scholarly journals Release from the cone ribbon synapse under bright light conditions can be controlled by the opening of only a few Ca2+ channels

2011 ◽  
Vol 106 (6) ◽  
pp. 2922-2935 ◽  
Author(s):  
Theodore M. Bartoletti ◽  
Skyler L. Jackman ◽  
Norbert Babai ◽  
Aaron J. Mercer ◽  
Richard H. Kramer ◽  
...  
Keyword(s):  
Diffusion Barrier ◽  
Single Channel ◽  
Bright Light ◽  
Vesicle Fusion ◽  
Channel Number ◽  
Release Mechanism ◽  
Ribbon Synapse ◽  
Channel Current ◽  
Replicated Observations ◽  
Vesicle Pool

Light hyperpolarizes cone photoreceptors, causing synaptic voltage-gated Ca2+ channels to open infrequently. To understand neurotransmission under these conditions, we determined the number of L-type Ca2+ channel openings necessary for vesicle fusion at the cone ribbon synapse. Ca2+ currents ( ICa) were activated in voltage-clamped cones, and excitatory postsynaptic currents (EPSCs) were recorded from horizontal cells in the salamander retina slice preparation. Ca2+ channel number and single-channel current amplitude were calculated by mean-variance analysis of ICa. Two different comparisons—one comparing average numbers of release events to average ICa amplitude and the other involving deconvolution of both EPSCs and simultaneously recorded cone ICa—suggested that fewer than three Ca2+ channel openings accompanied fusion of each vesicle at the peak of release during the first few milliseconds of stimulation. Opening fewer Ca2+ channels did not enhance fusion efficiency, suggesting that few unnecessary channel openings occurred during strong depolarization. We simulated release at the cone synapse, using empirically determined synaptic dimensions, vesicle pool size, Ca2+ dependence of release, Ca2+ channel number, and Ca2+ channel properties. The model replicated observations when a barrier was added to slow Ca2+ diffusion. Consistent with the presence of a diffusion barrier, dialyzing cones with diffusible Ca2+ buffers did not affect release efficiency. The tight clustering of Ca2+ channels, along with a high-Ca2+ affinity release mechanism and diffusion barrier, promotes a linear coupling between Ca2+ influx and vesicle fusion. This may improve detection of small light decrements when cones are hyperpolarized by bright light.

Permeation and gating properties of a cloned renal K+ channel

1995 ◽  
Vol 268 (2) ◽  
pp. C389-C401 ◽  
Author(s):  
S. Chepilko ◽  
H. Zhou ◽  
H. Sackin ◽  
L. G. Palmer
Keyword(s):  
Single Channel ◽  
Reversal Potential ◽  
Cation Binding ◽  
K Channel ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Membrane Hyperpolarization ◽  
Channel Current ◽  
Inward Currents ◽  
Kinetics Of

The renal K+ channel (ROMK2) was expressed in Xenopus oocytes, and the patch-clamp technique was used to assess its conducting and gating properties. In cell-attached patches with 110 mM K+ in the bath and pipette, the reversal potential was near zero and the inward conductance (36 pS) was larger than the outward conductance (17 pS). In excised inside-out patches the channels showed rectification in the presence of 5 mM Mg2+ on the cytoplasmic side but not in Mg(2+)-free solution. Inward currents were also observed when K+ was replaced in the pipette by Rb+, NH4+, or thallium (Tl+). The reversal potentials under these conditions yielded a selectivity sequence of Tl+ > K+ > Rb+ > NH4+. On the other hand, the slope conductances for inward current gave a selectivity sequence of K+ = NH4+ > Tl+ > Rb+. The differences in the two sequences can be explained by the presence of cation binding sites within the channel, which interact with Rb+ and Tl+ more strongly and with NH4+ less strongly than with K+. Two other ions, Ba2+ and Cs+, blocked the channel from the outside. The effect of Ba2+ (1 mM) was to reduce the open probability of the channels, whereas Cs+ (10 mM) reduced the apparent single-channel current. The effects of both blockers are enhanced by membrane hyperpolarization. The kinetics of the channel were also studied in cell-attached patches. With K+ in the pipette the distribution of open times could be described by a single exponential (tau 0 = 25 ms), whereas two exponentials (tau 1 = 1 ms, tau 2 = 30 ms) were required to describe the closed-time distribution. Hyperpolarization of the oocyte membrane decreased the open probability and tau 0, and increased tau 1, tau 2, and the number of long closures. The presence of Tl+ in the pipette significantly altered the kinetics, reducing tau 0 and eliminating the long-lived closures. These results suggest that the gating of the channel may depend on the nature of the ion in the pore.

