scholarly journals Long-Term and Immediate Effect of Testosterone on Single T-Type Calcium Channel in Neonatal Rat Cardiomyocytes

Endocrinology ◽  
2006 ◽  
Vol 147 (11) ◽  
pp. 5160-5169 ◽  
Author(s):  
Guido Michels ◽  
Fikret Er ◽  
Michael Eicks ◽  
Stefan Herzig ◽  
Uta C. Hoppe
Keyword(s):  
Calcium Channels ◽  
Single Channel ◽  
Heart Diseases ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Channel Activity ◽  
Open Probability ◽  
Rat Cardiomyocytes ◽  
Single Channel Activity

In the cardiovascular system, T-type calcium channels play an important role for the intracellular calcium homeostasis and spontaneous pacemaker activity and are involved in the progression of structural heart diseases. Androgens influence the cardiovascular physiology and pathophysiology. However, their effect on native T-type calcium currents (ICa,T) remains unclear. To test the chronic effect of testosterone on the cardiac ICa,T, cultured neonatal rat ventricular cardiomyocytes were treated with testosterone (1 nm-10 μm) for 24–30 h. Current measurements were performed after testosterone washout to exclude any acute testosterone effects. Testosterone (100 nm) pretreatment significantly increased whole-cell ICa,T density from 1.26 ± 0.48 pA/pF (n = 8) to 5.06 ± 1.75 pA/pF (n = 7; P < 0.05) and accelerated beating rate. This was attributed to both increased expression levels of the pore-forming subunits Cav3.1 and Cav3.2 and increased T-type single-channel activity. On single-channel level, the increase of the ensemble average current by testosterone vs. time-matched controls was due to an increased availability (58.1 ± 4.2 vs. 21.5 ± 4.0%, P < 0.01) and open probability (2.78 ± 0.29 vs. 0.85 ± 0.23%, P < 0.01). Cotreatment with the selective testosterone receptor antagonist flutamide (10 μm) prevented these chronic testosterone-induced effects. Conversely, acute application of testosterone (10 μm) decreased T-type single-channel activity in testosterone pretreated cells by reducing the open probability (0.78 ± 0.13 vs. 2.91 ± 0.38%, P < 0.01), availability (23.6 ± 3.3 vs. 57.6 ± 4.5%, P < 0.01), and peak current (−20 ± 4 vs. −58 ± 4 fA, P < 0.01). Flutamide (10 μm) did not abolish the testosterone-induced acute block of T-type calcium channels. Our results indicate that long-term testosterone treatment increases, whereas acute testosterone decreases neonatal rat T-type calcium currents. These effects seem to be mediated by a genomic chronic stimulation and a nongenomic acute inhibitory action.

scholarly journals MicroRNA-204 Is Necessary for Aldosterone-Stimulated T-Type Calcium Channel Expression in Cardiomyocytes

2018 ◽  
Vol 19 (10) ◽  
pp. 2941 ◽  
Author(s):  
Riko Koyama ◽  
Tiphaine Mannic ◽  
Jumpei Ito ◽  
Laurence Amar ◽  
Maria-Christina Zennaro ◽  
...  
Keyword(s):  
Calcium Channels ◽  
Molecular Mechanisms ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Messenger Rnas ◽  
Screening Assay ◽  
Rat Cardiomyocytes ◽  
Ventricular Cardiomyocytes ◽  
Channel Expression ◽  
Beating Frequency

Activation of the mineralocorticoid receptor (MR) in the heart is considered to be a cardiovascular risk factor. MR activation leads to heart hypertrophy and arrhythmia. In ventricular cardiomyocytes, aldosterone induces a profound remodeling of ion channel expression, in particular, an increase in the expression and activity of T-type voltage-gated calcium channels (T-channels). The molecular mechanisms immediately downstream from MR activation, which lead to the increased expression of T-channels and, consecutively, to an acceleration of spontaneous cell contractions in vitro, remain poorly investigated. Here, we investigated the putative role of a specific microRNA in linking MR activation to the regulation of T-channel expression and cardiomyocyte beating frequency. A screening assay identified microRNA 204 (miR-204) as one of the major upregulated microRNAs after aldosterone stimulation of isolated neonatal rat cardiomyocytes. Aldosterone significantly increased the level of miR-204, an effect blocked by the MR antagonist spironolactone. When miR-204 was overexpressed in isolated cardiomyocytes, their spontaneous beating frequency was significantly increased after 24 h, like upon aldosterone stimulation, and messenger RNAs coding T-channels (CaV3.1 and CaV3.2) were increased. Concomitantly, T-type calcium currents were significantly increased upon miR-204 overexpression. Specifically repressing the expression of miR-204 abolished the aldosterone-induced increase of CaV3.1 and CaV3.2 mRNAs, as well as T-type calcium currents. Finally, aldosterone and miR-204 overexpression were found to reduce REST-NRSF, a known transcriptional repressor of CaV3.2 T-type calcium channels. Our study thus strongly suggests that miR-204 expression stimulated by aldosterone promotes the expression of T-channels in isolated rat ventricular cardiomyocytes, and therefore, increases the frequency of the cell spontaneous contractions, presumably through the inhibition of REST-NRSF protein.

