Inhibitory interactions between 5-HT3 and P2X channels in submucosal neurons

2002 ◽  
Vol 283 (6) ◽  
pp. G1238-G1248 ◽  
Author(s):  
Carlos Barajas-López ◽  
Luis M. Montaño ◽  
Rosa Espinosa-Luna
Keyword(s):  
P2x Receptors ◽  
Disulfonic Acid ◽  
Pharmacological Properties ◽  
Pipette Solution ◽  
Outward Currents ◽  
C 23 ◽  
Cell Recording ◽  
Inward Currents ◽  
Inhibitory Interactions ◽  
Subtype 3

Inhibitory interactions between 5-HT subtype 3 (5-HT3) and P2X receptors were characterized using whole cell recording techniques. Currents induced by 5-HT ( I 5-HT) and ATP ( I ATP) were blocked by tropisetron (or ondansetron) and pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid, respectively. Currents induced by 5-HT + ATP ( I 5-HT+ATP) were only as large as the current induced by the most effective transmitter, revealing current occlusion. Occlusion was observed at membrane potentials of −60 and 0 mV (for inward currents), but it was not present at +40 mV (for outward currents). Kinetic and pharmacological properties of I 5-HT+ATP indicate that they are carried through 5-HT3 and P2X channels. Current occlusion occurred as fast as activation of I 5-HT and I ATP, was still present in the absence of Ca2+ or Mg2+, after adding staurosporine, genistein, K-252a, or N-ethylmaleimide to the pipette solution, after substituting ATP with ∝,β-methylene ATP or GTP with GTP-γ-S in the pipette, and was observed at 35°C, 23°C, and 8°C. These results are in agreement with a model that considers that 5-HT3 and P2X channels are in functional clusters and that these channels might directly inhibit each other.

scholarly journals Exocytotic release of ATP and activation of P2X receptors in dissociated guinea pig stellate neurons

2006 ◽  
Vol 291 (5) ◽  
pp. C1062-C1071 ◽  
Author(s):  
John D. Tompkins ◽  
Rodney L. Parsons
Keyword(s):  
Guinea Pig ◽  
Pyridoxal Phosphate ◽  
Purinergic Receptor ◽  
P2x Receptors ◽  
Disulfonic Acid ◽  
Inward Current ◽  
Protein Receptor ◽  
Vesicular Release ◽  
Cell Recording ◽  
Inward Currents

Activation of P2X receptors by a Ca2+- and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein-dependent release of ATP was measured using patch-clamp recordings from dissociated guinea pig stellate neurons. Asynchronous transient inward currents (ASTICs) were activated by depolarization or treatment with the Ca2+ ionophore ionomycin (1.5 and 3 μM). During superfusion with a HEPES-buffered salt solution containing 2.5 mM Ca2+, depolarizing voltage steps (−60 to 0 mV, 500 ms) evoked ASTICs on the decaying phase of a larger, transient inward current. Equimolar substitution of Ba2+ for Ca2+ augmented the postdepolarization frequency of ASTICs, while eliminating the larger transient current. Perfusion with an ionomycin-containing solution elicited a sustained activation of ASTICs, allowing quantitative analysis over a range of holding potentials. Under these conditions, increasing extracellular [Ca2+] to 5 mM increased ASTIC frequency, whereas no events were observed following replacement of Ca2+ with Mg2+, demonstrating a Ca2+ requirement. ASTICs were Na+ dependent, inwardly rectifying, and reversed near 0 mV. Treatment with the nonselective purinergic receptor antagonist pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS) (10 μM) blocked all events under both conditions, whereas the ganglionic nicotinic antagonist hexamethonium (100 μM and 1 mM) had no effect. PPADS also blocked the macroscopic inward current evoked by exogenously applied ATP (300 μM). The presence of botulinum neurotoxin E (BoNT/E) in the whole-cell recording electrode significantly attenuated the ionomycin-induced ASTIC activity, whereas phorbol ester treatment potentiated this activity. These results suggest that ASTICs are mediated by vesicular release of ATP and activation of P2X receptors.

