scholarly journals Gating by Cyclic Gmp and Voltage in the α Subunit of the Cyclic Gmp–Gated Channel from Rod Photoreceptors

1999 ◽  
Vol 114 (4) ◽  
pp. 477-490 ◽  
Author(s):  
Klaus Benndorf ◽  
Rolf Koopmann ◽  
Elisabeth Eismann ◽  
U. Benjamin Kaupp
Keyword(s):  
Cyclic Gmp ◽  
Rate Constants ◽  
Time Course ◽  
Rod Photoreceptor ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Α Subunit ◽  
Outward Rectification ◽  
Voltage Dependent ◽  
Gated Channel

Gating by cGMP and voltage of the α subunit of the cGMP-gated channel from rod photoreceptor was examined with a patch-clamp technique. The channels were expressed in Xenopus oocytes. At low [cGMP] (<20 μM), the current displayed strong outward rectification. At low and high (700 μM) [cGMP], the channel activity was dominated by only one conductance level. Therefore, the outward rectification at low [cGMP] results solely from an increase in the open probability, Po. Kinetic analysis of single-channel openings revealed two exponential distributions. At low [cGMP], the larger Po at positive voltages with respect to negative voltages is caused by an increased frequency of openings in both components of the open-time distribution. In macroscopic currents, depolarizing voltage steps, starting from −100 mV, generated a time-dependent current that increased with the step size (activation). At low [cGMP] (20 μM), the degree of activation was large and the time course was slow, whereas at saturating [cGMP] (7 mM) the respective changes were small and fast. The dose–response relation at −100 mV was shifted to the right and saturated at significantly lower Po values with respect to that at +100 mV (0.77 vs. 0.96). Po was determined as function of the [cGMP] (at +100 and −100 mV) and voltage (at 20, 70, and 700 μM, and 7 mM cGMP). Both relations could be fitted with an allosteric state model consisting of four independent cGMP-binding reactions and one voltage-dependent allosteric opening reaction. At saturating [cGMP] (7 mM), the activation time course was monoexponential, which allowed us to determine the individual rate constants for the allosteric reaction. For the rapid rate constants of cGMP binding and unbinding, lower limits are determined. It is concluded that an allosteric model consisting of four independent cGMP-binding reactions and one voltage-dependent allosteric reaction, describes the cGMP- and voltage-dependent gating of cGMP-gated channels adequately.

scholarly journals Slo3 K+ Channels: Voltage and pH Dependence of Macroscopic Currents

2006 ◽  
Vol 128 (3) ◽  
pp. 317-336 ◽  
Author(s):  
Xue Zhang ◽  
Xuhui Zeng ◽  
Christopher J. Lingle
Keyword(s):  
Time Course ◽  
Voltage Dependence ◽  
Ph Dependence ◽  
K Channel ◽  
Open Probability ◽  
Cytosolic Ph ◽  
Α Subunit ◽  
K Channels ◽  
C Terminus ◽  
Voltage Dependent

The mouse Slo3 gene (KCNMA3) encodes a K+ channel that is regulated by changes in cytosolic pH. Like Slo1 subunits responsible for the Ca2+ and voltage-activated BK-type channel, the Slo3 α subunit contains a pore module with homology to voltage-gated K+ channels and also an extensive cytosolic C terminus thought to be responsible for ligand dependence. For the Slo3 K+ channel, increases in cytosolic pH promote channel activation, but very little is known about many fundamental properties of Slo3 currents. Here we define the dependence of macroscopic conductance on voltage and pH and, in particular, examine Slo3 conductance activated at negative potentials. Using this information, the ability of a Horrigan-Aldrich–type of general allosteric model to account for Slo3 gating is examined. Finally, the pH and voltage dependence of Slo3 activation and deactivation kinetics is reported. The results indicate that Slo3 differs from Slo1 in several important ways. The limiting conductance activated at the most positive potentials exhibits a pH-dependent maximum, suggesting differences in the limiting open probability at different pH. Furthermore, over a 600 mV range of voltages (−300 to +300 mV), Slo3 conductance shifts only about two to three orders of magnitude, and the limiting conductance at negative potentials is relatively voltage independent compared to Slo1. Within the context of the Horrigan-Aldrich model, these results indicate that the intrinsic voltage dependence (zL) of the Slo3 closed–open equilibrium and the coupling (D) between voltage sensor movement are less than in Slo1. The kinetic behavior of Slo3 currents also differs markedly from Slo1. Both activation and deactivation are best described by two exponential components, both of which are only weakly voltage dependent. Qualitatively, the properties of the two kinetic components in the activation time course suggest that increases in pH increase the fraction of more rapidly opening channels.

