Browsing by Subject "Angiotensin"
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Item ANGIOTENSIN II RECEPTOR TYPE-1A KNOCKDOWN IN SUBFORNICAL ORGAN PREVENTS SUSTAINED INCREASE IN MEAN ARTERIAL PRESSURE ASSOCIATED WITH CHRONIC INTERMITTENT HYPOXIA(2014-03) Saxena, Ashwini; Little, Joel T.; Nedungadi, Thekkethil P.; Cunningham, J. ThomasPurpose (a): Sleep apnea (SA) is associated with a sustained increase in mean arterial pressure (MAP) even during waking hours. Chronic intermittent hypoxia (CIH) models the hypoxemia associated with SA and produces elevated MAP during CIH and normoxia. Angiotensin II (Ang II) is implicated in the CIH associated increase in MAP. Subfornical organ (SFO), a forebrain circumventricular organ, lacks blood brain barrier and is a major site for the central effects of circulating Ang II. We investigated the effects of Ang II type 1a receptor knockdown (AT1aRKD) in the SFO on CIH hypertension in adult male rats. Methods (b): Adeno-associated viral vectors carrying GFP and either AT1aR shRNA or scrambled shRNA (SCM) were injected in SFO. Continuous measurements of mean arterial pressure, heart rate, respiratory rate, and activity were measured using radio-telemetry device implanted in abdominal aorta. Rats were exposed to cyclic hypoxia (3 min 21% O2 - 3 min 10% O2) for 8 hours/day for 7 days. Rats were sacrificed on Day 8. Results (c): Using laser-capture microdissection and qRT-PCR of amino-allyl RNA, AT1aRKD rats showed decreased SFO AT1aRmRNA in comparison with SCM rats. During intermittently-hypoxic light phase, the AT1aRKD rats exposed to CIH (3 min 10% O2 and 3 min room air cycles for 8 h during light phase for 7 d) exhibited significant increases in MAP vs. AT1aRKD-Normoxia group (p(p<0.05). During the normoxic dark phase, there was no difference in MAP between CIH and normoxic AT1aRKD rats (p=0.69). SCM-CIH group showed significant increase in MAP from SCM-Normoxia group during light phase CIH (p<0.001) and the normoxic dark phase (p<0.001). Conclusions (d): Our data indicate that AT1aRs in SFO may play a role in the sustained increases in MAP during normoxia associated with CIH.Item AT1A RECEPTOR KNOCKDOWN IN THE MEDIAN PREOPTIC NUCLEUS ATTENUATES THE SUSTAINED COMPONENT OF HYPERTENSION RESULTING FROM CHRONIC INTERMITTENT HYPOXIA(2013-04-12) Shell, BrentPurpose: Obstructive sleep apnea produces hypertension and increases sympathetic nerve activity during the hypoxic sleeping events as well as during the waking hours. This study examined a possible receptor contribution to the sustained component of hypertension. Methods: To model the hypoxemia experienced during sleep apnea, rodents are exposed to chronic intermittent hypoxia (CIH). The median preoptic nucleus (MnPO) is a forebrain region that contributes to this sustained increase in mean arterial pressure (MAP) from CIH. This region integrates information from forebrain circumventricular organs and projects to the paraventricular nucleus to influence sympathetic nerve activity. Our lab has previously shown that in response to CIH there is increased ΔFosB expression in the MnPO and that after a 7 day hypoxia protocol MnPO AT1a receptor mRNA is increased 7 fold whereas AT1b shows no change. In this study, we tested the hypothesis that MnPO AT1a receptors contribute to CIH hypertension using adult male Sprague Dawley rats injected with a neuron specific adenoviral vectors with either shRNA to knock down expression of AT1a receptors in the MnPO or a scrambled RNA sequence. Results: All rats showed an increase in MAP during the hypoxic light period, but rats receiving the AT1a knock down did not exhibit the sustained increase in MAP during the normoxic dark phase (P<.05). Conclusions: This data indicates that the AT1a receptors in the MnPO is necessary for the sustained component of hypertension resulting from CIH.Item SRC-KINASE MEDIATES ANGIOTENSIN II INDUCED POTENTIATION IN TRPV4 AGONIST EVOKED CALCIUM TRANSIENTS IN HYPOTHALAMIC IMMORTALIZED NEURONAL CELL LINE 4B(2014-03) Saxena, Ashwini; Bachelor, Martha E.; Carreno, Flavia R.; Cunningham, J. ThomasWe have previously demonstrated that in bile duct ligated rats, an animal model of inappropriate vasopressin (AVP) release, TRPV4 protein expression and membrane trafficking is increased in AVP neurons. Here, we used an in vitro approach with an immortalized AVP expressing neuroendocrine cell line (4B neurons) to investigate the possible regulation of TRPV4 by angiotensin II (Ang II). We characterized the presence of TRPV4 mRNA and protein in 4B cells. Ang II (100nM;1 hr) treatment significantly increased TRPV4 abundance (p. Purpose (a): ØInappropriate Vasopressin (AVP) release causes dilutional hyponatremia associated with heart and liver failure. Although the central molecular mechanisms that mediate inappropriate AVP release are not clear, plasma angiotensin II (Ang II) has been implicated as a factor in the pathogenesis of dilutional hyponatremia. Our previous studies using a rodent model of liver failure, have shown that increased TRPV4 expression in vasopressinergic neurons and elevated circulating AVP were blunted by normalization of the renin angiotensin system (RAS). Effects of circulating Ang II on neural networks may mediate cellular adaptations associated with changes in TRPV4 expression and/or sorting ØBased on our in vivo studies we speculate that modulation of transient receptor potential vanilloid (TRPV4) channels by means of changes in its membrane sorting could alter its gating, and thus contribute to changes in neural excitability that would be consistent with increased AVP release in rats with liver failure. To examine the effects of AngII treatment upon TRPV4 we utilized the rat hypothalamic AVP expressing neuronal cell line 4B. Methods (b): We used Western Blots to detect changes in TRPV4 protein in membrane fraction after drug treatments. In addition, we used calcium sensitive dye Fura 2-AM to detect changes in intracellular calcium after administration of a selective TRPV4 agonist - GSK 1016790A. Results (c): We characterized the presence of TRPV4 mRNA and protein in 4B cells. After Ang II (100nM;1 hr) treatment significantly increased TRPV4 levels in crude membrane fractions (p<0.001) and tyrosine phosphorylation of TRPV4 (p<0.001). Using calcium sensitive dye Fura-2AM, we noted that Ang II treated cells exhibited increased calcium transients in response to TRPV4 agonist, GSK1016790A (20nM, p<0.05). This increase was blocked by the Losartan (Ang II receptor antagonist) and Src-kinase inhibitor, PP2, but not by its analog PP3. Conclusions (d): Our data indicate that Ang II may facilitate TRPV4 trafficking and alter the phosphorylation status of TRPV4 through Src-kinases.