scholarly journals Two kinds of calcium channels in canine atrial cells. Differences in kinetics, selectivity, and pharmacology.

1985 ◽  
Vol 86 (1) ◽  
pp. 1-30 ◽  
Author(s):  
B P Bean
Keyword(s):  
Single Channel ◽  
Cardiac Cells ◽  
Fluctuation Analysis ◽  
Ca Channel ◽  
Ca Channels ◽  
Ca Current ◽  
Channel Current ◽  
Atrial Cells ◽  
Cell Recording ◽  
Ca Channel Current

Currents through Ca channels were recorded in single canine atrial cells using whole-cell recording with patch pipettes. Two components of Ca channel current could be distinguished. One ("Ifast") was present only if cells were held at negative potentials, was most prominent for relatively small depolarizations, and inactivated within tens of milliseconds. The other ("Islow"), corresponding to the Ca current previously reported in single cardiac cells, persisted even at relatively positive holding potentials, required stronger depolarizations for maximal current, and inactivated much more slowly. Both currents were unaffected by tetrodotoxin and both were reduced by Co. Ifast had the same size and kinetics when Ca was exchanged for Ba, while Islow was bigger and slower with Ba as the charge carrier. In isotonic BaCl2, fluctuation analysis showed that Ifast had a smaller single channel current than Islow. Islow was much more sensitive to block by nitrendipine than was Ifast; also, Islow, but not Ifast, was increased by the dihydropyridine drug BAY K8644. Isoproterenol produced large increases in Islow but had no effect on Ifast.

scholarly journals The dihydropyridine-sensitive calcium channel subtype in cone photoreceptors.

1996 ◽  
Vol 107 (5) ◽  
pp. 621-630 ◽  
Author(s):  
M F Wilkinson ◽  
S Barnes
Keyword(s):  
Channel Blocker ◽  
Channel Blockers ◽  
Ca Channel ◽  
Ca Channels ◽  
Cone Photoreceptors ◽  
Channel Current ◽  
Ca Channel Blockers ◽  
Voltage Dependent ◽  
P Type ◽  
Ca Channel Current

High-voltage activated Ca channels in tiger salamander cone photoreceptors were studied with nystatin-permeabilized patch recordings in 3 mM Ca2+ and 10 mM Ba2+. The majority of Ca channel current was dihydropyridine sensitive, suggesting a preponderance of L-type Ca channels. However, voltage-dependent, incomplete block (maximum 60%) by nifedipine (0.1-100 microM) was evident in recordings of cones in tissue slice. In isolated cones, where the block was more potent, nifedipine (0.1-10 microM) or nisoldipine (0.5-5 microM) still failed to eliminate completely the Ca channel current. Nisoldipine was equally effective in blocking Ca channel current elicited in the presence of 10 mM Ba2+ (76% block) or 3 mM Ca2+ (88% block). 15% of the Ba2+ current was reversibly blocked by omega-conotoxin GVIA (1 microM). After enhancement with 1 microM Bay K 8644, omega-conotoxin GVIA blocked a greater proportion (22%) of Ba2+ current than in control. After achieving partial block of the Ba2+ current with nifedipine, concomitant application of omega-conotoxin GVIA produced no further block. The P-type Ca channel blocker, omega-agatoxin IVA (200 nM), had variable and insignificant effects. The current persisting in the presence of these blockers could be eliminated with Cd2+ (100 microM). These results indicate that photoreceptors express an L-type Ca channel having a distinguishing pharmacological profile similar to the alpha 1D Ca channel subtype. The presence of additional Ca channel subtypes, resistant to the widely used L-, N-, and P-type Ca channel blockers, cannot, however, be ruled out.

Calcium Channel Activation Facilitated by Nitric Oxide in Retinal Ganglion Cells

2000 ◽  
Vol 83 (1) ◽  
pp. 198-206 ◽  
Author(s):  
Kazuyuki Hirooka ◽  
Dmitri E. Kourennyi ◽  
Steven Barnes
Keyword(s):  
Protein Kinase ◽  
Retinal Ganglion Cells ◽  
Ganglion Cells ◽  
Peptide Inhibitor ◽  
Ca Channel ◽  
Retinal Ganglion ◽  
Ca Channels ◽  
Channel Current ◽  
Channel Currents ◽  
Ca Channel Current

