Ca2+ Currents in Central Insect Neurons: Electrophysiological and Pharmacological Properties

1997 ◽  
Vol 77 (1) ◽  
pp. 186-199 ◽  
Author(s):  
Dieter Wicher ◽  
Heinz Penzlin
Keyword(s):  
Periplaneta Americana ◽  
Low Voltage ◽  
Fast Time ◽  
Channel Blockers ◽  
Pharmacological Properties ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Blocking Effects ◽  
Dum Neurons ◽  
Maximal Peak

Wicher, Dieter, and Heinz Penzlin. Ca2+ currents in central insect neurons: electrophysiological and pharmacological properties. J. Neurophysiol. 77: 186–199, 1997. Ca2+ currents in dorsal unpaired median (DUM) neurons isolated from the fifth abdominal ganglion of the cockroach Periplaneta americana were investigated with the whole cell patch-clamp technique. On the basis of kinetic and pharmacological properties, two different Ca2+ currents were separated in these cells: mid/low-voltage-activated (M-LVA) currents and high-voltage-activated (HVA) currents. M-LVA currents had an activation threshold of −50 mV and reached maximal peak values at −10 mV. They were sensitive to depolarized holding potentials and decayed very rapidly. The decay was largely Ca2+ dependent. M-LVA currents were effectively blocked by Cd2+ median inhibiting concentration (IC50 = 9 μM), but they also had a remarkable sensitivity to Ni2+ (IC50 = 19 μM). M-LVA currents were insensitive to vertebrate LVA channel blockers like flunarizine and amiloride. The currents were, however, potently blocked by ω-conotoxin MVIIC (1 μM) and ω-agatoxin IVA (50 nM). The blocking effects of ω-toxins developed fast (time constant τ = 15 s) and were fully reversible after wash. HVA currents activated positive to −30 mV and showed maximal peak currents at +10 mV. They were resistant to depolarized holding potentials up to −50 mV and decayed in a less pronounced manner than M-LVA currents. HVA currents were potently blocked by Cd2+ (IC50 = 5 μM) but less affected by Ni2+ (IC50 = 40 μM). These currents were reduced by phenylalkylamines like verapamil (10 μM) and benzothiazepines like diltiazem (10 μM), but they were insensitive to dihydropyridines like nifedipine (10 μM) and BAY K 8644 (10 μM). Furthermore, HVA currents were sensitive to ω-conotoxin GVIA (1 μM). The toxin-induced reduction of currents appeared slowly (τ ∼ 120 s) and the recovery after wash was incomplete in most cases. The dihydropyridine insensitivity of the phenylalkylamine-sensitive HVA currents is a property the cockroach DUM cells share with other invertebrate neurons. Compared with Ca2+ currents in vertebrates, the DUM neuron currents differ considerably from the presently known types. Although there are some similarities concerning kinetics, the pharmacological profile of the cockroach Ca2+ currents especially is very different from profiles already described for vertebrate currents.

Inactivation Properties of Human Recombinant Class E Calcium Channels

2000 ◽  
Vol 83 (2) ◽  
pp. 671-684 ◽  
Author(s):  
Anne Jouvenceau ◽  
Federica Giovannini ◽  
Cath P. Bath ◽  
Emily Trotman ◽  
Emanuele Sher
Keyword(s):  
Calcium Channels ◽  
Low Voltage ◽  
Channel Blockers ◽  
Pharmacological Properties ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Hek 293 ◽  
Clear Cut ◽  
Dependent Inactivation ◽  
Voltage Dependent