Acetylcholine-sensitive potassium channels in human atrial myocytes

1990 ◽  
Vol 259 (6) ◽  
pp. H1730-H1735 ◽  
Author(s):  
R. Sato ◽  
I. Hisatome ◽  
J. A. Wasserstrom ◽  
C. E. Arentzen ◽  
D. H. Singer
Keyword(s):  
Single Channel ◽  
K Channel ◽  
Channel Activity ◽  
Atrial Myocytes ◽  
Open Probability ◽  
Human Atrial Myocytes ◽  
Open Time ◽  
Patch Pipette ◽  
Single Channel Activity ◽  
Gamma S

Single channel recording techniques were used to study acetylcholine (ACh)-sensitive K+ channel activity in human atrial myocytes isolated from specimens obtained during corrective cardiac surgery. Under conditions of cell-attached patch, the presence of ACh in the patch pipette activated K+ channels. Single channel activity occurred in periodic bursts. The channels exhibited a slope conductance of 46 +/- 2 pS inwardly (means +/- SD, n = 4). During a burst, both open and closed time histograms were fitted by a single exponential curve, suggesting the existence of one open and one closed state during a burst. Open probability increased directly with ACh concentration without affecting open time. The channel could be activated by GTP and guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) (in the presence and absence of ACh in the pipette, respectively). Slope conductance, the response to GTP and GTP gamma S, and the independence of activation from Ca2+ were similar to those for other species. In contrast, sensitivity to ACh appeared diminished compared with frog atrial myocytes.

Single-channel properties of high- and low-voltage-activated calcium channels in rat pituitary melanotropic cells

1994 ◽  
Vol 71 (3) ◽  
pp. 840-855 ◽  
Author(s):  
J. A. Keja ◽  
K. S. Kits
Keyword(s):  
Calcium Channels ◽  
Kinetic Analysis ◽  
Calcium Current ◽  
Time Course ◽  
Single Channel ◽  
Low Voltage ◽  
Channel Activity ◽  
Open Probability ◽  
Test Pulse ◽  
Channel Properties

1. Single-channel properties of voltage-dependent calcium channels were investigated in rat melanotropes in short-term primary culture. Unitary currents were resolved using the cell-attached configuration. 2. Depolarizations higher than -50 mV activated a population of 8.1-pS calcium channels [low-voltage activated (LVA)]. The LVA channel ensembles displayed a monoexponential time course of inactivation and a sigmoidal time course of activation fitted best by an m2h Hodgkin-Huxley-type model. Microscopic kinetic analysis suggested that at least one open state, two closed states, and one inactivated state are involved in channel gating. 3. At potentials positive to -20 mV a second class of calcium channels was activated with a conductance of 24.7 pS [high-voltage activated (HVA)]. HVA channels display different gating modes. Gating with high open probability (mode 2) and low open probability (mode 1) as well as blank traces (mode 0) are observed. The HVA channels were heterogeneous with respect to their inactivation properties. Ensembles that decayed entirely during a 300-ms test pulse as well as nondecaying ensembles were observed. Both HVA channel subtypes displayed sigmoidal activation, which was fitted by an m2 model. Microscopic kinetic analysis suggested that at least one open state and two closed states are involved in mode two gating of both HVA channel subtypes. 4. Depolarizing prepulses did not recruit or facilitate calcium channel activity in response to a test pulse, but inactivating HVA channel activity was strongly reduced. Depolarizing prepulses (+50 mV) did not affect the probability of opening of the noninactivating HVA channel. 5. The voltage dependence and kinetics of the LVA as well as both HVA channels are in good agreement with previously published data on the properties of the various calcium current components derived from whole-cell recordings of rat melanotropes. The data suggest that a T-type as well as two L-type channels (an inactivating and noninactivating channel) underlie the calcium current in these cells.

scholarly journals Voltage-dependent inactivation of T-tubular skeletal calcium channels in planar lipid bilayers.