We investigated the modulation of voltage-gated Ca channels by nitric oxide (NO) in isolated salamander retinal ganglion cells with the goals of determining the type of Ca channel affected and the signaling pathway by which modulation might occur. The NO donors, S-nitroso- N-acetyl-penicillamine (SNAP, 1 mM) and S-nitroso-cysteine (1 mM) induced modest increases in the amplitude of Ca channel currents recorded with ruptured- and permeabilized-patch techniques by causing a subpopulation of the Ca channels to activate at more negative potentials. The Ca channel antagonists ω-conotoxin GVIA and nisoldipine each reduced the Ca channel current partially, but only ω-conotoxin GVIA blocked the enhancement by SNAP. The SNAP-induced increase was blocked by oxadiazolo-quinoxaline (50 μM), suggesting that the NO generated by SNAP acts via a soluble guanylyl cyclase to raise levels of cGMP. The membrane-permeant cGMP analog 8-(4-chlorophenylthio) guanosine cyclic monophosphate also enhanced Ca channel currents and 8-bromo guanosine cyclic monophosphate (1 mM) occluded enhancement by SNAP. Consistent with these results, isobutyl-methyl-xanthine (IBMX, 10 μM), which can raise cGMP levels by inhibiting phosphodiesterase activity, increased Ca channel current by the same amount as SNAP and occluded subsequent enhancement by SNAP. Neither IBMX, the cGMP analogs, nor SNAP itself, led to activation of cGMP-gated channels. N-[2-(methylamino)ethyl]−5-isoquinoline-sulfonamide (2 μM), a broad spectrum inhibitor of protein kinase activity, KT5823 (1 μM), a specific protein kinase G (PKG) inhibitor, and a peptide inhibitor of PKG (200 μM) blocked SNAP enhancement, as did 5′-adenylylimidophosphate (1.5 mM), a nonhydrolyzable ATP analog that prevents protein phosphorylation. A peptide inhibitor of protein kinase A (10 nM) did not block the facilitory effects of SNAP. Okadaic acid (1 μM), a phosphatase inhibitor, had no effect by itself but increased the enhancement of Ca channel current by SNAP. These results suggest that NO modulates retinal ganglion cell N-type Ca channels by facilitating their voltage-dependent activation via a mechanism involving guanylyl cyclase/PKG-dependent phosphorylation. This effect could fine-tune neural integration in ganglion cells or play a role in ganglion cell disease by modulating intracellular calcium signaling.

Carbenoxolone Inhibition of Voltage-Gated Ca Channels and Synaptic Transmission in the Retina

2004 ◽  
Vol 92 (2) ◽  
pp. 1252-1256 ◽  
Author(s):  
John P. Vessey ◽  
Melanie R. Lalonde ◽  
Hossein A. Mizan ◽  
Nicole C. Welch ◽  
Melanie E. M. Kelly ◽  
...  
Keyword(s):  
Synaptic Transmission ◽  
Horizontal Cell ◽  
Transfer Model ◽  
Ca Channel ◽  
Ca Channels ◽  
Inhibitory Effects ◽  
Voltage Gated ◽  
Channel Current ◽  
Inhibitory Feedback ◽  
Ca Channel Current

We show that carbenoxolone, a drug used to block hemichannels in the retina to test the ephaptic model of horizontal cell inhibitory feedback, has strong inhibitory effects on voltage-gated Ca channels. Carbenoxolone (100 μM) reduced photoreceptor-to-horizontal cell synaptic transmission by 92%. Applied to patch-clamped, isolated cone photoreceptors, carbenoxolone inhibited Ca channels with an EC50 of 48 μM. At 100 μM, it reduced cone Ca channel current by 37%, reduced depolarization-evoked [Ca2+] signals in fluo-4 loaded retinal slices by 57% and inhibited Ca channels in Müller cells by 52%. A synaptic transfer model suggests that the degree of block of Ca channels accounts for the reduction in synaptic transmission. These results suggest broad inhibitory actions for carbenoxolone in the retina that must be considered when interpreting its effects on inhibitory feedback.

scholarly journals Permeation and interaction of divalent cations in calcium channels of snail neurons.