ATP is not required for anion current activated by cell swelling in multidrug-resistant lung cancer cells

1994 ◽  
Vol 267 (3) ◽  
pp. C688-C699 ◽  
Author(s):  
J. D. Jirsch ◽  
D. W. Loe ◽  
S. P. Cole ◽  
R. G. Deeley ◽  
D. Fedida
Keyword(s):  
Multidrug Resistant ◽  
Cell Swelling ◽  
Disulfonic Acid ◽  
Open Probability ◽  
Pipette Solution ◽  
Current Cell ◽  
Absolute Requirement ◽  
Cell Recording ◽  
Volume Response ◽  
Cytoskeletal Elements

During whole cell recording with 4 mM ATP and 0.1 mM GTP in the pipette, outwardly rectifying Cl- currents (155 +/- 20.5 pA/pF) were repetitively activated on reduction of bath solution osmolarity from 290 mosM (control) to 210 mosM. These currents were sensitive to 0.1-1 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. Omission of ATP from the pipette solution reduced the current magnitude to 42.7 +/- 9.5 pA/pF and prevented repetitive activation. More hyposmotic solutions (160 mosM) usually elicited current repetitively despite an ATP-free pipette solution. In cells depleted of ATP (to < 5% of control) by preincubation with 2-deoxyglucose (10 mM) and rotenone (100 nM), hyposmotic solutions failed to activate significant current. Cell volume increased to 230 +/- 18% of control (19.1 +/- 1.2 microns) in 210 mosM bath (normal cells) but only to 114 +/- 13% of control in ATP-depleted cells exposed to 160 mosM solution. This failure of ATP-depleted cells to swell in hypotonic external solutions was reversed by overnight pretreatment with cytochalasin D (2 micrograms/ml; n = 6) but not by colchicine (250 microM; n = 8). In outside-out patches of membrane dialyzed with zero ATP and excised from swollen cells, we observed sustained activation of a 53-pS outwardly rectifying channel (chord conductance, +100 mV; open probability approximately 1.0). In cell-attached patches from normal and ATP-depleted cells, we activated similar channels by suction. ATP does not appear to be an absolute requirement for the activation of this Cl- channel in H69AR cells but may be essential for the normal volume response and channel activation mediated through cytoskeletal elements within cells.

scholarly journals A Store-operated Calcium Channel in Drosophila S2 Cells

2004 ◽  
Vol 123 (2) ◽  
pp. 167-182 ◽  
Author(s):  
Andriy V. Yeromin ◽  
Jack Roos ◽  
Kenneth A. Stauderman ◽  
Michael D. Cahalan
Keyword(s):  
Mammalian Cells ◽  
Divalent Cations ◽  
Noise Analysis ◽  
Reversal Potential ◽  
S2 Cells ◽  
Pipette Solution ◽  
Crac Channels ◽  
Drosophila S2 Cells ◽  
Cell Recording ◽  
Inward Currents

Using whole-cell recording in Drosophila S2 cells, we characterized a Ca2+-selective current that is activated by depletion of intracellular Ca2+ stores. Passive store depletion with a Ca2+-free pipette solution containing 12 mM BAPTA activated an inwardly rectifying Ca2+ current with a reversal potential &gt;60 mV. Inward currents developed with a delay and reached a maximum of 20–50 pA at −110 mV. This current doubled in amplitude upon increasing external Ca2+ from 2 to 20 mM and was not affected by substitution of choline for Na+. A pipette solution containing ∼300 nM free Ca2+ and 10 mM EGTA prevented spontaneous activation, but Ca2+ current activated promptly upon application of ionomycin or thapsigargin, or during dialysis with IP3. Isotonic substitution of 20 mM Ca2+ by test divalent cations revealed a selectivity sequence of Ba2+ &gt; Sr2+ &gt; Ca2+ &gt;&gt; Mg2+. Ba2+ and Sr2+ currents inactivated within seconds of exposure to zero-Ca2+ solution at a holding potential of 10 mV. Inactivation of Ba2+ and Sr2+ currents showed recovery during strong hyperpolarizing pulses. Noise analysis provided an estimate of unitary conductance values in 20 mM Ca2+ and Ba2+ of 36 and 420 fS, respectively. Upon removal of all external divalent ions, a transient monovalent current exhibited strong selectivity for Na+ over Cs+. The Ca2+ current was completely and reversibly blocked by Gd3+, with an IC50 value of ∼50 nM, and was also blocked by 20 μM SKF 96365 and by 20 μM 2-APB. At concentrations between 5 and 14 μM, application of 2-APB increased the magnitude of Ca2+ currents. We conclude that S2 cells express store-operated Ca2+ channels with many of the same biophysical characteristics as CRAC channels in mammalian cells.