Cation fluxes and cation channels in outer segment membranes of bovine retinal rods: contamination by antibiotics applied to cattle?

1994 ◽  
Vol 72 (6) ◽  
pp. 650-658 ◽  
Author(s):  
Ricardo Lopez-Escalera ◽  
Robert J. French ◽  
Paul P. M. Schnetkamp
Keyword(s):  
Cyclic Gmp ◽  
Single Channel ◽  
Membrane Vesicles ◽  
Rod Photoreceptor ◽  
Channel Measurements ◽  
Open Probability ◽  
Retinal Rods ◽  
Gated Channel ◽  
Flux Measurements ◽  
Conductance States

Membrane vesicles were prepared from intact rod outer segments (ROSs) isolated from bovine retinas and were examined for the presence of cation-selective conductances. We performed macroscopic flux measurements in an ensemble of ROS membrane vesicles and single-channel measurements after fusion of ROS membrane vesicles with planar bilayer membranes. Two K+-permeable conductances were observed, the well-established cyclic GMP (cGMP) gated channel and an apparently new K+ channel with some unusual properties. Flux and single-channel data showed that the new conductance passed K+, Rb+, and Cs+ equally well but was much less permeable to Na+, Li+ and protons. Single-channel measurements revealed a linear current–voltage relationship and three unitary conductance states of 15, 11, and 8 pS, using symmetric 150 mM KCl solutions. Measured macroscopic K+ fluxes varied considerably among different preparations, suggesting some unknown regulation of the channel; the variability appeared to arise from variation in the channel's open probability, not the unit conductance or the number of channels present. The recorded single-channel events and the selectivity data are remarkably similar to those reported for antibiotic channel-forming ionophore gramicidin. We believe that the variability in both macroscopic permeability experiments and single-channel experiments may reflect a variable contamination with gramicidin applied to the animals as the topical antibiotic V-Sporin.Key words: cyclic GMP gated channel, ion channel, rod photoreceptor, gramicidin.

Voltage-dependent calcium channel current in isolated gallbladder smooth muscle cells of guinea pig

1993 ◽  
Vol 264 (6) ◽  
pp. G1066-G1076 ◽  
Author(s):  
T. Shimada
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Time Course ◽  
Reversal Potential ◽  
High Sensitivity ◽  
Muscle Cells ◽  
Patch Clamp Technique ◽  
Decay Component ◽  
Voltage Dependent

The voltage-dependent Ca2+ current was studied in enzymatically dispersed guinea pig gallbladder smooth muscle cells using the whole cell patch-clamp technique. Depolarizing voltage (V) steps induced an inward current (I) that was carried by Ca2+. The threshold potential was -40 to -30 mV, the maximal current was observed at +10 to +20 mV, and the reversal potential was around +80 mV. I-V curves obtained with holding potentials of -80 and -40 mV were not significantly different. This current had a high sensitivity to dihydropyridine drugs, and the Ba2+ or Sr2+ current was larger than the Ca2+ current. Activation was accelerated by increasing the membrane potential. In general, the time course of decay was well fitted by the sum of two exponentials, but consideration of a third (ultra-slow) decay component was also necessary when the current generated by a 2-s command pulse was analyzed. Superimposition of activation and inactivation curves showed the presence of a significant window current. Carbachol suppressed the Ca2+ current only when the pipette contained a low concentration of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. These results show that the L-type Ca2+ current is dominant in gallbladder smooth muscle cells and may contribute to excitation-contraction coupling.