Active NaCl absorption across split lamellae of posterior gills of the chinese crab Eriocheir sinensis: stimulation by eyestalk extract

2000 ◽  
Vol 203 (8) ◽  
pp. 1373-1381 ◽  
Author(s):  
H. Onken ◽  
A. Schobel ◽  
J. Kraft ◽  
M. Putzenlechner
Keyword(s):  
Electromotive Force ◽  
Single Channel ◽  
Short Circuit ◽  
Noise Analysis ◽  
Eriocheir Sinensis ◽  
Dialysis Tubing ◽  
Channel Current ◽  
Dose Dependent ◽  
Cl Conductance ◽  
Stimulation Of

Split lamellae of the posterior gills of freshwater-adapted Chinese crabs (Eriocheir sinensis) were mounted in a modified Ussing-type chamber, and active and electrogenic absorption of Na(+) and Cl(−) were measured as positive (I(Na)) or negative (I(Cl)) short-circuit currents. Haemolymph-side addition of eyestalk extract stimulated I(Cl) by increasing both the transcellular Cl(−) conductance and the electromotive force for Cl(−) absorption. The effect was dose-dependent. Boiling the eyestalk extract did not change its effectiveness. The stimulating factor passed through dialysis tubing, indicating that it has a molecular mass of less than 2 kDa. R(p)cAMPS, a blocker of protein kinase A, reduced the stimulated I(Cl). Eyestalk extract stimulated I(Na) by increasing the transcellular Na(+) conductance at constant electromotive force. Amiloride-induced current-noise analysis revealed that stimulation of I(Na) was accompanied by an increase in the apparent number of open apical Na(+) channels at a slightly reduced single-channel current. In addition to the electrophysiological experiments, whole gills were perfused in the presence and in the absence of putative transport stimulators, and the specific activities of the V-ATPase and the Na(+)/K(+)-ATPase were measured. Eyestalk extract, theophylline or dibutyryl-cyclic AMP stimulated the activity of the V-ATPase, whereas the activity of the Na(+)/K(+)-ATPase was unaffected. The simultaneous presence of R(p)cAMPS prevented the stimulation of V-ATPase by eyestalk extract or theophylline.

Effect of Prozac on whole cell ionic currents in lens and corneal epithelia

1995 ◽  
Vol 269 (1) ◽  
pp. C250-C256 ◽  
Author(s):  
J. L. Rae ◽  
A. Rich ◽  
A. C. Zamudio ◽  
O. A. Candia
Keyword(s):  
Potassium Channels ◽  
Single Channel ◽  
Ionic Currents ◽  
Current Amplitude ◽  
Human Lens ◽  
Delayed Rectifier ◽  
Open Probability ◽  
Channel Current ◽  
Single Channel Current ◽  
Ionic Conductances

Prozac (fluoxetine), a compound used therapeutically in humans to combat depression, has substantial effects on ionic conductances in rabbit corneal epithelial cells and in cultured human lens epithelium. In corneal epithelium, it reduces the current due to the large-conductance potassium channels that dominate this preparation. Its effects seem largely to decrease the open probability while leaving the single-channel current amplitude unaltered. In cultured human epithelium, currents from calcium-activated potassium channels and inward rectifiers are unaffected by Prozac. Delayed-rectifier potassium currents are reduced by Prozac in a complicated way that involves both gating and single-channel current amplitude. Fast tetrodotoxin-blockable sodium currents are also decreased by Prozac in this preparation. For all of these ion conductance effects, Prozac concentrations of 10(-5) to 10(-4) M are required. Whereas these levels are 10- to 100-fold higher than the plasma levels achieved in therapeutic use in humans, they are comparable to or less than levels needed for many other blockers of the ionic conductances studied here.