KATP channel conductance of descending vasa recta pericytes

2005 ◽  
Vol 289 (6) ◽  
pp. F1235-F1245 ◽  
Author(s):  
Chunhua Cao ◽  
Whaseon Lee-Kwon ◽  
Erik P. Silldorff ◽  
Thomas L. Pallone
Keyword(s):  
Significant Contribution ◽  
Receptor Subtype ◽  
Ang Ii ◽  
Channel Conductance ◽  
Endothelin 1 ◽  
Outward Currents ◽  
Vasa Recta ◽  
Cell Recording ◽  
Inward Currents ◽  
Perforated Patch Clamp

Using nystatin-perforated patch-clamp and whole cell recording, we tested the hypothesis that KATP channels contribute to resting conductance of rat descending vasa recta (DVR) pericytes and are modulated by vasoconstrictors. The KATP blocker glybenclamide (Glb; 10 μM) depolarized pericytes and inhibited outward currents of cells held at −40 mV. KATP openers pinacidil (Pnc; 10 μM) and P-1075 (1 μM) hyperpolarized pericytes and transiently augmented outward currents. All effects of Pnc and P-1075 were fully reversed by Glb. Inward currents of pericytes held at −60 mV in symmetrical 140 mM K+ were markedly augmented by Pnc and fully reversed by Glb. Ramp depolarizations in symmetrical K+, performed in Pnc and Pnc + Glb, yielded a Pnc-induced, Glb-sensitive KATP difference current that lacked rectification and reversed at 0 mV. Immunostaining identified both KIR6.1, KIR6.2 inward rectifier subunits and sulfonurea receptor subtype 2B. ANG II (1 and 10 nM) and endothelin-1 (10 nM) but not vasopressin (100 nM) significantly lowered holding current at −40 mV and abolished Pnc-stimulated outward currents. We conclude that DVR pericytes express KATP channels that make a significant contribution to basal K+ conductance and are inhibited by ANG II and endothelin-1.

Muscarinic and Nicotinic ACh Receptor Activation Differentially Mobilize Ca2+ in Rat Intracardiac Ganglion Neurons

2003 ◽  
Vol 90 (3) ◽  
pp. 1956-1964 ◽  
Author(s):  
Friederike Beker ◽  
Martin Weber ◽  
Rainer H. A. Fink ◽  
David J. Adams
Keyword(s):  
Receptor Activation ◽  
Neonatal Rat ◽  
Induced Response ◽  
Induced Responses ◽  
Outward Currents ◽  
Cell Recording ◽  
Intracellular Ca ◽  
Inward Currents ◽  
Ganglion Neurons ◽  
Intracardiac Neurons

The origin of intracellular Ca2+ concentration ([Ca2+]i) transients stimulated by nicotinic (nAChR) and muscarinic (mAChR) receptor activation was investigated in fura-2-loaded neonatal rat intracardiac neurons. ACh evoked [Ca2+]i increases that were reduced to ∼60% of control in the presence of either atropine (1 μM) or mecamylamine (3 μM) and to <20% in the presence of both antagonists. Removal of external Ca2+ reduced ACh-induced responses to 58% of control, which was unchanged in the presence of mecamylamine but reduced to 5% of control by atropine. The nAChR-induced [Ca2+]i response was reduced to 50% by 10 μM ryanodine, whereas the mAChR-induced response was unaffected by ryanodine, suggesting that Ca2+ release from ryanodine-sensitive Ca2+ stores may only contribute to the nAChR-induced [Ca2+]i responses. Perforated-patch whole cell recording at –60 mV shows that the rise in [Ca2+]i is concomitant with slow outward currents on mAChR activation and with rapid inward currents after nAChR activation. In conclusion, different signaling pathways mediate the rise in [Ca2+]i and membrane currents evoked by ACh binding to nicotinic and muscarinic receptors in rat intracardiac neurons.

Nitric oxide mediates outward potassium currents in opossum esophageal circular smooth muscle

1996 ◽  
Vol 270 (6) ◽  
pp. G932-G938 ◽  
Author(s):  
J. Jury ◽  
K. R. Boev ◽  
E. E. Daniel
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Circular Muscle ◽  
Outward Current ◽  
Equilibrium Potential ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Circular Smooth Muscle ◽  
Outward Currents

Single smooth muscle cells from the opossum body circular muscle were isolated and whole cell currents were characterized by the whole cell patch-clamp technique. When the cells were held at -50 mV and depolarized to 70 mV in 20-mV increments, initial small inactivating inward currents were evoked (-30 to 30 mV) followed by larger sustained outward currents. Depolarization from a holding potential of -90 mV evoked an initial fast inactivating outward current sensitive to 4-aminopyridine but not to high levels of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). The outward currents reversed near K+ equilibrium potential and were abolished when KCl was replaced by CsCl in the pipette solution. The sustained outward current was inhibited by quinine and cesium. High EGTA in the pipette solution reduced but did not abolish the sustained outward currents, suggesting that both Ca(2+)-dependent and -independent currents were evoked. The nitric oxide (NO)-releasing agents Sin-1 and sodium nitroprusside increased outward K+ currents. High levels of EGTA in the pipette solution abolished the increase in outward current induced by Sin-1. The presence of cyclopiazonic acid, an inhibitor of the sarcoplasmic reticulum (SR) Ca2+ pump, blocked the effects of NO-releasing agents. We conclude that NO release activates K+ outward currents in opossum esophagus circular muscle, which may depend on Ca2+ release from the SR stores.