The electrophysiological and pharmacological properties of α1E-containing Ca2+ channels were investigated by using the patch-clamp technique in the whole cell configuration, in HEK 293 cells stably expressing the human α1E together with α2b and β1baccessory subunits. These channels had current-voltage ( I-V) characteristics resembling those of high-voltage–activated (HVA) Ca2+ channels (threshold at −30 mV and peak amplitude at +10 mV in 5 mM Ca2+). The currents activated and deactivated with a fast rate, in a time- and voltage-dependent manner. No difference was found in their relative permeability to Ca2+ and Ba2+. Inorganic Ca2+ channel blockers (Cd2+, Ni2+) blocked completely and potently the α1E,/α2bδ/β1b mediated currents (IC50 = 4 and 24.6 μM, respectively). α1E-mediated currents inactivated rapidly and mainly in a non–Ca2+-dependent manner, as evidenced by the fact that 1) decreasing extracellular Ca2+ from 10 to 2 mM and 2) changing the intracellular concentration of the Ca2+ chelator 1.2-bis(2-aminophenoxy) ethane- N,N,N′,N′-tetraacetic acid (BAPTA), did not affect the inactivation characteristics; 3) there was no clear-cut bell-shaped relationship between test potential and inactivation, as would be expected from a Ca2+-dependent event. Although Ba2+ substitution did not affect the inactivation of α1E channels, Na+substitution revealed a small but significant reduction in the extent and rate of inactivation, suggesting that besides the presence of dominant voltage-dependent inactivation, α1E channels are also affected by a divalent cation-dependent inactivation process. We have analyzed the Ca2+ currents produced by a range of imposed action potential–like voltage protocols (APVPs). The amplitude and area of the current were dependent on the duration of the waveform employed and were relatively similar to those described for HVA calcium channels. However, the peak latency resembled that obtained for low-voltage–activated (LVA) calcium channels. Short bursts of APVPs applied at 100 Hz produced a depression of the Ca2+ current amplitude, suggesting an accumulation of inactivation likely to be calcium dependent. The human α1E gene seems to participate to a Ca2+ channel type with biophysical and pharmacological properties partly resembling those of LVA and those of HVA channels, with inactivation characteristics more complex than previously believed.

Characterization of voltage-dependent calcium currents in mouse motoneurons

1992 ◽  
Vol 68 (1) ◽  
pp. 85-92 ◽  
Author(s):  
M. Mynlieff ◽  
K. G. Beam
Keyword(s):  
Voltage Dependence ◽  
Low Voltage ◽  
Calcium Currents ◽  
Patch Clamp Technique ◽  
Maximal Amplitude ◽  
Time To Peak ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent ◽  
Channel Currents

1. Calcium channel currents were measured with the whole-cell patch clamp technique in cultured, identified mouse motoneurons. Three components of current were operationally defined on the basis of voltage dependence, kinetics, and pharmacology. 2. Test potentials to -50 mV or greater (10 mM external Ca2+) elicited a low-voltage activated T-type current that was transient (decaying to baseline in less than 200 ms) and had a relatively slow time to peak (20-50 ms). A 1-s prepulse to -45 mV produced approximately half-maximal inactivation of this T current. 3. Two high-voltage activated (HVA) components of current (1 transient and 1 sustained) were activated by test potentials to -20 mV or greater (10 mM external Ca2+). A 1-s prepulse to -35 mV produced approximately half-maximal inactivation of the transient component without affecting the sustained component. 4. When Ba2+ was substituted for Ca2+ as the charge carrier, activation of the HVA components was shifted in the hyperpolarizing direction, and the relative amplitude of the transient HVA component was reduced. 5. Amiloride (1-2 mM) caused a reversible, partial block of the T current without affecting the HVA components. 6. The dihydropyridine agonist isopropyl 4-(2,1,3-benzoxadiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5-nitro-3- pyridine-carboxylate [(+)-SDZ 202-791, 100 nM-1 microM)] shifted the activation of the sustained component of HVA current to more negative potentials and increased its maximal amplitude. Additionally, (+)-SDZ 202-791 caused the appearance of a slowed component of tail current.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Differential Interactions of Na+ Channel Toxins with T-type Ca2+ Channels

2008 ◽  
Vol 132 (1) ◽  
pp. 101-113 ◽  
Author(s):  
Hui Sun ◽  
Diego Varela ◽  
Denis Chartier ◽  
Peter C. Ruben ◽  
Stanley Nattel ◽  
...  
Keyword(s):  
Ventricular Myocytes ◽  
Low Voltage ◽  
Parallel Evolution ◽  
Na Channel ◽  
Patch Clamp Technique ◽  
Hek 293 ◽  
Inward Current ◽  
Whole Cell Patch Clamp ◽  
The Right ◽  
Ca2 Channels