Properties and distribution of single voltage-gated calcium channels in adult hippocampal neurons

1990 ◽  
Vol 64 (1) ◽  
pp. 91-104 ◽  
Author(s):  
R. E. Fisher ◽  
R. Gray ◽  
D. Johnston
Keyword(s):  
Guinea Pig ◽  
Calcium Channels ◽  
Single Channel ◽  
Cell Types ◽  
Voltage Gated ◽  
Channel Current ◽  
Single Channel Current ◽  
Voltage Gated Calcium Channels ◽  
Large Conductance Channel ◽  
Small Conductance

1. The properties of single voltage-gated calcium channels were investigated in acutely exposed CA3 and CA1 pyramidal neurons and granule cells of area dentata in the adult guinea pig hippocampal formation. 2. Guinea pig hippocampal slices were prepared in a conventional manner, then treated with proteolytic enzymes and gently shaken to expose the somata of the three cell types studied. Standard patch-clamp techniques were used to record current flow through calcium channels in cell-attached membrane patches with isotonic barium as the charge carrier. 3. Single-channel current amplitudes were measured at different membrane potentials. Single-channel current-voltage plots were constructed and single-channel slope conductances were found to fall into three classes. These were (approximately) 8, 14, and 25 pS, and were observed in all three cell types. 4. The three groups of channels differed from each other in voltage dependence of activation: from a holding potential of -80, the small-conductance channel began to activate at about -40 to -30 mV, the medium-conductance channel at about -20 mV, and the large-conductance channel at approximately 0 mV. 5. Ensemble averages of single-channel currents during voltage steps revealed differences in voltage-dependent inactivation. The small-conductance channel inactivated completely within approximately 50 ms during steps from -80 to -10 mV or more positive. Steps to less positive potentials resulted in less inactivation. The medium-conductance channel displayed variable inactivation during steps from -80 to 0 mV. Inactivation of this channel during a 160-ms step ranged from virtually zero to approximately 100%. The large-conductance channel displayed no significant inactivation during steps as long as 400 ms. 6. The large-conductance channel was strikingly affected by the dihydropyridine agonist Bay K8644 (0.5-2.0 microM), resulting in a high probability of channel opening, prolonged openings, and an apparent increase in the number of channels available for activation. The medium and small-conductance channels were not noticeably affected by the drug. 7. The large-conductance channel could be induced to open at very negative membrane potentials by holding the patch for several seconds at 20 or 30 mV and stepping to -30 or -40 mV. This process was enhanced by Bay K8644, resulting in prolonged openings at potentials as negative as -100 mV.(ABSTRACT TRUNCATED AT 400 WORDS)

Time-dependent expression of ryanodine receptors in sea urchin eggs, zygotes and early embryos

Zygote ◽  
2021 ◽  
pp. 1-4
Author(s):  
G. Percivale ◽  
C. Angelini ◽  
C. Falugi ◽  
C. Picco ◽  
G. Prestipino
Keyword(s):  
Single Channel ◽  
Ryanodine Receptors ◽  
Paracentrotus Lividus ◽  
Current Fluctuations ◽  
Calcium Dependent ◽  
Channel Current ◽  
Single Channel Current ◽  
Electrophysiological Recordings ◽  
Early Embryos ◽  
Zygote Stage

Summary In this work, the presence of calcium-dependent calcium channels and their receptors (RyR) has been investigated in Paracentrotus lividus eggs and early embryos, from unfertilized egg to four-blastomere stages. Electrophysiological recordings of RyR single-channel current fluctuations showed that RyRs are functional during the first developmental events with a maximum at zygote stage, c. 40 min after fertilization, corresponding to the first cleavage. The nature of vertebrate-like RyRs active at this stage was established by specific activation/blockade experiments.

A rapidly activating delayed rectifier K+ channel in rabbit sinoatrial node cells

1995 ◽  
Vol 269 (2) ◽  
pp. H443-H452 ◽  
Author(s):  
H. Ito ◽  
K. Ono
Keyword(s):  
Steady State ◽  
Sinoatrial Node ◽  
Single Channel ◽  
Ensemble Average ◽  
Negative Slope ◽  
Delayed Rectifier ◽  
Channel Current ◽  
Single Channel Current ◽  
Sinoatrial Node Cells ◽  
Rabbit Sinoatrial Node

The single-channel current of the delayed rectifier K+ current (IK) was recorded in rabbit sinoatrial node cells. In the cell-attached patch, depolarization from -70 mV to potentials more positive than -50 mV activated the IK channel while repolarization deactivated it. The single-channel conductance was 7.8 pS for the outward current and 10.8 pS for the inward current (n = 6). The steady-state open probability (NPo) was maximum at around -30 mV and markedly decreased at more positive potentials. On repolarization from positive potentials, the channel was initially closed and then rapidly opened. The ensemble average showed an initial rise to a peak followed by the deactivation time course. Because the channel events were completely blocked by E-4031, the drug-sensitive component was examined in the whole cell current. The steady-state current-voltage relation of the drug-sensitive current showed a marked negative slope at potentials more positive than -10 mV. Upon repolarization, the drug-sensitive current initially increased (removal of inactivation) to the peak of the outward tail current, which was in agreement with the ensemble average of the single-channel current. We conclude that IK in the sinoatrial node cells is largely composed of the rapidly activating IK (IK,r) channels and that the inward rectification of IK,r, which is more marked than had been assumed in previous studies, is due to the decrease in NPo.