Lysophosphatidic acid, serum, and hyposmolarity activate Cl- currents in corneal keratocytes

1995 ◽  
Vol 269 (6) ◽  
pp. C1385-C1393 ◽  
Author(s):  
M. A. Watsky
Keyword(s):  
Lysophosphatidic Acid ◽  
Reversal Potential ◽  
Flufenamic Acid ◽  
Disulfonic Acid ◽  
Hypotonic Stress ◽  
Outward Currents ◽  
Fetal Bovine ◽  
Patch Technique ◽  
Corneal Keratocytes ◽  
In Cells

The influence of serum, lysophosphatidic acid (LPA), and hyposmotic stress on the ion channel activity of normal and cryo-injured rabbit corneal keratocytes was investigated. Whole cell currents were examined using the amphotericin perforated-patch technique. In cells from wounded corneas, fetal bovine serum activated large, holding voltage-insensitive, fast-activating, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)-, flufenamic acid-, and 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB)-blockable outward currents showing inactivation at depolarized voltages. LPA activated identical currents, also only in cells from wounded corneas. Blocker and reversal potential experiments characterized the current as a Cl- currents (Icl). Lysophosphatidylcholine (10 microM) failed to activate the current. An identical current was activated by hyposmotic stimulation in cells from control and wounded corneas. Hyposmotic stimulation also activated Icl in cells from wounded corneas that were unresponsive to LPA. We conclude that serum, LPA, and hypotonic stress activate Icl in keratocytes from wounded corneas. We also conclude that LPA is a serum factor that can activate Icl and that hyposmotic activation may work through a signaling pathway separate from that of LPA.

ATP-sensitive K(+)-selective channels of inner medullary collecting duct cells

1994 ◽  
Vol 267 (3) ◽  
pp. F489-F496 ◽  
Author(s):  
S. C. Sansom ◽  
T. Mougouris ◽  
S. Ono ◽  
T. D. DuBose
Keyword(s):  
Single Channel ◽  
Collecting Duct ◽  
K Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Inner Medullary Collecting Duct ◽  
Outward Currents ◽  
Inward Currents ◽  
Excised Patches ◽  
Medullary Collecting Duct

The inner medullary collecting duct (IMCD) in vivo has the capacity to either secrete or reabsorb K+. However, a selective K+ conductance has not been described previously in the IMCD. In the present study, the patch-clamp method was used to determine the presence and properties of K(+)-selective channels in the apical membrane of the inner medullary collecting duct cell line, mIMCD-3. Two types of K(+)-selective channels were observed in both cell-attached and excised patches. The most predominant K+ channel, a smaller conductance K+ channel (SK), was present in cell-attached patches with 140 mM KCl (high bath K+) but not with 135 mM NaCl plus 5 mM KCl (low bath K+) in the bathing solution. The single-channel conductance of SK was 36 pS with inward currents and 29 pS with outward currents in symmetrical 140 mM KCl. SK was insensitive to both voltage and Ca2+. However, SK was inhibited significantly by millimolar concentrations of ATP in excised patches. A second K(+)-selective channel [a larger K+ channel (BK)] displayed a single-channel conductance equal to 132 pS with inward currents and 90 pS with outward currents in symmetrical 140 mM KCl solutions. BK was intermittently activated in excised inside-out patches by Mg(2+)-ATP in concentrations from 1 to 5 mM. With complete removal of Mg2+, BK was insensitive to ATP. BK was also insensitive to potential and Ca2+ and was observed in cell-attached patches with 140 mM KCl in the bath solution. Both channels were blocked reversibly by 1 mM Ba2+ from the intracellular surface but not by external Ba2+.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Effect of FMRFamide on voltage-dependent currents in identified centrifugal neurons of the optic lobe of the cuttlefish, Sepia officinalis