1985 ◽  
Vol 85 (4) ◽  
pp. 491-518 ◽  
Author(s):  
L Byerly ◽  
P B Chase ◽  
J R Stimers
Keyword(s):  
Mole Fraction ◽  
Surface Potential ◽  
Divalent Cation ◽  
Lymnaea Stagnalis ◽  
Divalent Cations ◽  
Ca Channel ◽  
Ca Current ◽  
Channel Current ◽  
Gating Mechanism ◽  
Ca Channel Current

We have studied the current-carrying ability and blocking action of various divalent cations in the Ca channel of Lymnaea stagnalis neurons. Changing the concentration or species of the permeant divalent cation shifts the voltage dependence of activation of the Ca channel current in a manner that is consistent with the action of the divalent cation on an external surface potential. Increasing the concentration of the permeant cation from 1 to 30 mM produces a twofold increase in the maximum Ca current and a fourfold increase in the maximum Ba current; the maximum Ba current is twice the size of the maximum Ca current for 10 mM bulk concentration. Correcting for the changing surface potential seen by the gating mechanism, the current-concentration relation is almost linear for Ba2+, and shows only moderate saturation for Ca2+; also, Ca2+, Ba2+, and Sr2+ are found to pass through the channel almost equally well. These conclusions are obtained for either of two assumptions: that the mouth of the channel sees (a) all or (b) none of the surface potential seen by the gating mechanism. Cd2+ blocks Lymnaea and Helix Ca channels at concentrations 200 times smaller than those required for Co2+ or Ni2+. Ca2+ competes with Cd2+ for the blocking site; Ba2+ binds less strongly than Ca2+ to this site. Mixtures of Ca2+ and Ba2+ produce an anomalous mole fraction effect on the Ca channel current. After correction for the changing surface potential (using either assumption), the anomalous mole fraction effect is even more prominent, which suggests that Ba2+ blocks Ca current more than Ca2+ blocks Ba current.

scholarly journals Inositol (1,4,5)-trisphosphate (InsP3)-gated Ca channels from cerebellum: conduction properties for divalent cations and regulation by intraluminal calcium.

1994 ◽  
Vol 104 (5) ◽  
pp. 821-856 ◽  
Author(s):  
I Bezprozvanny ◽  
B E Ehrlich
Keyword(s):  
Ryanodine Receptor ◽  
Single Channel ◽  
Divalent Cations ◽  
Reversal Potential ◽  
Ca Channel ◽  
Ca Channels ◽  
Ca Current ◽  
Open Probability ◽  
Inositol 1,4,5 Trisphosphate ◽  
Conduction Properties

The conduction properties of inositol (1,4,5)-trisphosphate (InsP3)-gated calcium (Ca) channels (InsP3R) from canine cerebellum for divalent cations and the regulation of the channels by intraluminal Ca were studied using channels reconstituted into planar lipid bilayers. Analysis of single-channel recordings performed with different divalent cations present at 55 mM on the trans (intraluminal) side of the membrane revealed that the current amplitude at 0 mV and the single-channel slope conductance fell in the sequence: Ba (2.2 pA, 85 pS) > Sr (2.0 pA, 77 pS) > Ca (1.4 pA, 53 pS) > Mg (1.1 pA, 42 pS). The mean open time of the InsP3R recorded with Ca (2.9 ms) was significantly shorter than with other divalent cations (approximately 5.5 ms). The "anomalous mole fraction effect" was not observed in mixtures of divalent cations (Mg and Ba), suggesting that these channels are single-ion pores. Measurements of InsP3R activity at different intraluminal Ca levels demonstrated that Ca in the submillimolar range did not potentiate channel activity, and that very high levels of intraluminal Ca (> or = 10 mM) decreased channel open probability 5-10-fold. When InsP3R were measured with Ba as a current carrier in the presence of 110 mM cis potassium, a PBa/PK of 6.3 was estimated from the extrapolated value for the reversal potential. When the unitary current through the InsP3R at 0 mV was measured as a function of the permeant ion (Ba) concentration, the half-maximal current occurred at 10 mM trans Ba. The following conclusions are drawn from these data: (a) the conduction properties of InsP3R are similar to the properties of the ryanodine receptor, another intracellular Ca channel, and differ dramatically from the properties of voltage-gated Ca channels of the plasma membrane. (b) The estimated size of the Ca current through the InsP3R under physiological conditions is 0.5 pA, approximately four times less than the Ca current through the ryanodine receptor. (c) The potentiation of InsP3R by intraluminal Ca in the submillimolar range remains controversial. (d) A quantitative model that explains the inhibitory effects of high trans Ca on InsP3R activity was developed and the kinetic parameters of InsP3R gating were determined.

