scholarly journals ATP stimulates rat hypothalamic sympathetic neurons by enhancing AMPA receptor-mediated currents

2015 ◽  
Vol 114 (1) ◽  
pp. 159-169 ◽  
Author(s):  
Hildebrando Candido Ferreira-Neto ◽  
Vagner R. Antunes ◽  
Javier E. Stern
Keyword(s):  
Ampa Receptor ◽  
Ampa Receptors ◽  
Kynurenic Acid ◽  
Sympathetic Neurons ◽  
Synaptic Function ◽  
Ventrolateral Medulla ◽  
Cellular Mechanisms ◽  
Receptor Coupling ◽  
Whole Cell Patch Clamp ◽  
Intracellular Ca

We have previously shown that ATP within the paraventricular nucleus (PVN) induces an increase in sympathetic activity, an effect attenuated by the antagonism of P2 and/or glutamatergic receptors. Here, we evaluated precise cellular mechanisms underlying the ATP-glutamate interaction in the PVN and assessed whether this receptor coupling contributed to osmotically driven sympathetic PVN neuronal activity. Whole-cell patch-clamp recordings obtained from PVN-rostral ventrolateral medulla neurons showed that ATP (100 μM, 1 min, bath applied) induced an increase in firing rate (89%), an effect blocked by kynurenic acid (1 mM) or 4-[[4-Formyl-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]-2-pyridinyl]azo]-1,3-benzenedisulfonic acid tetrasodium salt (PPADS) (10 μM). Whereas ATP did not affect glutamate synaptic function, α-amino-3-hydroxy-5-methylisoxazole propionic acid (AMPA) receptor-mediated currents evoked by focal application of AMPA (50 μM, n = 13) were increased in magnitude by ATP (AMPA amplitude: 33%, AMPA area: 52%). ATP potentiation of AMPA currents was blocked by PPADS ( n = 12) and by chelation of intracellular Ca2+ (BAPTA, n = 10). Finally, a hyperosmotic stimulus (mannitol 1%, +55 mosM, n = 8) potentiated evoked AMPA currents (53%), an effect blocked by PPADS ( n = 6). Taken together, our data support a functional stimulatory coupling between P2 and AMPA receptors (likely of extrasynaptic location) in PVN sympathetic neurons, which is engaged in response to an acute hyperosmotic stimulus, which might contribute in turn to osmotically driven sympathoexcitatory responses by the PVN.

scholarly journals Hypertension Induced by Angiotensin II and a High Salt Diet Involves Reduced SK Current and Increased Excitability of RVLM Projecting PVN Neurons

2010 ◽  
Vol 104 (5) ◽  
pp. 2329-2337 ◽  
Author(s):  
Qing-Hui Chen ◽  
Mary Ann Andrade ◽  
Alfredo S. Calderon ◽  
Glenn M. Toney
Keyword(s):  
Angiotensin Ii ◽  
Input Resistance ◽  
High Salt ◽  
Ventrolateral Medulla ◽  
Sk Channels ◽  
Hypertensive Rats ◽  
Cellular Mechanisms ◽  
Hypertensive Group ◽  
Whole Cell Patch Clamp ◽  
The Impact

Although evidence indicates that activation of presympathetic paraventricular nucleus (PVN) neurons contributes to the pathogenesis of salt-sensitive hypertension, the underlying cellular mechanisms are not fully understood. Recent evidence indicates that small conductance Ca2+-activated K+ (SK) channels play a significant role in regulating the excitability of a key group of sympathetic regulatory PVN neurons, those with axonal projections to the rostral ventrolateral medulla (RVLM; i.e., PVN-RVLM neurons). In the present study, rats consuming a high salt (2% NaCl) diet were made hypertensive by systemic infusion of angiotensin II (AngII), and whole cell patch-clamp recordings were made in brain slice from retrogradely labeled PVN-RVLM neurons. To determine if the amplitude of SK current was altered in neurons from hypertensive rats, voltage-clamp recordings were performed to isolate SK current. Results indicate that SK current amplitude ( P < 0.05) and density ( P < 0.01) were significantly smaller in the hypertensive group. To investigate the impact of this on intrinsic excitability, current-clamp recordings were performed in separate groups of PVN-RVLM neurons. Results indicate that the frequency of spikes evoked by current injection was significantly higher in the hypertensive group ( P < 0.05–0.01). Whereas bath application of the SK channel blocker apamin significantly increased discharge of neurons from normotensive rats ( P < 0.05–0.01), no effect was observed in the hypertensive group. In response to ramp current injections, subthreshold depolarizing input resistance was greater in the hypertensive group compared with the normotensive group ( P < 0.05). Blockade of SK channels increased depolarizing input resistance in normotensive controls ( P < 0.05) but had no effect in the hypertensive group. On termination of current pulses, a medium afterhyperpolarization potential (mAHP) was observed in most neurons of the normotensive group. In the hypertensive group, the mAHP was either small or absent. In the latter case, an afterdepolarization potential (ADP) was observed that was unaffected by apamin. Apamin treatment in the normotensive group blocked the mAHP and revealed an ADP resembling that seen in the hypertensive group. We conclude that diminished SK current likely underlies the absence of mAHPs in PVN-RVLM neurons from hypertensive rats. Both the ADP and greater depolarizing input resistance likely contribute to increased excitability of PVN-RVLM neurons from rats with AngII-Salt hypertension.