1991 ◽  
Vol 97 (2) ◽  
pp. 393-412 ◽  
Author(s):  
R Mejía-Alvarez ◽  
M Fill ◽  
E Stefani
Keyword(s):  
Calcium Channels ◽  
Lipid Bilayers ◽  
Single Channel ◽  
Channel Activity ◽  
T Tube ◽  
Dependent Inactivation ◽  
Voltage Dependent ◽  
Single Channel Activity ◽  
Kinetics Of ◽  
Channel Properties

Single-channel properties of dihydropyridine (DHP)-sensitive calcium channels isolated from transverse tubular (T-tube) membrane of skeletal muscle were explored. Single-channel activity was recorded in planar lipid bilayers after fusion of highly purified rabbit T-tube microsomes. Two populations of DHP-sensitive calcium channels were identified. One type of channel (noninactivating) was active (2 microM +/- Bay K 8644) at steady-state membrane potentials and has been studied in other laboratories. The second type of channel (inactivating) was transiently activated during voltage pulses and had a very low open probability (Po) at steady-state membrane potentials. Inactivating channel activity was observed in 47.3% of the experiments (n = 84 bilayers). The nonstationary kinetics of this channel was determined using a standard voltage pulse (HP = -50 mV, pulse to 0 mV). The time constant (tau) of channel activation was 23 ms. During the mV). The time constant (tau) of channel activation was 23 ms. During the pulse, channel activity decayed (inactivated) with a tau of 3.7 s. Noninactivating single-channel activity was well described by a model with two open and two closed states. Inactivating channel activity was described by the same model with the addition of an inactivated state as proposed for cardiac muscle. The single-channel properties were compared with the kinetics of DHP-sensitive inward calcium currents (ICa) measured at the cellular level. Our results support the hypothesis that voltage-dependent inactivation of single DHP-sensitive channels contributes to the decay of ICa.

A calcium-activated chloride channel in sarcoplasmic reticulum vesicles from rabbit skeletal muscle

1996 ◽  
Vol 270 (6) ◽  
pp. C1675-C1686 ◽  
Author(s):  
J. I. Kourie ◽  
D. R. Laver ◽  
G. P. Ahern ◽  
A. F. Dulhunty
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Lipid Bilayers ◽  
Chloride Channel ◽  
Single Channel ◽  
Single Channels ◽  
Channel Activity ◽  
Open Probability ◽  
Single Channel Activity ◽  
Maximum Conductance

A Ca(2+)-activated Cl- channel is described in sarcoplasmic reticulum (SR) enriched vesicles of skeletal muscle incorporated into lipid bilayers. Small chloride (SCl) channels (n = 20) were rapidly and reversibly activated when cis- (cytoplasmic) [Ca2+] was increased above 10(-7) M, with trans-(luminal) [Ca2+] at either 10(-3) or 10(-7) M. The open probability of single channels increased from zero when cis-[Ca2+] was 10(-7) M to 0.61 +/- 0.12 when [Ca2+] was 10(-4) M. High- and low-conductance levels in single-channel activity were activated at different cis-[Ca2+]. Channel openings to the maximum conductance, 65-75 pS (250/50 mM Cl-, cis/ trans), were active when cis-[Ca2+] was increased above 5 x 10(-6) M. In contrast to the maximum conductance, channel openings to submaximal levels between 5 and 40 pS were activated at a lower cis-[Ca2+] and dominated channel activity between 5 x 10(-7) and 5 x 10(-6) M. Activation of SCl channels was Ca2+ specific and not reproduced by cytoplasmic Mg2+ concentrations of 10(-3) M. We suggest that the SCl channel arises in the SR membrane. The Ca2+ dependence of this channel implies that it is active at [Ca2+] achieved during muscle contraction.

Activation of cardiac ryanodine receptors by the calcium channel agonist FPL-64176

2002 ◽  
Vol 283 (1) ◽  
pp. H331-H338 ◽  
Author(s):  
J. Andrew Wasserstrom ◽  
Leslie A. Wasserstrom ◽  
Andrew J. Lokuta ◽  
James E. Kelly ◽  
Sireen T. Reddy ◽  
...  
Keyword(s):  
Single Channel ◽  
Ryanodine Receptors ◽  
Channel Activity ◽  
Open Probability ◽  
Release Channel ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Significant Difference ◽  
Single Channel Activity ◽  
Control Versus ◽  
Channel Agonist