Two types of voltage-dependent Ca2+ channels have been identified in heart: high (ICaL) and low (ICaT) voltage-activated Ca2+ channels. In guinea pig ventricular myocytes, low voltage–activated inward current consists of ICaT and a tetrodotoxin (TTX)-sensitive ICa component (ICa(TTX)). In this study, we reexamined the nature of low-threshold ICa in dog atrium, as well as whether it is affected by Na+ channel toxins. Ca2+ currents were recorded using the whole-cell patch clamp technique. In the absence of external Na+, a transient inward current activated near −50 mV, peaked at −30 mV, and reversed around +40 mV (HP = −90 mV). It was unaffected by 30 μM TTX or micromolar concentrations of external Na+, but was inhibited by 50 μM Ni2+ (by ∼90%) or 5 μM mibefradil (by ∼50%), consistent with the reported properties of ICaT. Addition of 30 μM TTX in the presence of Ni2+ increased the current approximately fourfold (41% of control), and shifted the dose–response curve of Ni2+ block to the right (IC50 from 7.6 to 30 μM). Saxitoxin (STX) at 1 μM abolished the current left in 50 μM Ni2+. In the absence of Ni2+, STX potently blocked ICaT (EC50 = 185 nM) and modestly reduced ICaL (EC50 = 1.6 μM). While TTX produced no direct effect on ICaT elicited by expression of hCaV3.1 and hCaV3.2 in HEK-293 cells, it significantly attenuated the block of this current by Ni2+ (IC50 increased to 550 μM Ni2+ for CaV3.1 and 15 μM Ni2+ for CaV3.2); in contrast, 30 μM TTX directly inhibited hCaV3.3-induced ICaT and the addition of 750 μM Ni2+ to the TTX-containing medium led to greater block of the current that was not significantly different than that produced by Ni2+ alone. 1 μM STX directly inhibited CaV3.1-, CaV3.2-, and CaV3.3-mediated ICaT but did not enhance the ability of Ni2+ to block these currents. These findings provide important new implications for our understanding of structure–function relationships of ICaT in heart, and further extend the hypothesis of a parallel evolution of Na+ and Ca2+ channels from an ancestor with common structural motifs.

Two distinct low-voltage-activated Ca2+ currents contribute to the pacemaker mechanism in cockroach dorsal unpaired median neurons

1996 ◽  
Vol 76 (2) ◽  
pp. 963-976 ◽  
Author(s):  
F. Grolleau ◽  
B. Lapied
Keyword(s):  
Voltage Dependence ◽  
Low Voltage ◽  
Neurosecretory Cells ◽  
Pacemaker Activity ◽  
Tetraacetic Acid ◽  
Current Clamp ◽  
Patch Clamp Technique ◽  
Time To Peak ◽  
Whole Cell Patch Clamp ◽  
Dorsal Unpaired Median

1. The contribution of Ca2+ currents to the endogenous firing properties of cockroach isolated adult dorsal unpaired median neurons was investigated using whole cell patch-clamp technique with 5 mM Ca2+ as the charge carrier. At least three types of Ca2+ currents, a high-voltage-activated Ca2+ current and two low-voltage-activated (LVA) Ca2+ currents, have been found in these neurons. This study focused on the LVA Ca2+ currents, which are suitable candidates in the generation of the slow predepolarization because of their low threshold of activation. 2. The global LVA Ca2+ current could be dissociated by means of nickel sensitivity, deactivation time constant and voltage dependence of time to peak, tail current amplitude and inactivation, as transient and maintained LVA Ca2+ currents. 3. The transient LVA Ca2+ current, sensitive to 100 microM Ni2+, was isolated by using a subtraction procedure. It was activated at -70 mV and half-inactivated at -59.5 mV. The inactivation was purely voltage dependent. Current-clamp experiments performed with 150 microM Ni2+ indicated that this current was involved in the initial part of the predepolarization. 4. The maintained LVA Ca2+ current, resistant to 100 microM Ni2+, was activated in a range of potential 10 mV more positive than the transient LVA Ca2+ current, and its voltage dependence of inactivation displayed a U-shaped-curve. 5. Replacing Ca2+ with Ba2+ in equimolar amount or low internal Ca2+ concentration [5 mM bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA) in the pipette] induced a monotonic voltage dependence of inactivation and increased the rate of relaxation of this current. These effects were mimicked by high internal Ca2+ concentration [0.1 mM Ca2+ and no ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid in the pipette]. This demonstrated an unusual Ca2+-sensitive inactivation process that varied over a narrow range of Ca2+ concentrations. 6. Current-clamp experiments performed under 150 microM Ni2+, with 15 mM external Ca2+ concentration (which potentiated the maintained LVA current within 30 s of superfusion) or with 5 mM BAPTA in the pipette demonstrated the participation of this current in the last two-thirds of the slow predepolarizing phase. 7. Our findings demonstrated, for the first time in neurosecretory cells, the coexistence of two distinct LVA Ca2+ currents, which have specialized function in the generation of the pacemaker activity.