scholarly journals Identifying cellular mechanisms of zinc-induced relaxation in isolated cardiomyocytes

2013 ◽  
Vol 305 (5) ◽  
pp. H706-H715 ◽  
Author(s):  
Ting Yi ◽  
Jonathan S. Vick ◽  
Marc J. H. Vecchio ◽  
Kelly J. Begin ◽  
Stephen P. Bell ◽  
...  
Keyword(s):  
Calcium Channel ◽  
Relaxation Function ◽  
Competitive Inhibition ◽  
Systolic Function ◽  
Cellular Mechanisms ◽  
Tension Development ◽  
Whole Cell Patch Clamp ◽  
Intracellular Ca ◽  
Channel Reduction ◽  
Future Work

We tested several molecular and cellular mechanisms of cardiomyocyte contraction-relaxation function that could account for the reduced systolic and enhanced diastolic function observed with exposure to extracellular Zn2+. Contraction-relaxation function was monitored in isolated rat and mouse cardiomyocytes maintained at 37°C, stimulated at 2 or 6 Hz, and exposed to 32 μM Zn2+ or vehicle. Intracellular Zn2+ detected using FluoZin-3 rose to a concentration of ∼13 nM in 3–5 min. Peak sarcomere shortening was significantly reduced and diastolic sarcomere length was elongated after Zn2+ exposure. Peak intracellular Ca2+ detected by Fura-2FF was reduced after Zn2+ exposure. However, the rate of cytosolic Ca2+ decline reflecting sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA2a) activity and the rate of Na+/Ca2+ exchanger activity evaluated by rapid Na+-induced Ca2+ efflux were unchanged by Zn2+ exposure. SR Ca2+ load evaluated by rapid caffeine exposure was reduced by ∼50%, and L-type calcium channel inward current measured by whole cell patch clamp was reduced by ∼70% in cardiomyocytes exposed to Zn2+. Furthermore, ryanodine receptor (RyR) S2808 and phospholamban (PLB) S16/T17 were markedly dephosphorylated after perfusing hearts with 50 μM Zn2+. Maximum tension development and thin-filament Ca2+ sensitivity in chemically skinned cardiac muscle strips were not affected by Zn2+ exposure. These findings suggest that Zn2+ suppresses cardiomyocyte systolic function and enhances relaxation function by lowering systolic and diastolic intracellular Ca2+ concentrations due to a combination of competitive inhibition of Ca2+ influx through the L-type calcium channel, reduction of SR Ca2+ load resulting from phospholamban dephosphorylation, and lowered SR Ca2+ leak via RyR dephosphorylation. The use of the low-Ca2+-affinity Fura-2FF likely prevented the detection of changes in diastolic Ca2+ and SERCA2a function. Other strategies to detect diastolic Ca2+ in the presence of Zn2+ are essential for future work.