Voltage-dependent kinetics of Na-Ca exchange current in Ca(2+)-loaded guinea pig heart cells

1993 ◽  
Vol 265 (5) ◽  
pp. C1258-C1265 ◽  
Author(s):  
K. Benndorf ◽  
C. Biskup ◽  
M. Friedrich
Keyword(s):  
Guinea Pig ◽  
Time Course ◽  
Exchange Current ◽  
Heart Cells ◽  
Current Component ◽  
Patch Clamp Technique ◽  
Initial Current ◽  
Current Increase ◽  
Voltage Dependent ◽  
Kinetics Of

Voltage-dependent properties of Na-Ca exchange current were revealed with the patch-clamp technique in Ca(2+)-overloaded guinea pig ventricular myocytes in the whole cell configuration. With the assumption that the transient inward current (Iti) is mediated by the Na-Ca exchanger, oscillations of internal Ca2+ concentration ([Ca2+]i) were used to investigate voltage-dependent kinetics of exchange current differences at two [Ca2+]i values. After Iti was elicited by clamping from -45 mV to basic pulses of +10 mV, pairs of equipotential short test pulses were applied during the basic pulse at both the phase of low [Ca2+]i (between two neighboring Iti values) and the phase of high [Ca2+]i (at the peak of Iti). The test pulses were short enough to leave the time course of Iti during the basic pulse approximately unchanged, which allowed study of the voltage dependence of the respective current differences without disturbing the underlying oscillation of [Ca2+]i. The current differences were inward at all potentials between -140 and +70 mV, started from an equal initial value, and obeyed characteristic voltage-dependent time courses: hyperpolarization to potentials negative to -70 mV caused an initial current increase, which was followed by a decay to very small amplitudes or zero with a decay time constant decreasing toward hyperpolarization e-fold per 45.6 mV. Depolarizing pulses caused a decay of the current differences to smaller levels. Respective current differences formed during a slowly decaying current component, following the Ca current spike, showed equal voltage-dependent properties. This indicates that the slowly decaying current component is preferentially also carried by the Na-Ca exchanger.(ABSTRACT TRUNCATED AT 250 WORDS)

Two types of voltage-dependent potassium channels in outer hair cells from the guinea pig cochlea

1999 ◽  
Vol 277 (5) ◽  
pp. C913-C925 ◽  
Author(s):  
Thierry van den Abbeele ◽  
Jacques Teulon ◽  
Patrice Tran Ba Huy
Keyword(s):  
Guinea Pig ◽  
Hair Cells ◽  
Basolateral Membrane ◽  
Outer Hair Cells ◽  
Catalytic Subunit ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Guinea Pig Cochlea ◽  
Voltage Dependent ◽  
Inside Out

Cell-attached and cell-free configurations of the patch-clamp technique were used to investigate the conductive properties and regulation of the major K+channels in the basolateral membrane of outer hair cells freshly isolated from the guinea pig cochlea. There were two major voltage-dependent K+ channels. A Ca2+-activated K+ channel with a high conductance (220 pS, P K/ P Na= 8) was found in almost 20% of the patches. The inside-out activity of the channel was increased by depolarizations above 0 mV and increasing the intracellular Ca2+concentration. External ATP or adenosine did not alter the cell-attached activity of the channel. The open probability of the excised channel remained stable for several minutes without rundown and was not altered by the catalytic subunit of protein kinase A (PKA) applied internally. The most frequent K+ channel had a low conductance and a small outward rectification in symmetrical K+ conditions (10 pS for inward currents and 20 pS for outward currents, P K/ P Na= 28). It was found significantly more frequently in cell-attached and inside-out patches when the pipette contained 100 μM acetylcholine. It was not sensitive to internal Ca2+, was inhibited by 4-aminopyridine, was activated by depolarization above −30 mV, and exhibited a rundown after excision. It also had a slow inactivation on ensemble-averaged sweeps in response to depolarizing pulses. The cell-attached activity of the channel was increased when adenosine was superfused outside the pipette. This effect also occurred with permeant analogs of cAMP and internally applied catalytic subunit of PKA. Both channels could control the cell membrane voltage of outer hair cells.

scholarly journals Potassium channel types in arterial chemoreceptor cells and their selective modulation by oxygen.