Electroporation and recovery of cardiac cell membrane with rectangular voltage pulses

1992 ◽  
Vol 263 (4) ◽  
pp. H1128-H1136 ◽  
Author(s):  
O. Tovar ◽  
L. Tung
Keyword(s):  
Cell Membrane ◽  
Time Course ◽  
Low Voltage ◽  
Experimental Studies ◽  
Membrane Conductance ◽  
Cardiac Cells ◽  
Channel Blockers ◽  
Cardiac Cell ◽  
Patch Clamp Technique ◽  
Pipette Solution

Electroporation of the cardiac cell membrane may result from intense electric fields applied to cardiac muscle, associated for example with defibrillation and cardioversion. We analyzed the distribution of voltage levels sufficient to cause electroporation in enzymatically isolated frog cardiac cells, using the cell-attached patch-clamp technique with rectangular pulses similar to those used in experimental studies of cardiac defibrillation. Five-millisecond monophasic or ten-millisecond biphasic symmetric (1/1) and asymmetric (1/0.5) rectangular pulses of either polarity were applied to the cell membrane in 100-mV steps from 0.2 to 0.8 V. The membrane conductance was continuously monitored by a low-voltage pulse train. In a total of 77 cells, we observed a step increase in conductance, occurring in 21% of cells at a transmembrane potential of 0.3 V, 52% at 0.4 V, 14% at 0.5 V, and 13% at 0.6-0.8 V. Electroporation occurred with this voltage distribution regardless of pulse shape, polarity, or the presence of all of the following ionic channel blockers: tetrodotoxin, barium, tetraethylammonium, 4-aminopyridine, cadmium, nickel, and gadolinium. The time course of membrane recovery was highly variable. The maintenance of a high membrane conductance after the shock pulse was associated with irreversible cell contracture provided that Ca2+ was included in the patch-pipette solution. However, with biphasic asymmetric pulses, the conductance recovered very quickly (< or = 37 ms).(ABSTRACT TRUNCATED AT 250 WORDS)

Voltage-Gated Channels and Calcium Homeostasis in Mammalian Rod Photoreceptors

2005 ◽  
Vol 93 (3) ◽  
pp. 1468-1475 ◽  
Author(s):  
David Cia ◽  
Agnès Bordais ◽  
Carolina Varela ◽  
Valérie Forster ◽  
José A. Sahel ◽  
...  
Keyword(s):  
Channel Blockers ◽  
Rod Photoreceptor ◽  
Patch Clamp Technique ◽  
Rod Photoreceptors ◽  
Voltage Gated ◽  
Whole Cell Patch Clamp ◽  
Voltage Gated Channels ◽  
High Concentrations ◽  
Electrophysiological Characterization ◽  
Low Sensitivity

Recent reports on rod photoreceptor neuroprotection by Ca2+ channel blockers have pointed out the need to assess the effect of these blockers on mammalian rods. However, in mammals, rod electrophysiological characterization has been hampered by the small size of these photoreceptors, which were instead extensively studied in nonmammalian vertebrates. To further characterize ionic conductances and to assess the pharmacology of Ca2+ channels in mammalian rods, freshly dissociated pig rod photoreceptors were recorded with the whole cell patch-clamp technique. Rod cells expressed 1) a hyperpolarization-activated inward-rectifying conductance ( Ih) sensitive to external Cs+; 2) a sustained outward K+ current ( IK) sensitive to tetraethylammonium; 3) a sustained voltage-gated Ca2+ current ( ICa) sensitive to benzothiazepine (diltiazem) and phenylalkylamine (verapamil) derivatives; 4) a Ca2+-activated Cl− current ( ICl(Ca)); and 5) a plasma membrane Ca2+-ATPase. The Ca2+ current showed a range of activation from positive potentials to –60 mV with a maximum between –30 and –20 mV. In contrast to other L-type Ca2+ channels, rod Ca2+ channels were blocked at similar and relatively high concentrations by the diltiazem isomers and verapamil. The biphasic dose-response for d-diltiazem confirmed the low sensitivity of Ca2+ channels for the molecule. The ATPase, which was localized at the axon terminal, was found to contribute to Ca2+ extrusion. These results suggest that the electrophysiological features of rod photoreceptors had been preserved during evolution from nonmammalian vertebrates to mammals. This work indicates further that mammalian rods express nonclassic L-type Ca2+ channels, showing a low sensitivity to the diltiazem isomers used in neuroprotective studies.