2020 ◽  
Author(s):  
Abdesslam Chrachri
Keyword(s):  
Visual Processing ◽  
Calcium Current ◽  
Optic Lobe ◽  
Low Voltage ◽  
Sodium Current ◽  
Calcium Currents ◽  
Delayed Rectifier ◽  
Outward Currents ◽  
Voltage Dependent ◽  
Inward Currents

AbstractWhole-cell patch-clamp recordings from identified centrifugal neurons of the optic lobe in a slice preparation allowed the characterization of five voltage-dependent currents; two outward and three inward currents. The outward currents were; the 4-aminopyridine-sensitive transient potassium or A-current (IA), the TEA-sensitive sustained current or delayed rectifier (IK). The inward currents were; the tetrodotoxin-sensitive transient current or sodium current (INa). The second is the cobalt- and cadmium-sensitive sustained current which is enhanced by barium and blocked by the dihydropyridine antagonist, nifedipine suggesting that it could be the L-type calcium current (ICaL). Finally, another transient inward current, also carried by calcium, but unlike the L-type, this current is activated at more negative potentials and resembles the low-voltage-activated or T-type calcium current (ICaT) of other preparations.Application of the neuropeptide FMRFamide caused a significant attenuation to the peak amplitude of both sodium and sustained calcium currents without any apparent effect on the transient calcium current. Furthermore, FMRFamide also caused a reduction of both outward currents in these centrifugal neurons. The fact that FMRFamide reduced the magnitude of four of five characterized currents could suggest that this neuropeptide may act as a strong inhibitory agent on these neurons.SummaryFMRFamide modulate the ionic currents in identified centrifugal neurons in the optic lobe of cuttlefish: thus, FMRFamide could play a key role in visual processing of these animals.

scholarly journals THE VARIATION OF ELECTRICAL RESISTANCE WITH APPLIED POTENTIAL

1930 ◽  
Vol 14 (1) ◽  
pp. 139-162 ◽  
Author(s):  
L. R. Blinks
Keyword(s):  
Electrical Resistance ◽  
Sea Water ◽  
Single Layer ◽  
Applied Potential ◽  
Intact Cells ◽  
Outward Currents ◽  
Toxic Agents ◽  
Constant State ◽  
Inward Currents ◽  
Apparent Resistance

Electrical resistance and polarization were measured during the passage of direct current across a single layer of protoplasm in the cells of Valonia ventricosa impaled upon capillaries. These were correlated with five stages of the P.D. existing naturally across the protoplasm, as follows: 1. A stage of shock after impalement, when the P.D. drops from 5 mv. to zero and then slowly recovers. There is very little effective resistance in the protoplasm, and polarization is slight. 2. The stage of recovery and normal P.D., with values from 8 to 25 mv. (inside positive). The average is 15 mv. At first there is little or no polarization when small potentials are applied in either direction across the protoplasm, nor when very large currents pass outward (from sap to sea water). But when the positive current passes inward there is a sudden response at a critical applied potential ranging from 0.5 to 2.0 volts. The resistance then apparently rises as much as 10,000 ohms in some cases, and the rise occurs more quickly in succeeding applications after the first. When the potential is removed there is a back E.M.F. displayed. Later there is also an effect of such inward currents which persists into the first succeeding outward flow, causing a brief polarization at the first application of the reverse potential. Still later this polarization occurs at every exposure, and at increasingly lower values of applied potentials. Finally there is a "constant" state reached in which the polarization occurs with currents of either direction, and the apparent resistance is nearly uniform over a considerable range of applied potential. 3. A state of increased P.D.; to 100 mv. (inside positive) in artificial sap; and to 35 or 40 mv. in dilute sea water or 0.6 M MgSO4. The polarization response and apparent resistance are at first about as in sea water, but later decrease. 4. A reversed P.D., to 50 mv. (outside positive) produced by a variety of causes, especially by dilute sea water, and also by large flows of current in either direction. This stage is temporary and the cells promptly recover from it. While it persists the polarization appears to be much greater to outward currents than to inward. This can largely be ascribed to the reduction of the reversed P.D. 5. Disappearance of P.D. caused by death, and various toxic agents. The resistance and polarization of the protoplasm are negligible. The back E.M.F. of polarization is shown to account largely for the apparent resistance of the protoplasm. Its calculation from the observed resistance rises gives values up to 150 mv. in the early stages of recovery, and later values of 50 to 75 mv. in the "constant" state. These are compared with the back E.M.F. similarly calculated from the apparent resistance of intact cells. The electrical capacitance of the protoplasm is shown by the time curves to be of the order of 1 microfarad per cm.2 of surface.

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