1992 ◽  
Vol 100 (3) ◽  
pp. 401-426 ◽  
Author(s):  
M D Ganfornina ◽  
J López-Barneo
Keyword(s):  
Time Course ◽  
Single Channel ◽  
K Channel ◽  
Open Probability ◽  
K Channels ◽  
Glomus Cells ◽  
Control Value ◽  
Voltage Dependent ◽  
K Current ◽  
Chemoreceptor Cells

Single K+ channel currents were recorded in excised membrane patches from dispersed chemoreceptor cells of the rabbit carotid body under conditions that abolish current flow through Na+ and Ca2+ channels. We have found three classes of voltage-gated K+ channels that differ in their single-channel conductance (gamma), dependence on internal Ca2+ (Ca2+i), and sensitivity to changes in O2 tension (PO2). Ca(2+)-activated K+ channels (KCa channels) with gamma approximately 210 pS in symmetrical K+ solutions were observed when [Ca2+]i was greater than 0.1 microM. Small conductance channels with gamma = 16 pS were not affected by [Ca2+]i and they exhibited slow activation and inactivation time courses. In these two channel types open probability (P(open)) was unaffected when exposed to normoxic (PO2 = 140 mmHg) or hypoxic (PO2 approximately 5-10 mmHg) external solutions. A third channel type (referred to as KO2 channel), having an intermediate gamma(approximately 40 pS), was the most frequently recorded. KO2 channels are steeply voltage dependent and not affected by [Ca2+]i, they inactivate almost completely in less than 500 ms, and their P(open) reversibly decreases upon exposure to low PO2. The effect of low PO2 is voltage dependent, being more pronounced at moderately depolarized voltages. At 0 mV, for example, P(open) diminishes to approximately 40% of the control value. The time course of ensemble current averages of KO2 channels is remarkably similar to that of the O2-sensitive K+ current. In addition, ensemble average and macroscopic K+ currents are affected similarly by low PO2. These observations strongly suggest that KO2 channels are the main contributors to the macroscopic K+ current of glomus cells. The reversible inhibition of KO2 channel activity by low PO2 does not desensitize and is not related to the presence of F-, ATP, and GTP-gamma-S at the internal face of the membrane. These results indicate that KO2 channels confer upon glomus cells their unique chemoreceptor properties and that the O2-K+ channel interaction occurs either directly or through an O2 sensor intrinsic to the plasma membrane closely associated with the channel molecule.

Voltage-dependent K+ current in capillary endothelial cells isolated from guinea pig heart

1999 ◽  
Vol 277 (1) ◽  
pp. H119-H127 ◽  
Author(s):  
Michael Dittrich ◽  
Jürgen Daut
Keyword(s):  
Guinea Pig ◽  
Time Course ◽  
Single Channel ◽  
Patch Clamp Technique ◽  
Guinea Pig Heart ◽  
Potential Oscillations ◽  
Time To Peak ◽  
Voltage Dependent ◽  
Capillary Endothelial Cells ◽  
Single Channel Currents

Capillary fragments were isolated from guinea pig hearts, and their electrical properties were studied using the perforated-patch and cell-attached mode of the patch-clamp technique. A voltage-dependent K+ current was discovered that was activated at potentials positive to −20 mV and showed a sigmoid rising phase. For depolarizing voltage steps from −128 to +52 mV, the time to peak was 71 ± 5 ms (mean ± SE) and the amplitude of the current was 3.7 ± 0.5 pA/pF in the presence of 5 mM external K+. The time course of inactivation was exponential with a time constant of 7.2 ± 0.5 s at +52 mV. The current was blocked by tetraethylammonium (inhibitory constant ∼3 mM) but was not affected by charybdotoxin (1 μM) or apamin (1 μM). In the cell-attached mode, depolarization-activated single-channel currents were found that inactivated completely within 30 s; the single-channel conductance was 12.3 ± 2.4 pS. The depolarization-activated K+current described here may play a role in membrane potential oscillations of the endothelium.

