Modulation by endogenously released ATP and opioids of chromaffin cell calcium channels in mouse adrenal slices

2011 ◽  
Vol 300 (3) ◽  
pp. C610-C623 ◽  
Author(s):  
A. Hernández ◽  
P. Segura-Chama ◽  
N. Jiménez ◽  
A. G. García ◽  
J. M. Hernández-Guijo ◽  
...  
Keyword(s):  
Calcium Channels ◽  
Chromaffin Cells ◽  
Opiate Receptors ◽  
Tonic Inhibition ◽  
Inactivation Kinetics ◽  
High Threshold ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Voltage Dependent ◽  
Patch Configuration

Modulation of high-threshold voltage-dependent calcium channels by neurotransmitters has been the subject of numerous studies in cultures of neurons and chromaffin cells. However, no studies on such modulation exist in chromaffin cells in their natural environment, the intact adrenal medullary tissue. Here we performed such a study in voltage-clamped chromaffin cells of freshly prepared mouse adrenal slices under the whole cell configuration of the patch-clamp technique. The subcomponents of the whole cell inward Ca2+ current ( ICa) accounted for 49% for L-, 28% for N-, and 36% for P/Q-type channels. T-type Ca2+ channels or residual R-type Ca2+ currents were not seen. However, under the perforated-patch configuration, 20% of ICa accounted for a toxin-resistant R-type Ca2+ current. Exogenously applied ATP and methionine-enkephalin (Met-enk) inhibited ICa by 33%. Stop-flow and Ca2+ replacement by Ba2+, which favored the release of endogenous ATP and opioids, also inhibited ICa, with no changes in activation or inactivation kinetics. This inhibition was partially voltage independent and insensitive to prepulse facilitation. Furthermore, in about half of the cells, suramin and naloxone augmented ICa in the absence of exogenous application of ATP/Met-enk. No additional modulation of ICa was obtained after bath application of exogenous ATP and opioids to these already inhibited cells. Augmentation of ICa was also seen upon intracellular dialysis of guanosine 5′-[β-thio]diphosphate (GDPβS), indicating the existence in the intact slice of a tonic inhibition of ICa in resting conditions. These results suggest that in the intact adrenal tissue a tonic inhibition of ICa exists, mediated by purinergic and opiate receptors.

scholarly journals Ascorbic acid modulation of calcium channels in pancreatic beta cells.

1993 ◽  
Vol 102 (3) ◽  
pp. 503-523 ◽  
Author(s):  
R V Parsey ◽  
D R Matteson
Keyword(s):  
Ascorbic Acid ◽  
Calcium Channels ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Inactivation Kinetics ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Voltage Dependent ◽  
Voltage Dependent Calcium Channels ◽  
Metal Catalyzed

We have studied the effect of ascorbic acid on voltage-dependent calcium channels in pancreatic beta cells. Using the whole-cell and perforated-patch variants of the patch clamp technique to record calcium tail currents, we have shown that the slowly deactivating (SD) calcium channel, which is similar to the T-type channel in other cells, is inhibited in a voltage-dependent manner by ascorbic acid (AA). The other channels that carry inward current in beta cells, FD calcium channels and sodium channels, are unaffected by AA. Ascorbic acid causes a voltage-dependent decrease in the magnitude of the SD channel conductance which can be explained by the hypothesis that approximately 50-60% of the channels have their voltage dependence shifted by approximately 62 mV in the depolarizing direction. Thus, ascorbate appears to modify only a fraction of the SD channels. The activation kinetics of the ascorbate-modified channels are slower than control channels in a manner that is consistent with this hypothesis. Deactivation and inactivation kinetics are unaffected by ascorbate. These effects of ascorbate require metal ions, and it appears that some of the activity of ascorbate is due to a product of its metal catalyzed oxidation, perhaps dehydroascorbate.