We investigated the possibility that the Ca2+ channel agonist FPL-64176 (FPL) might also activate the cardiac sarcoplasmic reticulum (SR) Ca2+ release channel ryanodine receptor (RyR). The effects of FPL were tested on single channel activity of purified and crude vesicular RyR (RyR2) isolated from human and dog hearts using the planar lipid bilayer technique. FPL (100–200 μM) increased single channel open probability ( P o) when added to the cytoplasmic side of the channel ( P o = 0.070 ± 0.021 in control RyR2; 0.378 ± 0.086 in 150 μM FPL, n = 9, P < 0.01) by prolonging open times and decreasing closed times without changing current magnitude. FPL had no effect on P o when added to the trans (luminal) side of the bilayer ( P o = 0.079 ± 0.036 in control and 0.103 ± 0.066 in FPL, n = 4, no significant difference). The bell-shaped [Ca2+] dependence of [3H]ryanodine binding and of P o was altered by FPL, suggesting that the mechanism by which FPL increases channel activity is by an increase in Ca2+-induced activation at low [Ca2+] (without a change in threshold) and suppression of Ca2+-induced inactivation at high [Ca2+]. However, the fact that inactivation was restored at elevated [Ca2+] suggests a competitive interaction between Ca2+ and FPL on inactivation. FPL had no effect on RyR skeletal channels (RyR1), where P o was 0.039 ± 0.005 in control versus 0.030 ± 0.006 in 150 μM FPL (no significant difference). These results suggest that, in addition to its ability to activate the L-type Ca2+channels, FPL activates cardiac RyR2 primarily by reducing the Ca2+ sensitivity of inactivation.

scholarly journals Nonmodal gating of cardiac calcium channels as revealed by dihydropyridines.

1989 ◽  
Vol 93 (6) ◽  
pp. 1243-1273 ◽  
Author(s):  
A E Lacerda ◽  
A M Brown
Keyword(s):  
Calcium Channels ◽  
Single Channel ◽  
Calcium Currents ◽  
Neonatal Rat ◽  
Whole Cell ◽  
Frequency Distributions ◽  
Time Frequency ◽  
Channel Current ◽  
Open Time ◽  
Open States

The hypothesis that dihydropyridine (DHP)-sensitive calcium channels have three distinct modes of gating has been examined. The major prediction is that the relative frequencies among modes depend on DHP concentration while the kinetics within a mode do not. We tested this by studying whole-cell and single-channel calcium currents in neonatal rat and adult guinea pig cardiac myocytes in different concentrations of several DHPs. In the absence of DHPs calcium currents declined with time but the kinetics, which are the focus of this study, were unchanged. Open-time frequency distributions had insignificant numbers of prolonged openings and were well fit by single tau's. Agonist DHP stereoisomers produced concentration-dependent changes in whole-cell tail current tau's. The frequency distribution of single calcium channel current open times became biexponential and the tau's were concentration dependent. The average number of openings per trace of channels with customary open times increased with increases in DHP concentration. Latencies to first opening for the customary openings and for prolonged openings were shorter in the presence of DHPs. A second larger conductance is another important feature of DHP-bound single calcium channels. Thus DHPs not only caused prolonged openings; they produced numerous changes in the kinetics of customary openings and increased channel conductance. It follows that these effects of DHPs do not support the hypothesis of modal gating of calcium channels. The mode model is not the only model excluded by the results; models in which DHPs are allowed to act only or mainly on open states are excluded, as are models in which the effects are restricted to inactivated states. We suggest a different type of model in which cooperative binding of DHPs at two sites produces the essential changes in kinetics and conductance.

Inwardly rectifying chloride channel activity in intestinal pacemaker cells

2005 ◽  
Vol 288 (4) ◽  
pp. G809-G821 ◽  
Author(s):  
Yaohui Zhu ◽  
Andrea Mucci ◽  
Jan D. Huizinga
Keyword(s):  
Single Channel ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Pacemaker Activity ◽  
Channel Activity ◽  
Pacemaker Cells ◽  
Open Probability ◽  
Whole Cell ◽  
Single Channel Activity ◽  
Inwardly Rectifying

Cl− channels are proposed to play a role in gut pacemaker activity, but little is known about the characteristics of Cl− channels in interstitial cells of Cajal (ICC), the intestinal pacemaker cells. The objective of the present study was to identify whole cell Cl− currents in ICC associated with previously observed single-channel activity and to characterize its inward rectification. Whole cell patch-clamp studies showed that ICC express an inwardly rectifying Cl− current that was not sensitive to changes in cation composition of the extracellular solutions. Currents were not affected by replacing all cations with N-methyl-d-glucamine (NMDG+). Whole cell currents followed the Cl− equilibrium potential and were inhibited by DIDS and 9-anthracene carboxylic acid. Ramp protocols of single-channel activity showed that inward rectification was due to reduction in single-channel open probability, not a reduction in single-channel conductance. Single-channel data led to the hypothesis that strong cooperation exists between 30-pS channels that show less cooperation at potentials positive to the reversal potential. Hence, an inwardly rectifying Cl− channel plays a prominent role in determining pacemaker activity in the gut.