Enhanced single-channel K+ current in arterial membranes from genetically hypertensive rats

1993 ◽  
Vol 264 (5) ◽  
pp. H1337-H1345 ◽  
Author(s):  
S. K. England ◽  
T. A. Wooldridge ◽  
W. J. Stekiel ◽  
N. J. Rusch
Keyword(s):  
Single Channel ◽  
Muscle Cells ◽  
Ca Channel ◽  
Ca Channels ◽  
Hypertensive Rats ◽  
Genetically Hypertensive Rats ◽  
K Current ◽  
Inside Out ◽  
Genetically Hypertensive ◽  
Arterial Muscle

Arterial smooth muscle from hypertensive rats shows an increased membrane permeability to K+ that depends on Ca2+ influx. To define the mechanism of this membrane alteration, we tested the hypothesis that Ca(2+)-dependent K+ current (IK(Ca)) is increased in arterial muscle membranes from genetically hypertensive rats. Single-channel K+ currents measured in cell-attached and inside-out aortic membrane patches from spontaneously hypertensive rats (SHR) were compared with those from normotensive Wistar-Kyoto rats (WKY). Inside-out patches from both rat strains showed a predominant 225 pS, Ca(2+)- and voltage-dependent K+ channel in symmetrical 145 mM KCl solutions, which was blocked by tetraethylammonium [concentration for half-maximal block (IC50) < or = 0.3 mM]. In cell-attached patches of aortic muscle cells bathed in physiological salt solution, this channel [IK(Ca) channel] showed a fivefold higher open-state probability (NPo) in SHR as compared with WKY. This increased NPo of SHR IK(Ca) channels in membranes of intact aortic muscle cells was not correlated with an altered membrane potential in current-clamped SHR myocytes or with changes in cytosolic free Ca2+ concentration in fura-2-loaded aortic muscle cells. However, inside-out aortic membrane patches from SHR showed more detected IK(Ca) channels per patch, a higher IK(Ca) channel NPo, and a greater total patch current than their WKY counterparts. Further analysis revealed a greater Ca2+ sensitivity of SHR than WKY IK(Ca) channels. These results suggest that IK(Ca) channel function is altered in isolated membrane patches of arterial muscle from genetically hypertensive rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential responses of Ca-activated K channels to bradykinin in sensory neurons and F-11 cells

1992 ◽  
Vol 262 (2) ◽  
pp. C453-C460 ◽  
Author(s):  
K. Naruse ◽  
D. S. McGehee ◽  
G. S. Oxford
Keyword(s):  
Intracellular Signaling ◽  
Single Channel ◽  
Outward Current ◽  
Transient Outward Current ◽  
Ca Channel ◽  
Ca Channels ◽  
Drg Neurons ◽  
Open Probability ◽  
K Channels ◽  
Drg Neuron

The nonapeptide bradykinin (BK) excites a subset of dorsal root ganglion (DRG) neurons with putative nociceptive functions by stimulating an inward cation current. In addition, BK stimulates various intracellular signaling pathways including an elevation of intracellular Ca2+. In a DRG neuron x neuroblastoma hybrid cell (F-11), BK stimulates similar increases in intracellular [Ca2+] and inward current but also elicits a large transient outward current through Ca(2+)-activated K channels. We have investigated the mechanisms underlying differential expression of outward current responses in the two cell types at the single channel level. Although K(Ca) channel activity appears in inside-out patches from both cells exposed to Ca2+, BK applied to the extrapatch membrane of cell-attached patches activates K(Ca) channels in F-11 but not DRG neurons. Whereas single K(Ca) channels are quantitatively similar in terms of conductance, voltage-dependence, and sensitivity to tetraethylammonium, they differ in sensitivity to intracellular Ca2+. Channel activation in both cells requires at least four Ca2+ ions, but half-maximal activation occurs at slightly higher [Ca2+] for DRG neurons. The shift in the Ca2+ dose-response curve combined with the steep [Ca2+] dependence of channel open probability makes it less likely that a BK-induced rise in internal [Ca2+] induced will trigger a transient outward current and resultant hyperpolarization in a DRG neuron.

scholarly journals Cardiac calcium channels in planar lipid bilayers. L-type channels and calcium-permeable channels open at negative membrane potentials.

