Integrative Physiology

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    Role of A1/A2 Neurons in Increased Vasopressin Release in Male Bile Duct Ligated Rats
    (2020) Little, Joel; Cunningham, J. Thomas; Aikins, Ato
    Bile duct ligation (BDL), a rat model of liver failure causes increased vasopressin release leading to water retention and a decrease in plasma osmolality. Water retained in ascites leads to a perceived decrease in plasma volume by the A1/A2 norepinephrine neurons in the caudal ventrolateral medulla (CVLM) and the nucleus tractus solitarius (NTS) respectively, increasing vasopressin release. We hypothesized that lesioning norepinephrine A1/A2 neurons will prevent increased plasma vasopressin (measured as a function of plasma copeptin concentrations), normalize hematocrit and plasma osmolality, and decrease the number of immunoreactive ΔFosB and DβH cells in the NTS and CVLM. To investigate the role of A1/A2 neurons in the increase in vasopressin release observed in male BDL rats, anti-DβH-Saporin [IT-03] (Advanced Targeting Systems), or vehicle was injected into the SON. BDL surgery or sham surgery was performed after two weeks. Four weeks following the BDL/Sham surgery, the rats were sacrificed, and blood samples were taken for copeptin, hematocrit and plasma osmolality measurements. Hindbrains were processed for immunohistochemistry. The results showed a significant decrease in CVLM ΔFosB reactive cells in the Saporin/BDL group (n=2) as compared to the Vehicle/BDL group (n=3) (p< 0.001). DβH immunoreactive cells in both the CVLM and NTS were significantly decreased in the Saporin/BDL group as compared to the vehicle/BDL group (p< 0.001). However, differences were not observed for the copeptin concentration, hematocrit and plasma osmolality. The results suggest that lesions of SON-projecting A1/A2 neurons may not be sufficient to prevent vasopressin release associated with BDL.
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    Visit-to-Visit Reproducibility of Cerebral Vascular Reactivity to CO2 in Healthy Young Humans
    (2020) Anderson, Garen; Rickards, Caroline; Rosenberg, Alexander; Barnes, Haley; Hua, Vincent
    Partial pressure of arterial CO2 (PaCO2) is an important regulator of cerebral blood flow. The magnitude of change in cerebral blood flow per unit change in PaCO2 represents the cerebral vascular responsiveness to a CO2 stimulus, and is used as an index of cerebrovascular health. Accordingly, it is important to assess the reproducibility of this technique for clinical diagnoses. Purpose: To assess visit-to-visit reproducibility of a cerebral vascular reactivity to CO2 test in healthy young humans. Methods: Healthy adults (n=6, 25±2 y) performed a 5-min cerebral vascular reactivity to CO2 protocol (+5 mmHg end-tidal CO2 (etCO2) above eucapnic baseline) on two separate visits (7-234 days between visits). EtCO2 and middle cerebral artery velocity (MCAv) were measured continuously. Coefficient of variation (CV) was calculated for cerebral vascular reactivity to CO2 ((%Δ MCAv)/Δ etCO2), etCO2, and mean MCAv responses between the two visits. Results: While the CO2 stimulus between visits was similar (Visit 1: 5.3±0.2 mmHg vs. Visit 2: 5.1±0.3 mmHg, p=0.54) with a CV of 2.8%, there was a difference in the MCAv response to this stimulus (Visit 1: +15±1 cm/s vs. Visit 2: +12±2 cm/s, p=0.09; CV=19%). This resulted in a large variation in cerebral vascular reactivity to CO2 between visits (Visit 1: 4.6±0.3 %/mmHg vs. Visit 2: 3.6±0.5 %/mmHg, p=0.06; CV=18%). Conclusion: These findings suggest that despite a similar magnitude of change in the CO2 stimulus, physiological variations in cerebral vascular reactivity to CO2 occur in young healthy adults between two experimental visits.
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    Rats with Placental Ischemia and Preeclampsia-like Symptoms Have Increased Circulating Cell-free Mitochondrial DNA
    (2020) Goulopoulou, Styliani; Phillips, Nicole; Davidge, Sandra; Cushen, Spencer; Morton, Jude; Spaans, Floortje; Kirschenman, Raven; Gardner, Jennifer
    Preeclampsia (PE) is a hypertensive disorder of pregnancy, which is characterized by placental mitochondrial dysfunction. Increased circulating cell-free mitochondrial DNA (mtDNA) has been also reported in PE. Animal models are commonly used to study the role of placental dysfunction in the maternal syndrome of PE. The objective of this study was to determine the concentrations of circulating mtDNA in rat models of placental ischemia. Placental ischemia was induced in rats on gestational day (GD) 14 by placing clips on a) the abdominal aorta and ovarian arteries (reduced uterine perfusion pressure (RUPP)) and b) ovarian arteries and uterine arteries (selective RUPP (sRUPP)). Sham rats had clips placed on intraabdominal fat. Different groups of rats were exposed to hypoxia (11% O2) or maintained at atmospheric conditions (21% O2) from GD6 to GD21. Blood samples were collected on GD21. Real time PCR quantification of mtDNA was performed on DNA extracts from serum using TaqMan™ probes and chemistry. mtDNA copy number (CN) was greater in RUPP and sRUPP rats compared to their respective controls (Sham (11) vs. RUPP (11): 0.18 ± 0.04 CN/μl vs. 0.30 ± 0.04 CN/μl, p-value: 0.04; Sham (8) vs. sRUPP (10): 24.84 ± 3.29 CN/μl vs. 54.38 ± 3.29 CN/μl, p-value: 0.016)). Hypoxia did not affect mtDNA CN (Control (7) vs. Hypoxia (9): 0.28 ± 0.05 CN/μl vs. 0.36 ± 0.04 CN/μl, p-value: 0.28). Rats with placental ischemia have increased circulating cell-free mtDNA similar to what is seen in pregnant women with PE.
