46 research outputs found
Ryanodine receptor and capacitative Ca2+ entry in fresh preglomerular vascular smooth muscle cells
Get PDFRyanodine receptor and capacitative Ca2+ entry in fresh preglomerular vascular smooth muscle cells.BackgroundA multiplicity of hormonal, neural, and paracrine factors regulates preglomerular arterial tone by stimulating calcium entry or mobilization. We have previously provided evidence for capacitative (store-operated) Ca2+ entry in fresh renal vascular smooth muscle cells (VSMCs). Ryanodine-sensitive receptors (RyRs) have recently been identified in a variety of nonrenal vascular beds.MethodsWe isolated fresh rat preglomerular VSMCs with a magnetized microsphere/sieving technique; cytosolic Ca2+ ([Ca2+]i) was measured with fura-2 ratiometric fluorescence.ResultsRyanodine (3 μmol/L) increased [Ca2+]i from 79 to 138 nmol/L (P = 0.01). Nifedipine (Nif), given before or after ryanodine, was without effect. The addition of calcium (1 mmol/L) to VSMCs in calcium-free buffer did not alter resting [Ca2+]i. In Ca-free buffer containing Nif, [Ca2+]i rose from 61 to 88 nmol/L after the addition of the Ca2+-ATPase inhibitor cyclopiazonic acid and to 159 nmol/L after the addition of Ca2+ (1 mmol/L). Mn2+ quenched the Ca/fura signal, confirming divalent cation entry. In Ca-free buffer with Nif, [Ca2+]i increased from 80 to 94 nmol/L with the addition of ryanodine and further to 166 nmol/L after the addition of Ca2+ (1 mmol/L). Mn2+ quenching was again shown. Thus, emptying of the sarcoplasmic reticulum (SR) with ryanodine stimulated capacitative Ca2+ entry.ConclusionPreglomerular VSMCs have functional RyR, and a capacitative (store-operated) entry mechanism is activated by the depletion of SR Ca2+ with ryanodine, as is the case with inhibitors of SR Ca2+-ATPase
Modulation of the myogenic mechanism: concordant effects of NO synthesis inhibition and O 2 − dismutation on renal autoregulation in the time and frequency domains
Get PDFRenal blood flow autoregulation was investigated in anesthetized C57Bl6 mice using time- and frequency-domain analyses. Autoregulation was reestablished by 15 s in two stages after a 25-mmHg step increase in renal perfusion pressure (RPP). The renal vascular resistance (RVR) response did not include a contribution from the macula densa tubuloglomerular feedback mechanism. Inhibition of nitric oxide (NO) synthase [NG-nitro-l-arginine methyl ester (l-NAME)] reduced the time for complete autoregulation to 2 s and induced 0.25-Hz oscillations in RVR. Quenching of superoxide (SOD mimetic tempol) during l-NAME normalized the speed and strength of stage 1 of the RVR increase and abolished oscillations. The slope of stage 2 was unaffected by l-NAME or tempol. These effects of l-NAME and tempol were evaluated in the frequency domain during random fluctuations in RPP. NO synthase inhibition amplified the resonance peak in admittance gain at 0.25 Hz and markedly increased the gain slope at the upper myogenic frequency range (0.06–0.25 Hz, identified as stage 1), with reversal by tempol. The slope of admittance gain in the lower half of the myogenic frequency range (equated with stage 2) was not affected by l-NAME or tempol. Our data show that the myogenic mechanism alone can achieve complete renal blood flow autoregulation in the mouse kidney following a step increase in RPP. They suggest also that the principal inhibitory action of NO is quenching of superoxide, which otherwise potentiates dynamic components of the myogenic constriction in vivo. This primarily involves the first stage of a two-stage myogenic response
<html>Activation of Phospholipase Cγ<sub>1</sub> Protects Renal Arteriolar VSMCs from H<sub>2</sub>O<sub>2</sub>-Induced Cell Death</html>
Get PDFWe evaluated the effect of hydrogen peroxide (H2O2) on viability of vascular smooth muscle cells (VSMCs) of renal resistance arterioles and determined whether responses are modulated by activation of PLCγ1
Endothelin A and B receptors of preglomerular vascular smooth muscle cells
Get PDFBACKGROUND: The endothelin (ET) receptors are subclassified into ET(A,) which are purely vasoconstrictive, and ET(B). The ET(B) receptors may cause either vasodilation by stimulating the release of nitric oxide from endothelial cells, or vasoconstriction of vascular smooth muscle cells (VSMC). The relative contribution of ET(A) and ET(B) receptors to calcium signaling and vasoconstriction in the renal microcirculation is not clear. Our goal was to study the cytosolic calcium concentration ([Ca(2+)](i)) responses of fresh rat preglomerular VSMC and afferent arterioles to agonists and antagonists of ET(A) and ET(B) receptors in rats. METHODS: Fresh VSMC and afferent arterioles were isolated using the magnetized microsphere/sieving