scholarly journals Primary structure and functional expression from complementary DNA of the rod photoreceptor cyclic GMP-gated channel

Nature ◽  
1989 ◽  
Vol 342 (6251) ◽  
pp. 762-766 ◽  
Author(s):  
U. Benjamin Kaupp ◽  
Tetsuhiro Niidome ◽  
Tsutomu Tanabe ◽  
Shoichi Terada ◽  
Wolfgang Bönigk ◽  
...  
Keyword(s):  
Cyclic Gmp ◽  
Primary Structure ◽  
Functional Expression ◽  
Rod Photoreceptor ◽  
Complementary Dna ◽  
Gated Channel

L- and T-type voltage-gated Ca2+ currents in adrenal medulla endothelial cells

1999 ◽  
Vol 276 (4) ◽  
pp. H1313-H1322 ◽  
Author(s):  
Raul Vinet ◽  
Fernando F. Vargas
Keyword(s):  
Endothelial Cells ◽  
Adrenal Medulla ◽  
Time Course ◽  
Maximum Amplitude ◽  
Secretory Cells ◽  
Activation Time ◽  
Patch Clamp Technique ◽  
Activation Threshold ◽  
Current Voltage ◽  
Voltage Dependent

We investigated voltage-dependent Ca2+ channels of bovine adrenal medulla endothelial cells with the whole cell version of the patch-clamp technique. Depolarization elicited an inward current that was carried by Ca2+ and was composed of a transient (T) current, present in all the cells tested, and a sustained (L) current, present in 65% of them. We separated these currents and measured their individual kinetic and gating properties. The activation threshold for T current was approximately −50 mV, and its maximum amplitude was −49.8 ± 4.8 pA (means ± SE, n = 19) at 0 mV. The time constant was 10.2 ± 1.5 ms ( n= 4) for activation and 18.4 ± 2.8 ms ( n = 4) for inactivation. The L current activated at −40 mV, and it reached a plateau at −20.1 ± 2.3 pA ( n = 6). Its activation time course was a single exponential with an activation time contant of 26.8 ± 2.3 ms ( n = 4). Current-voltage curves, kinetics, gating, response to BAY K 8644, nifedipine, amiloride, and different selectivity for Ba2+ and Ca2+ indicated that the underlying channels for the observed currents are only of the T- and L-types that resemble those of the endocrine secretory cells.

Cl- current in IMCD cells activated by hypotonicity: time course, ATP dependence, and inhibitors

1996 ◽  
Vol 271 (3) ◽  
pp. F552-F559 ◽  
Author(s):  
K. A. Volk ◽  
C. Zhang ◽  
R. F. Husted ◽  
J. B. Stokes
Keyword(s):  
Time Course ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Primary Cultures ◽  
Renal Medulla ◽  
Cell Types ◽  
Disulfonic Acid ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent

The hypertonic environment of the renal medulla can change rapidly according to the state of hydration of the animal. We used primary cultures of rat inner medullary collecting duct (IMCD) cells to investigate the characteristics of Cl- currents activated by an acute reduction in osmolarity (ICl(osm)). Using the whole cell patch-clamp technique, we identified an outwardly rectifying current that decayed slowly at strongly depolarizing voltages. The onset of ICl(osm) began 6.7 min after the fall in bath osmolarity, a delay longer than reported in other cell types. Hypotonicity did not induce an increase in intracellular Ca2+ concentration, and activation of ICl(osm) did not require the presence of Ca2+. Intracellular ATP was needed to evoke ICl(osm) when the hypotonic stimulus was modest (50 mosmol/l or less) but was not necessary when the stimulus was stronger (100 mosmol/ l). ICl(osm) was inhibited by 5-nitro-2-(3-phenylpropylamino)benzoic acid but not by tamoxifen or glibenclamide. 4,4'-Diisothiocyanostilbene-2,2'-disulfonic acid produced a voltage-dependent block. Acute reduction in osmolarity using cells grown on filters did not induce a Cl- secretory current. The ICl(osm) of IMCD cells appears to be on the basolateral membrane and displays some unique features.

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