Extracellular inhibition and intracellular enhancement of Ca2+ currents by Pb2+ in bovine adrenal chromaffin cells

1995 ◽  
Vol 74 (2) ◽  
pp. 574-581 ◽  
Author(s):  
L. R. Sun ◽  
J. B. Suszkiw
Keyword(s):  
Calcium Channels ◽  
Chromaffin Cells ◽  
Adrenal Chromaffin Cells ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent ◽  
Steady State Inactivation ◽  
Voltage Dependent Calcium Channels ◽  
Adrenal Chromaffin ◽  
Dose Dependent

1. Effects of highly neurotoxic, inorganic lead ions (Pb2+) on voltage-dependent calcium channels were investigated with the use of the whole cell patch-clamp technique in bovine adrenal chromaffin cells maintained in short-term primary culture (1–5 days). 2. Extracellularly applied Pb2+ induced a concentration-dependent, reversible inhibition of Ca2+ currents, with an estimated IC50 approximately equal to 3.0 x 10(-7) M free Pb2+. 3. Elevation of the intracellular free Ca2+ concentration above 10(-8) M dose-dependently reduced the amplitude of the initial Ca2+ current and increased the exponential rate of current rundown. 4. Intracellularly applied Pb2+ prevented the Ca(2+)-dependent reduction of the initial Ca2+ current amplitude and altered the current rundown kinetics from exponential to linear. The effect was dose dependent and saturable, with an estimated EC50 approximately equal to 2.0 x 10(-10) M free Pb2+. 5. These results indicate that in contrast to extracellular blockade, intracellular Pb2+ promotes Ca2+ currents by attenuating the Ca(2+)-dependent, steady-state inactivation of calcium channels. This provides a novel mechanism through which Pb2+ may disrupt calcium signaling in chronically lead-exposed cells.

Voltage-Dependent Sodium and Calcium Currents in Acutely Isolated Adult Rat Trigeminal Root Ganglion Neurons

1999 ◽  
Vol 81 (3) ◽  
pp. 1123-1134 ◽  
Author(s):  
Hyung-Chan Kim ◽  
Man-Kyo Chung
Keyword(s):  
Calcium Currents ◽  
Inactivation Kinetics ◽  
Type I ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Adult Rat ◽  
Voltage Dependent ◽  
Trigeminal Root ◽  
Ganglion Neurons ◽  
Drug Free

Voltage-dependent sodium and calcium currents in acutely isolated adult rat trigeminal root ganglion neurons. Voltage-dependent sodium ( I Na) and calcium ( I Ca) currents in small (<30 μM) neurons from adult rat trigeminal root ganglia were characterized with a standard whole cell patch-clamp technique. Two types of I Na showing different sensitivity to tetrodotoxin (TTX) were recorded, which showed marked differences in their activating and inactivating time courses. The activation and the steady-state inactivation kinetics of TTX-resistant I Na were more depolarized by about +20 and +30 mV, respectively, than those of TTX-sensitive I Na. Voltage-dependent I Ca was recorded under the condition that suppressed sodium and potassium currents with 10 mM Ca2+ as a charge carrier. Depolarizing step pulses from a holding potential of −80 mV evoked two distinct inward I Ca, low-voltage activated (LVA) and high-voltage activated (HVA) I Ca. LVA I Ca was first observed at −60 to −50 mV and reached a peak at about −30 mV. Amiloride (0.5 mM) suppressed ∼60% of the LVA I Ca, whereas ∼10% of HVA I Ca was inhibited by the same concentration of the amiloride. LVA I Ca was far less affected by the presence of external Cd2+ or the replacement of Ca2+ by 10 Ba2+ than HVA I Ca. The ω-conotoxin GVIA (ω-CgTx), an N-type I Ca blocker, suppressed ∼65% of the whole cell HVA I Ca at the concentration of 1 μM. The ω-CgTx-resistant HVA I Ca was sensitive to nifedipine (10 μM), a dihydropyridine (DHP) calcium channel antagonist, which produced an additional blockade by ∼25% of the drug-free control (∼70% of the ω-CgTx-resistant I Ca). The combination of 10 μM nifedipine and 1 μM ω-CgTx left ∼13% of the drug-free control I Ca unblocked. The DHP agonist S(−)-BayK8644 (5 μM) shifted the activation of the HVA I Ca to more negative potentials and increased its maximal amplitude. Additionally, S(−)-BayK8644 caused the appearance of a slowed component of the tail current. These results clearly demonstrate that the presence of two types of sodium channels, TTX sensitive and resistant, and three types of calcium channels, T, L, and N type, in the small-sized adult rat trigeminal ganglion neurons.