scholarly journals Ionic Species Involved in the Electrical Activity of Single Adult Aminergic Neurones Isolated from the Sixth Abdominal Ganglion of the Cockroach Periplaneta Americana

1989 ◽  
Vol 144 (1) ◽  
pp. 535-549 ◽  
Author(s):  
BRUNO LAPIED ◽  
CLAIRE O. MALÉCOT ◽  
MARCEL PELHATE
Keyword(s):  
Periplaneta Americana ◽  
Potassium Conductance ◽  
Ionic Species ◽  
Resting Potential ◽  
Channel Blockers ◽  
Patch Clamp Technique ◽  
Mgcl2 Concentration ◽  
Dorsal Area ◽  
Ca2 Channels

Adult neurones were obtained by dissociation of the dorsal area of the sixth abdominal (A6) ganglion of the cockroach, and electrical properties were studied with the patch-clamp technique. The neurones showed spontaneous fast action potentials, similar to those recorded with microelectrodes in neurones in situ along the dorsal median line of the A6 ganglion. Synthetic saxitoxin (sSTX) at concentrations of 10 × 10−8 to 1.0×10−7mol l−1 suppressed the action potential (AP) and induced a dose-dependent hyperpolarization of the resting potential, suggesting that two types of sSTX-sensitive Na+ channels are present. The resting potential was dependent on the external concentration of both Na+ and K+, with a similar sensitivity to each, yielding a slope of about 43 mV per 10-fold change in concentration. The delayed outward rectification present under control conditions was reduced by tetraethylammonium chloride (TEA-Cl, 1.0×10−2mol l−1). TEA-Cl or Ca2+-free saline abolished the afterhyperpolarization and increased the overshoot and duration of triggered APs, indicating that a calcium-activated potassium conductance contributes to the falling phase of the AP. At 3.0×10−3mol l−1, the Ca2+ channel blockers MnCl2, CoCl2 and NiCl2 lengthened the AP. A blocker-dependent increase in the overshoot and threshold of the AP and reduction of the afterhyperpolarization were observed, probably reflecting the relative potencies of these ions in blocking Ca2+ channels and thus the Ca2+- activated K+ conductance. Increasing MgCl2 concentration by 3.0 × 10−3mol l−1 had no effect on the AP, indicating that the positive shift of the threshold is due to the blockade of Ca2+ channels present at this potential. The results suggest that these isolated neurones are dorsal unpaired median neurones previously studied in a number of insect species.

K+ currents responsible for repolarization in mouse ventricle and their modulation by FK-506 and rapamycin

2000 ◽  
Vol 278 (3) ◽  
pp. H886-H897 ◽  
Author(s):  
W. H. DuBell ◽  
W. J. Lederer ◽  
T. B. Rogers
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Regulatory Proteins ◽  
Channel Blockers ◽  
Patch Clamp Technique ◽  
Fk 506 ◽  
Transient Component ◽  
Whole Cell Patch Clamp ◽  
Recovery From Inactivation ◽  
K Currents

Modulation of mouse ventricular action potentials and K+ currents was examined using the whole cell patch-clamp technique. The composite mouse ventricular K+ current (consisted of an outward transient followed by a slowly decaying sustained component. Use of the K+ channel blockers tetraethylammonium and 4-aminopyridine and a transgenic mouse model revealed three pharmacologically and kinetically distinct currents: I to, which contributed to the transient component; I K, which contributed to the sustained component; and a slowly activating current ( I slow), which contributed to both components. The immunosuppressant FK-506 increased action potential duration at 90% repolarization by 66.7% by decreasing the sustained component (−48% at +60 mV) and prolonging recovery from inactivation (by 26% at 200 ms) of the transient component. These effects were isolated to I K and I to, respectively. Rapamycin had strikingly similar effects on these currents. Both FK-506 and rapamycin are known to target the immunophilin FKBP12. Thus we conclude that FKBP12 modulates specific mouse K+ channels, and thus the mouse ventricular action potential, by interacting directly with K+ channel proteins or with other associated regulatory proteins.