Enhanced ETA-receptor-mediated inhibition of Kv channels in hypoxic hypertensive rat pulmonary artery myocytes

1999 ◽  
Vol 277 (1) ◽  
pp. H363-H370 ◽  
Author(s):  
Kai-Xun Li ◽  
Brian Fouty ◽  
Ivan F. McMurtry ◽  
David M. Rodman
Keyword(s):  
Pulmonary Hypertension ◽  
Pulmonary Artery ◽  
Hypoxic Pulmonary Hypertension ◽  
Voltage Gated ◽  
Receptor Coupling ◽  
Kv Channels ◽  
Whole Cell Patch Clamp ◽  
Intracellular Ca ◽  
Eta Receptor

Endothelin (ET)-1 has been implicated as a critical mediator in the pathogenesis of hypoxic pulmonary hypertension. We questioned whether, during exposure to chronic hypobaric hypoxia, rat pulmonary artery smooth muscle cells (PASMC) became sensitized to ET-1. Two effects of ET-1, inhibition of voltage-gated K+(Kv) channels and release of intracellular Ca2+, were studied using whole cell patch clamp and single cell indo 1 fluorescence, respectively. In both normotensive and chronically hypoxic-hypertensive PASMC, ET-1 caused concentration-dependent inhibition of voltage-gated K+ current [ I K(v)], with maximum inhibition of 12–18% seen at a concentration of 0.1–1 nM. Although the chronically hypoxic-hypertensive PASMC was no more susceptible to ET-1-mediated I K(v) inhibition, a switch in coupling between ET-1 and I K(v) from ETB to ETA receptors occurred. This switch in receptor coupling, combined with reduced I K(v) density and increased ET-1 production in the hypoxic rat lung, may help explain the ability of ETA-receptor blockers to attenuate the development of hypoxic pulmonary hypertension in vivo.

AMPA Receptor Antagonist CFM-2 Decreases Survivin Expression in Cancer Cells

2018 ◽  
Vol 18 (4) ◽  
pp. 591-596 ◽  
Author(s):  
Domingo Sanchez Ruiz ◽  
Hella Luksch ◽  
Marco Sifringer ◽  
Achim Temme ◽  
Christian Staufner ◽  
...  
Keyword(s):  
Cell Survival ◽  
Receptor Antagonist ◽  
Cancer Cells ◽  
Glutamate Receptors ◽  
Cancer Cell ◽  
Ampa Receptor ◽  
Ampa Receptors ◽  
Survivin Expression ◽  
Ampa Receptor Antagonist ◽  
Cancer Cell Survival

Background: Glutamate receptors are widely expressed in different types of cancer cells. α-Amino-3- hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors are ionotropic glutamate receptors which are coupled to intracellular signaling pathways that influence cancer cell survival, proliferation, and migration. Blockade of AMPA receptors by pharmacologic compounds may potentially constitute an effective tool in anticancer treatment strategies. Method: Here we investigated the impact of the AMPA receptor antagonist CFM-2 on the expression of the protein survivin, which is known to promote cancer cell survival and proliferation. We show that CFM-2 inhibits survivin expression at mRNA and protein levels and decreases the viability of cancer cells. Using a stably transfected cell line which overexpresses survivin, we demonstrate that over-expression of survivin enhances cancer cell viability and attenuates CFM-2–mediated inhibition of cancer cell growth. Result: These findings point towards suppression of survivin expression as a new mechanism contributing to anticancer effects of AMPA antagonists.

scholarly journals Control of AMPA receptor activity by the extracellular loops of auxiliary proteins

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Irene Riva ◽  
Clarissa Eibl ◽  
Rudolf Volkmer ◽  
Anna L Carbone ◽  
Andrew JR Plested
Keyword(s):  
Ampa Receptor ◽  
Ampa Receptors ◽  
De Novo ◽  
Dominant Role ◽  
Mammalian Brain ◽  
Binding Assays ◽  
Receptor Kinetics ◽  
Receptor Complexes ◽  
Extracellular Loops ◽  
Kinetics Of

At synapses throughout the mammalian brain, AMPA receptors form complexes with auxiliary proteins, including TARPs. However, how TARPs modulate AMPA receptor gating remains poorly understood. We built structural models of TARP-AMPA receptor complexes for TARPs γ2 and γ8, combining recent structural studies and de novo structure predictions. These models, combined with peptide binding assays, provide evidence for multiple interactions between GluA2 and variable extracellular loops of TARPs. Substitutions and deletions of these loops had surprisingly rich effects on the kinetics of glutamate-activated currents, without any effect on assembly. Critically, by altering the two interacting loops of γ2 and γ8, we could entirely remove all allosteric modulation of GluA2, without affecting formation of AMPA receptor-TARP complexes. Likewise, substitutions in the linker domains of GluA2 completely removed any effect of γ2 on receptor kinetics, indicating a dominant role for this previously overlooked site proximal to the AMPA receptor channel gate.