technique, followed by gentle collagenase digestion. [Ca(2+)](i) was measured with fura-2 ratiometric fluorescence. RESULTS: Afferent arterioles and VSMC responded to ET-1 stimulation with a rapid peak increase in [Ca(2+)](i) (Delta= 287 +/- 81 and 342 +/- 55 nmol/L, respectively). The ET(B) receptor agonist IRL 1620 stimulated a rise in [Ca(2+)](i) in afferent arterioles (106 +/- 35 nmol/L); subsequent addition of ET-1 at the IRL 1620 nadir to stimulate ET(A) receptors caused a second peak that was twice as large (213 +/- 44 nmol/L). In VSMC, the ET(B) agonist peak increase was 99 +/- 12 nmol/L; addition of ET-1 then increased [Ca(2+)](i) by 294 +/- 23 nmol/L. The ET(B) inhibitor BQ-788 prevented stimulation of [Ca(2+)](i) by IRL 1620 in afferent arterioles and VSMC; subsequent stimulation of ET(A) receptors with ET-1 caused an increase in [Ca(2+)](i) (239 +/- 17 and 248 +/- 22 nmol/L). Pretreatment with the selective ET(A) inhibitor PD 156707 attenuated but did not abolish the responses to ET-1, suggesting that the residual [Ca(2+)](i) response was caused by ET(B) stimulation. CONCLUSION: These results indicate that fresh preglomerular VSMC as well as afferent arterioles have both ET(A) and ET(B) receptors, and that the rapid peak [Ca(2+)](i) responses to the ET(B) agonist IRL 1620 are less than half that of subsequent stimulation of ET(A) receptors with ET-1. The similarity of findings in isolated VSMC and afferent arterioles suggests that responses in VSMC in our arteriolar preparation overshadow any potential contribution of endothelial cells when reagents are administered abluminally
Superoxide Enhances Ca2+ Entry Through L-Type Channels in the Renal Afferent Arteriole
Get PDFReactive oxygen species regulate cardiovascular and renal function in health and disease. Superoxide participates in acute calcium signaling in afferent arterioles and renal vasoconstriction produced by angiotensin II, endothelin, thromboxane, and pressure-induced myogenic tone. Known mechanisms by which superoxide acts include quenching of nitric oxide and increased ADP ribosyl cyclase/ryanodine-mediated calcium mobilization. The effect(s) of superoxide on other calcium signaling pathways in the renal microcirculation is poorly understood. The present experiments examined the acute effect of superoxide generated by paraquat on calcium entry pathways in isolated rat afferent arterioles. The peak increase in cytosolic calcium concentration caused by KCl (40 mmol/L) was 99±14 nmol/L. The response to this membrane depolarization was mediated exclusively by L-type channels because it was abolished by nifedipine but was unaffected by the T-type channel blocker mibefradil. Paraquat increased superoxide production (dihydroethidium fluorescence), tripled the peak response to KCl to 314±68 nmol/L ( P <0.001) and doubled the plateau response. These effects were abolished by tempol and nitroblue tetrazolium, but not by catalase, confirming actions of superoxide and not of hydrogen peroxide. Unaffected by paraquat and superoxide was calcium entry through store-operated calcium channels activated by thapsigargin-induced calcium depletion of sarcoplasmic reticular stores. Also unresponsive to paraquat was ryanodine receptor–mediated calcium-induced calcium release from the sarcoplasmic reticulum. Our results provide new evidence that superoxide enhances calcium entry through L-type channels activated by membrane depolarization in rat cortical afferent arterioles, without affecting calcium entry through store-operated entry or ryanodine receptor–mediated calcium mobilization. # Novelty and Significance {#article-title-68
Validation of Dynamic Contrast-Enhanced Ultrasound in Rodent Kidneys as an Absolute Quantitative Method for Measuring Blood Perfusion
Get PDFContrast-enhanced ultrasound (CEUS) has demonstrated utility in the monitoring of blood flow in tissues, organs, and tumors. However, current CEUS methods typically provide only relative image-derived measurements, rather than quantitative values of blood flow in milliliters/minute per gram of tissue. In this study, CEUS derived parameters of blood flow are compared to absolute measurements of blood flow in rodent kidneys. Additionally, the effect of contrast agent infusion rate and transducer orientation on image-derived perfusion measurements are assessed. Both wash-in curve and time-to-refill algorithms are examined. Data illustrate that for all conditions, image-derived flow measurements were well-correlated with transit-time flow probe measurements (R > 0.9). However, we report differences in the sensitivity to flow across different transducer orientations as well as the contrast analysis algorithm utilized. Results also indicate that there exists a range of contrast agent flow rates for which image-derived estimates are consistent