Activation of purified cardiac ryanodine receptors by dihydropyridine agonists

2001 ◽  
Vol 280 (3) ◽  
pp. H1201-H1207 ◽  
Author(s):  
Toshio Sagawa ◽  
Manabu Nishio ◽  
Kazuko Sagawa ◽  
James E. Kelly ◽  
Andrew J. Lokuta ◽  
...  
Keyword(s):  
Single Channel ◽  
Ryanodine Receptors ◽  
Channel Activity ◽  
Allosteric Site ◽  
Open Probability ◽  
Release Channel ◽  
Inotropic Effects ◽  
Cytoplasmic Face ◽  
Single Channel Activity ◽  
Negative Inotropic Effects

Prior observations have raised the possibility that dihydropyridine (DHP) agonists directly affect the sarcoplasmic reticulum (SR) cardiac Ca2+ release channel [i.e., ryanodine receptor (RyR)]. In single-channel recordings of purified canine cardiac RyR, both DHP agonists (−)-BAY K 8644 and (+)-SDZ202-791 increased the open probability of the RyR when added to the cytoplasmic face of the channel. Importantly, the DHP antagonists nifedipine and (−)-SDZ202-791 had no competitive blocking effects either alone or after channel activation with agonist. Thus there is a stereospecific effect of SDZ202-791, such that the agonist activates the channel, whereas the antagonist has little effect on channel activity. Further experiments showed that DHP agonists changed RyR activation by suppressing Ca2+-induced inactivation of the channel. We concluded that DHP agonists can also influence RyR single-channel activity directly at a unique allosteric site located on the cytoplasmic face of the channel. Similar results were obtained in human purified cardiac RyR. An implication of these data is that RyR activation by DHP agonists is likely to cause a loss of Ca2+ from the SR and to contribute to the negative inotropic effects of these agents reported by other investigators. Our results support this notion that the negative inotropic effects of DHP agonists result in part from direct alteration in the activity of RyRs.

scholarly journals Membrane-delimited Inhibition of Maxi-K Channel Activity by the Intermediate Conductance Ca2+-activated K Channel

2006 ◽  
Vol 127 (2) ◽  
pp. 159-169 ◽  
Author(s):  
Jill Thompson ◽  
Ted Begenisich
Keyword(s):  
Single Channel ◽  
Expression System ◽  
Chemical Activation ◽  
K Channel ◽  
Channel Activity ◽  
Single Family ◽  
Open Probability ◽  
K Channels ◽  
Channel Proteins ◽  
K Activity

The complexity of mammalian physiology requires a diverse array of ion channel proteins. This diversity extends even to a single family of channels. For example, the family of Ca2+-activated K channels contains three structural subfamilies characterized by small, intermediate, and large single channel conductances. Many cells and tissues, including neurons, vascular smooth muscle, endothelial cells, macrophages, and salivary glands express more than a single class of these channels, raising questions about their specific physiological roles. We demonstrate here a novel interaction between two types of Ca2+-activated K channels: maxi-K channels, encoded by the KCa1.1 gene, and IK1 channels (KCa3.1). In both native parotid acinar cells and in a heterologous expression system, activation of IK1 channels inhibits maxi-K activity. This interaction was independent of the mode of activation of the IK1 channels: direct application of Ca2+, muscarinic receptor stimulation, or by direct chemical activation of the IK1 channels. The IK1-induced inhibition of maxi-K activity occurred in small, cell-free membrane patches and was due to a reduction in the maxi-K channel open probability and not to a change in the single channel current level. These data suggest that IK1 channels inhibit maxi-K channel activity via a direct, membrane-delimited interaction between the channel proteins. A quantitative analysis indicates that each maxi-K channel may be surrounded by four IK1 channels and will be inhibited if any one of these IK1 channels opens. This novel, regulated inhibition of maxi-K channels by activation of IK1 adds to the complexity of the properties of these Ca2+-activated K channels and likely contributes to the diversity of their functional roles.

Sign in / Sign up

Export Citation Format

Share Document