Neuropeptide galanin inhibits omega-conotoxin GVIA-sensitive calcium channels in parasympathetic neurons

1995 ◽  
Vol 73 (4) ◽  
pp. 1374-1382 ◽  
Author(s):  
L. A. Merriam ◽  
R. L. Parsons
Keyword(s):  
Calcium Channels ◽  
Patch Clamp Technique ◽  
Maximal Inhibition ◽  
Whole Cell ◽  
Recording Technique ◽  
Perforated Patch ◽  
Cell Recording ◽  
Parasympathetic Neurons ◽  
Current Activation ◽  
Patch Technique

1. We determined the effect of the neuropeptide galanin on barium currents (IBa) flowing through voltage-gated calcium channels. We voltage clamped parasympathetic neurons dissociated from mudpuppy cardiac ganglia using both the standard whole cell and the perforated-patch variations of the patch-clamp technique. 2. Galanin produced a concentration-dependent inhibition of IBa. The maximal inhibition was 50-60% and the concentration that produced half-maximal inhibition (IC50) was 0.42 nM. In mud-puppy parasympathetic neurons, omega-conotoxin-GVIA (CTX)-sensitive channels are the predominant type of calcium channels, and only a small portion of IBa is contributed by dihydropyridine-sensitive channels. Galanin preferentially inhibited a portion of the CTX-sensitive current. 3. In currents recorded with the standard whole cell technique, activation of IBa was slowed in the presence of galanin. In contrast, in the majority of neurons studied with the perforated-patch technique, galanin decreased IBa without altering the kinetics of current activation. With both recording methods, the decrease in IBa was greatest with voltage steps to 0 mV and persisted with steps to +50 mV. For control currents, large depolarizing voltage steps (+70 to +120 mV) did not markedly facilitate IBa when either recording technique was used. However, the degree of facilitation in galanin was significantly greater with the standard whole cell recording technique. 4. IBa exhibited inactivation under the conditions of these experiments. Inactivation of IBa recorded during a 900-ms depolarizing voltage step was fitted to a double exponential. Galanin decreased the amplitude of IBa but did not alter the time constants of inactivation.(ABSTRACT TRUNCATED AT 250 WORDS)

Voltage-dependent calcium channels in ventricular cells of rainbow trout: effect of temperature changes in vitro

2000 ◽  
Vol 278 (6) ◽  
pp. R1524-R1534 ◽  
Author(s):  
Catherine S. Kim ◽  
Mary D. Coyne ◽  
Judith K. Gwathmey
Keyword(s):  
Rainbow Trout ◽  
Calcium Channels ◽  
Temperature Sensitivity ◽  
Calcium Currents ◽  
Temperature Dependency ◽  
Patch Clamp Technique ◽  
Ventricular Cells ◽  
Voltage Dependent ◽  
Voltage Dependent Calcium Channels