Fluid pressure modulates L-type Ca2+ channel via enhancement of Ca2+-induced Ca2+ release in rat ventricular myocytes

2008 ◽  
Vol 294 (4) ◽  
pp. C966-C976 ◽  
Author(s):  
Sunwoo Lee ◽  
Joon-Chul Kim ◽  
Yuhua Li ◽  
Min-Jeong Son ◽  
Sun-Hee Woo
Keyword(s):  
Ventricular Myocytes ◽  
Fluid Pressure ◽  
Inhibitory Effect ◽  
Cellular Mechanism ◽  
Confocal Imaging ◽  
Rat Ventricular Myocytes ◽  
Whole Cell Patch Clamp ◽  
Current Voltage ◽  
Intracellular Ca ◽  
High Concentrations

This study examines whether fluid pressure (FP) modulates the L-type Ca2+ channel in cardiomyocytes and investigates the underlying cellular mechanism(s) involved. A flow of pressurized (∼16 dyn/cm2) fluid, identical to that bathing the myocytes, was applied onto single rat ventricular myocytes using a microperfusion method. The Ca2+ current ( ICa) and cytosolic Ca2+ signals were measured using a whole cell patch-clamp and confocal imaging, respectively. It was found that the FP reversibly suppressed ICa (by 25%) without altering the current-voltage relationships, and it accelerated the inactivation of ICa. The level of ICa suppression by FP depended on the level and duration of pressure. The Ba2+ current through the Ca2+ channel was only slightly decreased by the FP (5%), suggesting an indirect inhibition of the Ca2+ channel during FP stimulation. The cytosolic Ca2+ transients and the basal Ca2+ in field-stimulated ventricular myocytes were significantly increased by the FP. The effects of the FP on the ICa and on the Ca2+ transient were resistant to the stretch-activated channel inhibitors, GsMTx-4 and streptomycin. Dialysis of myocytes with high concentrations of BAPTA, the Ca2+ buffer, eliminated the FP-induced acceleration of ICa inactivation and reduced the inhibitory effect of the FP on ICa by ≈80%. Ryanodine and thapsigargin, abolishing sarcoplasmic reticulum Ca2+ release, eliminated the accelerating effect of FP on the ICa inactivation, and they reduced the inhibitory effect of FP on the ICa. These results suggest that the fluid pressure indirectly suppresses the Ca2+ channel by enhancing the Ca2+-induced intracellular Ca2+ release in rat ventricular myocytes.

scholarly journals Novel neuronal and astrocytic mechanisms in thalamocortical loop dynamics

2002 ◽  
Vol 357 (1428) ◽  
pp. 1675-1693 ◽  
Author(s):  
Vincenzo Crunelli ◽  
Kate L. Blethyn ◽  
David W. Cope ◽  
Stuart W. Hughes ◽  
H. Rheinallt Parri ◽  
...  
Keyword(s):  
Low Voltage ◽  
Electrical Coupling ◽  
Low Frequency ◽  
Network Activity ◽  
Thalamic Neuron ◽  
Cellular Mechanisms ◽  
Brain Areas ◽  
Sensory Stimuli ◽  
Loop Dynamics ◽  
Intracellular Ca

In this review, we summarize three sets of findings that have recently been observed in thalamic astrocytes and neurons, and discuss their significance for thalamocortical loop dynamics. (i) A physiologically relevant ‘window’ component of the low–voltage–activated, T–type Ca 2+ current ( I Twindow ) plays an essential part in the slow (less than 1 Hz) sleep oscillation in adult thalamocortical (TC) neurons, indicating that the expression of this fundamental sleep rhythm in these neurons is not a simple reflection of cortical network activity. It is also likely that I Twindow underlies one of the cellular mechanisms enabling TC neurons to produce burst firing in response to novel sensory stimuli. (ii) Both electrophysiological and dye–injection experiments support the existence of gap junction–mediated coupling among young and adult TC neurons. This finding indicates that electrical coupling–mediated synchronization might be implicated in the high and low frequency oscillatory activities expressed by this type of thalamic neuron. (iii) Spontaneous intracellular Ca 2+ ([Ca 2+ ] i ) waves propagating among thalamic astrocytes are able to elicit large and long–lasting N –methyl–D–aspartate–mediated currents in TC neurons. The peculiar developmental profile within the first two postnatal weeks of these astrocytic [Ca 2+ ] i transients and the selective activation of these glutamate receptors point to a role for this astrocyte–to–neuron signalling mechanism in the topographic wiring of the thalamocortical loop. As some of these novel cellular and intracellular properties are not restricted to thalamic astrocytes and neurons, their significance may well apply to (patho)physiological functions of glial and neuronal elements in other brain areas.