Brefeldin A inhibition of apical Na+ channels in epithelia

1996 ◽  
Vol 270 (1) ◽  
pp. C138-C147 ◽  
Author(s):  
R. S. Fisher ◽  
F. G. Grillo ◽  
S. Sariban-Sohraby
Keyword(s):  
Cultured Cells ◽  
Single Channel ◽  
Noise Analysis ◽  
Basolateral Membrane ◽  
Brefeldin A ◽  
Na Channel ◽  
Na Channels ◽  
Na Transport ◽  
Channel Current ◽  
Vacuolar System

Brefeldin A (BFA) is used to probe trafficking of proteins through the central vacuolar system (CVS) in a variety of cells. Transepithelial Na+ transport by high-resistance epithelia, such as A6 cultured cells, is inhibited by BFA. Apical Na+ channels, as well as basolateral pumps and K+ channels, are complex proteins that probably traverse the CVS for routing to the plasma membrane. BFA (5 micrograms/ml) decreases transepithelial Na+ current near zero and increases resistance reversibly after 4 h. Longer exposures are toxic. When tissues were treated for 20 h with 0.2 microgram/ml BFA, Na+ transport also was reversibly inhibited. Using noise analysis, we found that BFA drastically reduced apical Na+ channel density. The increase in single channel current was consistent with cell hyperpolarization. After apical permeabilization with nystatin, changes in transepithelial current reflect changes in basolateral membrane transport. Transport at this membrane was inhibited by ouabain and cycloheximide, but not by BFA. After BFA, aldosterone was ineffective, suggesting that an intact CVS is required for stimulation by this hormone. Thus BFA inhibition of Na+ transport is localized at the apical membrane. Implications for channel turnover as a mechanism for regulating the Na+ transport rate are discussed.

scholarly journals Stoichiometry of Recombinant N-Methyl-d-Aspartate Receptor Channels Inferred from Single-channel Current Patterns

1997 ◽  
Vol 110 (5) ◽  
pp. 485-502 ◽  
Author(s):  
Louis S. Premkumar ◽  
Anthony Auerbach
Keyword(s):  
Single Channel ◽  
Wild Type ◽  
Current Pattern ◽  
Channel Current ◽  
Single Channel Current ◽  
Aspartate Receptor ◽  
Single Channel Currents ◽  
Nr2 Subunits ◽  
Channel Currents ◽  
Straightforward Interpretation

Single-channel currents were recorded from mouse NR1-NR2B (ζ-ε2) receptors containing mixtures of wild-type and mutant subunits expressed in Xenopus oocytes. Mutant subunits had an asparagine-to-glutamine (N-to-Q) mutation at the N0 site of the M2 segment (NR1:598, NR2B:589). Receptors with pure N or Q NR1 and NR2 subunits generated single-channel currents with distinctive current patterns. Based on main and sublevel amplitudes, occupancy probabilities, and lifetimes, four patterns of current were identified, corresponding to receptors with the following subunit compositions (NR1/NR2): N/N, N/Q, Q/N, and Q/Q. Only one current pattern was apparent for each composition. When a mixture of N and Q NR2 subunits was coexpressed with pure mutant NR1 subunits, three single-channel current patterns were apparent. One pattern was the same as Q/Q receptors and another was the same as Q/N receptors. The third, novel pattern presumably arose from hybrid receptors having both N and Q NR2 subunits. When a mixture of N and Q NR1 subunits was coexpressed with pure mutant NR2 subunits, six single-channel current patterns were apparent. One pattern was the same as Q/Q receptors and another was the same as N/Q receptors. The four novel patterns presumably arose from hybrid receptors having both N and Q NR1 subunits. The relative frequency of NR1 hybrid receptor current patterns depended on the relative amounts of Q and N subunits that were injected into the oocytes. The number of hybrid receptor patterns suggests that there are two NR2 subunits per receptor and is consistent with either three or five NR1 subunits per receptor, depending on whether or not the order of mutant and wild-type subunits influences the current pattern. When considered in relation to other studies, the most straightforward interpretation of the results is that N-methyl-d-aspartate receptors are pentamers composed of three NR1 and two NR2 subunits.

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