Voltage-dependent calcium channels (VDCC) in ventricular myocytes from rainbow trout ( Oncorhynchus mykiss) were investigated in vitro using the perforated patch-clamp technique, which maintains the integrity of the intracellular milieu. First, we characterized the current using barium as the charge carrier and established the doses of various pharmacological agents to use these agents in additional studies. Second, we examined the current at several physiological temperatures to determine temperature dependency. The calcium currents at 10°C (acclimation temperature) were identified as l-type calcium currents based on their kinetic behavior and response to various calcium channel agonists and antagonists. Myocytes were chilled (4°C) and warmed (18 and 22°C), and the response of VDCC to varying temperatures was observed. There was no significant dependency of the current amplitude and kinetics on temperature. Amplitude decreased 25–36% at 4°C (Q10 ∼1.89) and increased 18% at 18°C (Q10 ∼1.23) in control, Bay K8644 (Bay K)-, and forskolin-enhanced currents. The inactivation rates (τi) did not demonstrate a temperature sensitivity for the VDCC (Q10 1.23–1.92); Bay K treatment, however, increased temperature sensitivity of τi between 10 and 18°C (Q10 3.98). The low Q10 values for VDCC are consistent with a minimal temperature sensitivity of trout myocytes between 4 and 22°C. This low-temperature dependency may provide an important role for sarcolemmal calcium channels in adaptation to varying environmental temperatures in trout.

scholarly journals Slow inactivation of L-type calcium current distorts the measurement of L- and T-type calcium current in Purkinje myocytes.

1993 ◽  
Vol 102 (5) ◽  
pp. 859-869 ◽  
Author(s):  
N B Datyner ◽  
I S Cohen
Keyword(s):  
Patch Clamp ◽  
Calcium Current ◽  
Slow Inactivation ◽  
Total Calcium ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Whole Cell Patch Clamp ◽  
Voltage Curve ◽  
Voltage Dependent ◽  
Method Of Measurement

We have examined slow inactivation of L-type calcium current in canine Purkinje myocytes with the whole cell patch clamp technique. Slow inactivation is voltage dependent. It is negligible at -50 mV but can inactivate more than half of available iCaL at -10 mV. There are two major consequences of this slow inactivation. First, standard protocols for the measurement of T-type current can dramatically overestimate its contribution to total calcium current, and second, the position and steepness of the inactivation versus voltage curve for iCaL will depend on the method of measurement. Given the widespread attempts to identify calcium current components and characterize them biophysically, an important first step should be to determine the extent of slow inactivation of calcium current in each preparation.

scholarly journals Single calcium channel behavior in native skeletal muscle.

1995 ◽  
Vol 105 (2) ◽  
pp. 227-247 ◽  
Author(s):  
R T Dirksen ◽  
K G Beam
Keyword(s):  
Skeletal Muscle ◽  
Calcium Channels ◽  
Single Channel ◽  
Bay K 8644 ◽  
Dependent Manner ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Whole Cell ◽  
Macroscopic Properties ◽  
Voltage Dependent

The purpose of this study was to use whole-cell and cell-attached patches of cultured skeletal muscle myotubes to study the macroscopic and unitary behavior of voltage-dependent calcium channels under similar conditions. With 110 mM BaCl2 as the charge carrier, two types of calcium channels with markedly different single-channel and macroscopic properties were found. One class was DHP-insensitive, had a single-channel conductance of approximately 9 pS, yielded ensembles that displayed an activation threshold near -40 mV, and activated and inactivated rapidly in a voltage-dependent manner (T current). The second class could only be well resolved in the presence of the DHP agonist Bay K 8644 (5 microM) and had a single-channel conductance of approximately 14 pS (L current). The 14-pS channel produced ensembles exhibiting a threshold of approximately -10 mV that activated slowly (tau act approximately 20 ms) and displayed little inactivation. Moreover, the DHP antagonist, (+)-PN 200-110 (10 microM), greatly increased the percentage of null sweeps seen with the 14-pS channel. The open probability versus voltage relationship of the 14-pS channel was fitted by a Boltzmann distribution with a VP0.5 = 6.2 mV and kp = 5.3 mV. L current recorded from whole-cell experiments in the presence of 110 mM BaCl2 + 5 microM Bay K 8644 displayed similar time- and voltage-dependent properties as ensembles of the 14-pS channel. Thus, these data are the first comparison under similar conditions of the single-channel and macroscopic properties of T current and L current in native skeletal muscle, and identify the 9- and 14-pS channels as the single-channel correlates of T current and L current, respectively.

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