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    Effect of Sleep Apnea Treatment on Urine Flow in a Patient with BPH
    (2020) Smith, Michael; Chandra, Michael
    Benign Prostatic Hyperplasia (BPH) is a chronic condition presenting with lower urinary tract symptoms that affects up to 70% of US men 60and69 years of age and 80% of those 70 years or older (Wei et. al. 2005). Obstructive Sleep Apnea (OSA) is a chronic condition with excessive daytime fatigue and chronically elevated sympathetic nerve activity that affects at least 5-7% of men and 2-5% of women and increases significantly with age (Punjabi 2008). Moreover, both disorders often go undiagnosed. The increased sympathetic activity accompanying OSA likely affects BPH symptoms, however this has not been systematically investigated. Thus, this case study was a preliminary investigation of whether treatment of OSA alters BPH symptoms. A 58 year old adult male with a history of BPH and concurrent OSA with moderate compliance to using BiPAP for OSA treatment recorded urine flow and volume throughout a 4 month period during which the patient either did not use BiPaP at all, or used it throughout the night. Peak urine flow (UFlow meter) and net urine volume were recorded in the morning of each data collection. Results: A Mann-Whitney Rank Sum analysis revealed that peak flow was significantly increased (p< 0.05) and total urine volume was increased (p< 0.01) on nights in which there was complete BiPAP treatment. Conclusion: These results demonstrate that effective OSA treatment has a positive impact on BPH symptoms. This study suggests an interaction between OSA, its treatment, and BPH symptoms and this association warrants further investigation.
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    White Mountain Expedition 2019: The Impact of Sustained Hypoxia on Cerebral Blood Flow Responses and Tolerance to Simulated Hemorrhage
    (2020) Anderson, Garen; Rickards, Caroline; Barnes, Haley; Bird, Jordan; Pentz, Brandon; Byman, Britta; Jendzjowsky, Nicholas; Wilson, Richard; Day, Trevor; Rosenberg, Alexander
    Trauma-induced hemorrhage can occur at high altitude (HA) from a variety of causes, including battlefield injuries, motor vehicle/air accidents, and major falls. Based on the known compensatory increases in cerebral blood flow that occur with exposure to hypoxia, we hypothesized that tolerance to simulated hemorrhage (via application of lower body negative pressure, LBNP) at HA would be similar compared to low altitude (LA) due to increased cerebral blood flow and oxygen delivery, and the subsequent preservation of cerebral tissue oxygenation. Healthy human subjects (N=8; 4F/4M) participated in LBNP protocols to presyncope at LA (1045 m) and at HA (3800 m) following 4-5 days of acclimatization. Arterial pressure, heart rate, stroke volume, internal carotid artery (ICA) blood flow, and cerebral oxygen saturation were measured continuously. Time to presyncope was similar between conditions (LA: 1276±108s vs. HA: 1208±108s; P=0.58). Similar responses to LBNP were observed at LA and HA in mean arterial pressure (LA: -16±2% vs. HA: -16±2%; P=0.85), stroke volume (LA: -57±5% vs. HA: -60±5%; P=0.39), and heart rate (LA: +69±12% vs. HA: +65±8%; P=0.71). ICA blood flow was higher at HA vs. LA (P=0.01), and decreased with LBNP under both conditions (P≤0.005), with no effect of altitude on cerebral oxygen saturation (P=0.73). These findings suggest that hypoxia with ascent to 3800 m does not affect tolerance to simulated hemorrhage in young healthy adults, which may be due to 1) similar cardiovascular reflex responses, and/or 2) compensatory increases in cerebral blood flow and subsequent preservation of cerebral tissue oxygenation.