Caffeine intake antagonizes salt sensitive hypertension through improvement of renal sodium handling
Get PDFHigh salt intake is a major risk factor for hypertension. Although acute caffeine intake produces moderate diuresis and natriuresis, caffeine increases the blood pressure (BP) through activating sympathetic activity. However, the long-term effects of caffeine on urinary sodium excretion and blood pressure are rarely investigated. Here, we investigated whether chronic caffeine administration antagonizes salt sensitive hypertension by promoting urinary sodium excretion. Dahl salt-sensitive (Dahl-S) rats were fed with high salt diet with or without 0.1% caffeine in drinking water for 15 days. The BP, heart rate and locomotor activity of rats was analyzed and urinary sodium excretion was determined. The renal epithelial Na+ channel (ENaC) expression and function were measured by in vivo and in vitro experiments. Chronic consumption of caffeine attenuates hypertension induced by high salt without affecting sympathetic nerve activity in Dahl-S rats. The renal α-ENaC expression and ENaC activity of rats decreased after chronic caffeine administration. Caffeine increased phosphorylation of AMPK and decrease α-ENaC expression in cortical collecting duct cells. Inhibiting AMPK abolished the effect of caffeine on α-ENaC. Chronic caffeine intake prevented the development of salt-sensitive hypertension through promoting urinary sodium excretion, which was associated with activation of renal AMPK and inhibition of renal tubular ENaC
Activation of TRPV1 by Dietary Capsaicin Improves Endothelium-Dependent Vasorelaxation and Prevents Hypertension
Get PDFSome plant-based diets lower the cardiometabolic risks and prevalence of hypertension. New evidence implies a role for the transient receptor potential vanilloid 1 (TRPV1) cation channel in the pathogenesis of cardiometabolic diseases. Little is known about impact of chronic TRPV1 activation on the regulation of vascular function and blood pressure. Here we report that chronic TRPV1 activation by dietary capsaicin increases the phosphorylation of protein kinase A (PKA) and eNOS and thus production of nitric oxide (NO) in endothelial cells, which is calcium dependent. TRPV1 activation by capsaicin enhances endothelium-dependent relaxation in wild-type mice, an effect absent in TRPV1-deficient mice. Long-term stimulation of TRPV1 can activate PKA, which contributes to increased eNOS phosphorylation, improves vasorelaxation, and lowers blood pressure in genetically hypertensive rats. We conclude that TRPV1 activation by dietary capsaicin improves endothelial function. TRPV1-mediated increase in NO production may represent a promising target for therapeutic intervention of hypertension
Oligoclonal CD8+ T Cells Play a Critical Role in the Development of Hypertension
Get PDFRecent studies have emphasized a role of adaptive immunity, and particularly T cells, in the genesis of hypertension. We sought to determine the T-cell subtypes that contribute to hypertension and renal inflammation in angiotensin II-induced hypertension. Using T-cell receptor spectratyping to examine T-cell receptor usage, we demonstrated that CD8(+) cells, but not CD4(+) cells, in the kidney exhibited altered T-cell receptor transcript lengths in Vβ3, 8.1, and 17 families in response to angiotensin II-induced hypertension. Clonality was not observed in other organs. The hypertension caused by angiotensin II in CD4(-/-) and MHCII(-/-) mice was similar to that observed in wild-type mice, whereas CD8(-/-) mice and OT1xRAG-1(-/-) mice, which have only 1 T-cell receptor, exhibited a blunted hypertensive response to angiotensin II. Adoptive transfer of pan T cells and CD8(+) T cells but not CD4(+)/CD25(-) cells conferred hypertension to RAG-1(-/-) mice. In contrast, transfer of CD4(+)/CD25(+) cells to wild-type mice receiving angiotensin II decreased blood pressure. Mice treated with angiotensin II exhibited increased numbers of kidney CD4(+) and CD8(+) T cells. In response to a sodium/volume challenge, wild-type and CD4(-/-) mice infused with angiotensin II retained water and sodium, whereas CD8(-/-) mice did not. CD8(-/-) mice were also protected against angiotensin-induced endothelial dysfunction and vascular remodeling in the kidney. These data suggest that in the development of hypertension, an oligoclonal population of CD8(+) cells accumulates in the kidney and likely contributes to hypertension by contributing to sodium and volume retention and vascular rarefaction
Review: Lessons on renal function from transgenic mice lacking different angiotensin II receptors
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