Rapidly Deactivating AMPA Receptors Determine Excitatory Synaptic Transmission to Interneurons in the Nucleus Tractus Solitarius From Rat

1997 ◽  
Vol 78 (1) ◽  
pp. 82-91 ◽  
Author(s):  
Stefan Titz ◽  
Bernhard U. Keller
Keyword(s):  
Synaptic Transmission ◽  
Ampa Receptor ◽  
Decay Time ◽  
Ampa Receptors ◽  
Time Course ◽  
Nucleus Tractus Solitarius ◽  
Synaptic Cleft ◽  
Excitatory Synaptic Transmission ◽  
Membrane Properties ◽  
Time Constants

Titz, Stefan and Bernhard U. Keller. Rapidly deactivating AMPA receptors determine excitatory synaptic transmission to interneurons in the nucleus tractus solitarius from rat. J. Neurophysiol. 78: 82–91, 1997. Excitatory synaptic transmission was investigated in interneurons of the parvocellular nucleus tractus solitarius (pNTS) by performing patch-clamp experiments in thin slice preparations from rat brain stem. Stimulation of single afferent fibers evoked excitatory postsynaptic currents (EPSCs) mediated by glutamate receptors of the dl-α-amino-3-hydroxy-5-methylisoxazole-propionic acid (AMPA) and N-methyl-d-aspartate types. AMPA-receptor-mediated EPSCs displayed decay time constants of 3.5 ± 1.2 (SD) ms (13 cells), which were slow compared with EPSC decay time constants in neurons of the cerebellum or hippocampus. Slow EPSC decay was not explained by dendritic filtering, because the passive membrane properties of pNTS interneurons provided favorable voltage-clamp conditions. Also, the slowness of EPSC decay did not result from slow deactivation of AMPA receptors (0.7 ± 0.2 ms, 5 cells), which was investigated during rapid application of agonist to outside-out patches. Comparison of AMPA receptor kinetics with EPSC decay time constants suggested that the slow time course of EPSCs resulted from the prolonged presence of glutamate in the synaptic cleft.

Effects of oxygen tension on energetics of cultured vascular smooth muscle

2002 ◽  
Vol 283 (1) ◽  
pp. H110-H117 ◽  
Author(s):  
Anders Lindqvist ◽  
Karl Dreja ◽  
Karl Swärd ◽  
Per Hellstrand
Keyword(s):  
Chronic Hypoxia ◽  
Lactate Production ◽  
Synthesis Rate ◽  
Protein Synthesis Rate ◽  
Adrenergic Stimulation ◽  
Cellular Mechanisms ◽  
Mitochondrial Uncoupling ◽  
Vascular Abnormalities ◽  
Intracellular Ca ◽  
Glycogen Stores

Chronic hypoxia is a clinically important condition known to cause vascular abnormalities. To investigate the cellular mechanisms involved, we kept rings of a rat tail artery for 4 days in hypoxic culture (HC) or normoxic culture (NC) (Po 2 = 14 vs. 110 mmHg) and then measured contractility, oxygen consumption ( J o2 ), and lactate production ( J lac) in oxygenated medium. Compared with fresh rings, basal ATP turnover ( J ATP) was decreased in HC, but not in NC, with a shift from oxidative toward glycolytic metabolism. J o2 during mitochondrial uncoupling was reduced by HC but not by NC. Glycogen stores were increased 40-fold by HC and fourfold by NC. Maximum tension in response to norepinephrine and the Jo2 versus tension relationship ( J o2 vs. high K+ elicited force) were unaffected by either HC or NC. Force transients in response to caffeine were increased in HC, whereas intracellular Ca2+ wave activity during adrenergic stimulation was decreased. Protein synthesis rate was reduced by HC. The results show that long-term hypoxia depresses basal energy turnover, impairs mitochondrial capacity, and alters Ca2+homeostasis, but does not affect contractile energetics. These alterations may form a basis for vascular damage by chronic hypoxia.

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