawley Rats ◽  
Resting Tone ◽  
Wall Distensibility

Although arterial dilator reactivity is severely impaired during exposure of animals to chronic intermittent hypoxia (CIH), few studies have characterized vasoconstrictor responsiveness in resistance arteries of this model of sleep-disordered breathing. Sprague-Dawley rats were exposed to CIH (10% inspired O2 fraction for 1 min at 4-min intervals; 12 h/day) for 14 days. Control rats were housed under normoxic conditions. Diameters of isolated gracilis muscle resistance arteries (GA; 120–150 μm) were measured by television microscopy before and during exposure to norepinephrine (NE) and angiotensin II (ANG II) and at various intraluminal pressures between 20 and 140 mmHg in normal and Ca2+-free physiological salt solution. There was no difference in the ability of GA to constrict in response to ANG II ( P = 0.42; not significant; 10−10–10−7 M). However, resting tone, myogenic activation, and vasoconstrictor responses to NE ( P < 0.001; 10−9–10−6 M) were reduced in CIH vs. controls. Treatment of rats with the superoxide scavenger 4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (tempol; 1 mM) in the drinking water restored myogenic responses and NE-induced constrictions of CIH rats, suggesting that elevated superoxide production during exposure to CIH attenuates vasoconstrictor responsiveness to NE and myogenic activation in skeletal muscle resistance arteries. CIH also leads to an increased stiffness and reduced vessel wall distensibility that were not correctable with oral tempol treatment.

scholarly journals Molecular Mechanism of the Inhibition of Estradiol-Induced Endometrial Epithelial Cell Proliferation by Clomiphene Citrate

Endocrinology ◽  
2010 ◽  
Vol 151 (1) ◽  
pp. 394-405 ◽  
Author(s):  
Mitsuyoshi Amita ◽  
Toshifumi Takahashi ◽  
Seiji Tsutsumi ◽  
Tsuyoshi Ohta ◽  
Keiko Takata ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Epithelial Cell ◽  
Transcriptional Activation ◽  
Molecular Mechanisms ◽  
Fluorescent Protein ◽  
Clomiphene Citrate ◽  
Immunofluorescence Assay ◽  
Epithelial Cell Proliferation ◽  
Endometrial Epithelial Cell ◽  
Green Fluorescent

Abstract We examined the molecular mechanisms of the antiestrogenic effects of clomiphene citrate (CC) in the endometrium using two types of cell lines, Ishikawa and EM-E6/E7/hTERT cells. CC or ICI182780 inhibited 17β-estradiol (E2)-induced endometrial cell proliferation and transcriptional activation of the estrogen response element (ERE) gene. We directly visualized the ligand-estrogen receptor (ER)α interaction using green fluorescent protein (GFP)-tagged ERα in a single living cell. Whereas E2 changed the nuclear localization of GFP-ERα to a punctate distribution within 5 min, CC or ICI182780 changed the slower and less mobilization of GFP-ERα compared with E2. Pretreatment with CC or ICI182780 partly prevented the E2-induced nuclear redistribution of GFP-ERα. Fluorescence recovery after photobleaching revealed that GFP-ERα mobility treated with E2 was more rapid than that treated by CC or ICI182780. As coactivator recruitment to the ER is essential for ER-dependent transcription, we examined the interaction between ERα and steroid receptor coactivator-1 (SRC-1). The complex formation between ERα and SRC-1 was significantly increased by E2 but was prevented in the presence of CC or ICI182780 by coimmunoprecipitation. Moreover, the E2-induced colocalization of GFP-ERα and SRC-1 was prevented in the presence of CC or ICI182780 according to an immunofluorescence assay. We also observed that the reduction of SRC-1 using small interfering RNA for SRC-1 resulted in the inhibition of E2-induced cell proliferation and transcriptional activation of the ERE gene. Collectively, these results suggest that CC may inhibit E2-induced endometrial epithelial cell proliferation and ERE transactivation by inhibiting the recruitment of SRC-1 to ERα.

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