1988 ◽  
Vol 92 (1) ◽  
pp. 27-54 ◽  
Author(s):  
R L Rosenberg ◽  
P Hess ◽  
R W Tsien
Keyword(s):  
Lipid Bilayers ◽  
Single Channel ◽  
Detailed Comparison ◽  
Membrane Potentials ◽  
Bay K 8644 ◽  
Ca Channel ◽  
Ca Channels ◽  
Channel Activity ◽  
Intact Cells ◽  
Free System

Planar lipid bilayer recordings were used to study Ca channels from bovine cardiac sarcolemmal membranes. Ca channel activity was recorded in the absence of nucleotides or soluble enzymes, over a range of membrane potentials and ionic conditions that cannot be achieved in intact cells. The dihydropyridine-sensitive L-type Ca channel, studied in the presence of Bay K 8644, was identified by a detailed comparison of its properties in artificial membranes and in intact cells. L-type Ca channels in bilayers showed voltage dependence of channel activation and inactivation, open and closed times, and single-channel conductances in Ba2+ and Ca2+ very similar to those found in cell-attached patch recordings. Open channels were blocked by micromolar concentrations of external Cd2+. In this cell-free system, channel activity tended to decrease during the course of an experiment, reminiscent of Ca2+ channel "rundown" in whole-cell and excised-patch recordings. A purely voltage-dependent component of inactivation was observed in the absence of Ca2+ stores or changes in intracellular Ca2+. Millimolar internal Ca2+ reduced unitary Ba2+ influx but did not greatly increase the rate or extent of inactivation or the rate of channel rundown. In symmetrical Ba2+ solutions, unitary conductance saturated as the Ba2+ concentration was increased up to 500 mM. The bilayer recordings also revealed activity of a novel Ca2+-permeable channel, termed "B-type" because it may contribute a steady background current at negative membrane potentials, which is distinct from L-type or T-type Ca channels previously reported. Unlike L-type channels, B-type channels have a small unitary Ba2+ conductance (7 pS), but do not discriminate between Ba2+ and Ca2+, show no obvious sensitivity to Bay K 8644, and do not run down. Unlike either L- or T-type channels, B-type channels did not require a depolarization for activation and displayed mean open times of greater than 100 ms.

scholarly journals Endogenous Protease Activation of ENaC

2005 ◽  
Vol 126 (4) ◽  
pp. 339-352 ◽  
Author(s):  
Adedotun Adebamiro ◽  
Yi Cheng ◽  
John P. Johnson ◽  
Robert J. Bridges
Keyword(s):  
Single Channel ◽  
Apical Membrane ◽  
Open Channels ◽  
Na Channel ◽  
Fluctuation Analysis ◽  
Protease Inhibition ◽  
Open Probability ◽  
Channel Current ◽  
Single Channel Current ◽  
Channel Properties

Endogenous serine proteases have been reported to control the reabsorption of Na+ by kidney- and lung-derived epithelial cells via stimulation of electrogenic Na+ transport mediated by the epithelial Na+ channel (ENaC). In this study we investigated the effects of aprotinin on ENaC single channel properties using transepithelial fluctuation analysis in the amphibian kidney epithelium, A6. Aprotinin caused a time- and concentration-dependent inhibition (84 ± 10.5%) in the amiloride-sensitive sodium transport (INa) with a time constant of 18 min and half maximal inhibition constant of 1 μM. Analysis of amiloride analogue blocker–induced fluctuations in INa showed linear rate–concentration plots with identical blocker on and off rates in control and aprotinin-inhibited conditions. Verification of open-block kinetics allowed for the use of a pulse protocol method (Helman, S.I., X. Liu, K. Baldwin, B.L. Blazer-Yost, and W.J. Els. 1998. Am. J. Physiol. 274:C947–C957) to study the same cells under different conditions as well as the reversibility of the aprotinin effect on single channel properties. Aprotinin caused reversible changes in all three single channel properties but only the change in the number of open channels was consistent with the inhibition of INa. A 50% decrease in INa was accompanied by 50% increases in the single channel current and open probability but an 80% decrease in the number of open channels. Washout of aprotinin led to a time-dependent restoration of INa as well as the single channel properties to the control, pre-aprotinin, values. We conclude that protease regulation of INa is mediated by changes in the number of open channels in the apical membrane. The increase in the single channel current caused by protease inhibition can be explained by a hyperpolarization of the apical membrane potential as active Na+ channels are retrieved. The paradoxical increase in channel open probability caused by protease inhibition will require further investigation but does suggest a potential compensatory regulatory mechanism to maintain INa at some minimal threshold value.

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