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    The Effect of Sex on GABAA Receptor Activation on Vasopressin Neurons from the Supraoptic Nucleus
    (2020) Little, Joel; Cunningham, J. Thomas; Farmer, George; Bachelor, Martha; Balapattabi, Kirthikaa; Brock, Courtney
    Arginine Vasopressin (AVP), a hormone produced by the magnocellular neurosecretory cells (MNC) of the supraoptic nucleus (SON) and paraventricular nucleus (PVN) of the hypothalamus regulates fluid balance and plasma osmolality in healthy individuals. During congestive heart failure (CHF) and liver failure, however, AVP becomes dysregulated leading to increased renal water reabsorption and consequently persistent hyponatremia. Previous work done in the laboratory suggests that female rats may be protected from such dysregulation, perhaps by hormones such as estrogen. Our hypothesis is that under normal physiological conditions, activation of the GABAA receptor inhibits vasopressin neurons within the SON by causing chloride influx, but that under pathophyisiological conditions, activation of the GABAA receptor induces chloride efflux thereby causing activation of the vasopressin neurons in males. It is not known if AVP neurons from female rats behave in a similar manner to AVP neurons from males or are influenced by the estrous cycle. To test our hypothesis, the SON of adult, intact Sprague Dawley rats of both sexes were bilaterally injected with the ClophensorN virus which is a genetically modified virus used to detect Chloride flux in vasopressin neurons using fluorescence. Dissociated neurons from the SON were tested to assess chloride flux in response to the GABAA receptor agonist muscimol. Results show that under normal physiological conditions, muscimol induces Chloride influx in both males and females. Future experiments will assess the effect of GABAA activation in pathophysiological conditions by using the bile-duct ligation model of liver failure.
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    Sex Differences in the Development of Hypertension in the Setting of Autoimmunity
    (2020) Morales, Jessica; Mathis, Keisa W.; Young-Stubbs, Cassandra M.; D'Souza, Bradley
    Purpose: Systemic lupus erythematosus (SLE) is a chronic autoimmune inflammatory disease associated with high risks of hypertension. We previously confirmed the disease develops later in life in male SLE mice compared to females (35 vs.< 30 weeks). We also found that both male and female SLE mice are hypertensive by 35 weeks and that this hypertension is linked to renal injury in females but not males; therefore, we aimed to investigate potential contributors to the latent sex difference. Toll-like receptor 7 (TLR7) is an immune mediator active in autoimmunity that instigates widespread tissue damage. We hypothesized that increased TLR7 promotes renal injury in female SLE mice and a different mechanism, potentially increased renal vascular resistance (RVR), is responsible for the hypertension in male SLE mice. Methods: Renal cortical expression of TLR7 was assessed via Western blot in male and female SLE mice (NZBWF1) at 35 weeks. Renal blood flow and mean arterial pressure were measured in anesthetized male and female SLE mice to determine RVR. Results: Female SLE mice had higher (p< 0.05) expression of TLR7 (2.6e6 ±5.4e5; normalized to total protein) than males (1.7e6 ± 3.3e5). Male SLE mice had lower RVR than females (5.15 ±0.60 vs. 10.07 ±1.23 mmHg·min·kg·mL-1). Conclusion: Our data suggest that while the hypertension in female SLE mice may be due to renal mechanisms, male SLE mice develop hypertension through other mechanisms. Future studies will continue to dissect sex-specific factors that should be considered when treating hypertensive patients with underlying chronic inflammation.
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    Role of Estrogen Receptors in a Model of Dilutional Hyponatremia
    (2020) Balapattabi, Kirthikaa; Little, Joel; Cunningham, J. Thomas; Nguyen, John-Bosco; Brock, Courtney; Nguyen, Dianna
    Purpose: Hyponatremia is the most frequently occurring electrolyte disorder and independent risk factor for increased patient mortality. Dilutional hyponatremia in liver failure due to inappropriate arginine vasopressin (AVP) release can be studied using rodent bile-duct ligation (BDL) model. Our previous sex differences studies in BDL rats show compared to males, female and ovariectomized (OVX) BDL rats did not develop hyponatremia, AVP neuron activation, or increased plasma copeptin (CPP; a marker for AVP), compared to sham ligated females. Due to increased adrenal and circulating estradiol (E2) in OVX BDL rats, the role of E2 was unclear. Intracerebroventricular infusion of estrogen receptor (ER) antagonist, ICI 182,780 (ICI) in female BDL rats increased CPP concentration compared to controls. These data suggest ER involvement in prevention of hyponatremia in female BDL rats. However, ICI is also a G protein-coupled estrogen receptor 1 (GPER) agonist. We tested GPER expression within hypothalamo-neurohypophyseal system of female rats. Methods: Immunohistochemistry was performed on three separate sets of forebrain sections from adult female Sprague-Dawley rats. All sets processed for GPER, and the separate sets stained for either AVP, oxytocin (OXY), or glia fibrillary acidic protein (GFAP). Results: Co-localization of GPER+AVP and GPER+OXY was observed in neurohypophyseal neurons (GPER+AVP, 64.8% and GPER+OXY, 64.0%). GPER+GFAP co-localization was not observed. Conclusion: GPER is expressed on subset of AVP and OXY neurons and not astrocytes in hypothalamo-neurohypophyseal system of female rats. Future studies in BDL rats will provide further insight about sex differences in neurohypophyseal function.