Phorbol Ester-Induced Inhibition of Potassium Currents in Rat Sensory Neurons Requires Voltage-Dependent Entry of Calcium

2001 ◽  
Vol 85 (1) ◽  
pp. 362-373 ◽  
Author(s):  
Yi-Hong Zhang ◽  
J. L. Kenyon ◽  
G. D. Nicol
Keyword(s):  
Sensory Neurons ◽  
Phorbol Ester ◽  
Potassium Currents ◽  
Channel Blockers ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Rat Pups ◽  
Whole Cell Patch Clamp ◽  
Potassium Channel Blockers ◽  
Voltage Dependent

The whole cell patch-clamp technique was used to examine the effects of protein kinase C (PKC) activation (via the phorbol ester, phorbol 12,13 dibutyrate, PDBu) on the modulation of potassium currents ( I K) in cultured capsaicin-sensitive neurons isolated from dorsal root ganglia from embryonic rat pups and grown in culture. PDBu, in a concentration- and time-dependent manner, reduced I K measured at +60 mV by ∼30% if the holding potential ( V h) was −20 or −47 mV but had no effect if V h was −80 mV. The PDBu-induced inhibition of I K was blocked by pretreatment with the PKC inhibitor bisindolylmaleimide I and I K was unaffected by 4-α phorbol, indicating that the suppression of I Kwas mediated by PKC. The inhibition of I K by 100 nM PDBu at a V h of −50 mV was reversed over several minutes if V h was changed to −80 mV. In addition, intracellular perfusion with 5 mM bis-( o-aminophenoxy)- N,N,N′,N′-tetraacetic acid (BAPTA) or pretreatment with ω-conotoxin GVIA or Cd2+-Ringer, but not nifedipine, prevented the PDBu-induced suppression of I K at −50 mV, suggesting that a voltage-dependent influx of calcium through N-type calcium channels was necessary for the activation of PKC. The potassium channel blockers tetraethylammonium (TEA, 10 mM) and 4-aminopyridine (4-AP, 3 mM and 30 μM) reduced I K, but only TEA attenuated the ability of PDBu to further inhibit the current, suggesting that the I K modified by PDBu was sensitive to TEA. Interestingly, in the presence of 3 mM or 30 μM 4-AP, 100 nM PDBu inhibited I K when V h was −80 mV. Thus 4-AP promotes the capacity of PDBu to reduce I K at −80 mV. We find that activation of PKC inhibits I K in rat sensory neurons and that voltage-dependent calcium entry is necessary for the development and maintenance of this inhibition.

Temperature-sensitive intracellular Mg2+ block of L-type Ca2+ channels in cardiac myocytes

2002 ◽  
Vol 282 (3) ◽  
pp. H1092-H1101 ◽  
Author(s):  
Kaoru Yamaoka ◽  
Tsunetsugu Yuki ◽  
Kayoko Kawase ◽  
Makoto Munemori ◽  
Issei Seyama
Keyword(s):  
Cardiac Myocytes ◽  
Late Phase ◽  
Slow Phase ◽  
Temperature Sensitive ◽  
Patch Clamp Technique ◽  
Rapid Phase ◽  
Channel Current ◽  
Whole Cell Patch Clamp ◽  
Blocking Effects ◽  
Ca2 Channels

We examined the concentration-dependent blocking effects of intracellular Mg2+ on L-type Ca2+ channels in cardiac myocytes using the whole cell patch-clamp technique. The increase of L-type Ca2+ channel current ( I Ca) (due to relief of Mg2+ block) occurred in two temporal phases. The rapid phase (runup) transiently appeared early (<5 min) in dialysis of the low-Mg2+ solution; the slow phase began later in dialysis (>10 min). Runup was not blocked by intracellular GTP (GTPi). The late phase of the I Ca increase (late I Ca) was suppressed by GTPi (0.4 mM) and was observed in myocytes of the guinea pig or frog at higher (32 or 24°C, respectively) rather than lower temperatures (24 or 17.5°C, respectively). At pMg = 6.0, raising the temperature from 24 to 32°C evoked late I Ca with a Q10 of 14.5. Restoring the temperature to 24°C decreased I Ca with a Q10 of only 2.4. The marked difference in the Q10 values indicated that late I Ca (pMg = 5–6) is an irreversible phenomenon. Phosphorylation suppressed the intracellular [Mg2+] dependency of late I Ca. This effect of phosphorylation together with the inhibitory action of GTPi on Mg2+-dependent blocking of I Ca are common properties of mammalian and amphibian cardiomyocytes.

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