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A Tale of Two (Calcium) Channels
Joël Nargeot
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properties of ICaT to those of the recombinant calcium
currents generated by ␣1G and ␣1H expressed in human
embryonic kidney cells. Most of the investigations are devoted to a comparison of the biophysical properties between
ICaT from AT-1 cells and ␣1G/␣1H currents. The results
indicate that most basic electrophysiological properties, such
as current-voltage relationship and activation, inactivation,
and deactivation properties, are rather similar and not discriminative. However, the recovery from inactivation of
T-type currents, characterized by the sum of a fast and a slow
time constant, is described as a functional signature of T-type
channel isoform expression. The major difference between
the two T-type channel isoforms relates to the relative
amplitude of the slow recovery rate (␶s) as a strong functional
criterion to establish a linkage between ICaT from AT-1 cells
and ␣1G current. Such linkage is confirmed by reverse
transcription–polymerase chain reaction (RT-PCR) experiments from cultured AT-1 cells using primers designed to
amplify the III-IV loop of the three isoforms. The results
show that only ␣1G, and predominantly a specific variant
thereof, is revealed from sequencing of the subcloned PCR
product corresponding to a single major band.
The results presented on AT-1 cells are convincing, but it
is reasonable to wonder whether they can be extended to any
cardiac cell. Comparing the properties of native ICaT in
cardiovascular cells with those of recombinant currents may
be difficult because many studies have been conducted either
under different experimental conditions or in different species. Cardiac T-type currents have often been studied in
cultured embryonic or neonatal myocytes because of their
low expression (or absence) in adult atrial (or ventricular)
myocytes, except in several species such as chicken and
guinea pig. Other studies report the characterization of
cardiac ICaT induced by hormone treatment9 or pathology.10,11 On the other hand, T-type channel isoforms have also
been cloned from different species, and the use of various
external concentrations of calcium or barium ions to study
their related currents sometimes confounds the comparisons
between native and recombinant currents. Satin and Cribbs8
provide a comparison between the properties of the recombinant rat (␣1G) and human (␣1H) currents and ICaT from
AT-1 (mouse) cells in a physiological calcium concentration.
However, a large number of splice variants have been
described for ␣1G in the different species3 and as discussed
by the authors, it cannot be ruled out that splice variations in
regions other than the III-IV loop may influence biophysical
properties including recovery from inactivation. All of these
splice variations involve connecting loops or the C-terminus.
Curiously, nickel sensitivity was not investigated by Satin
and Cribbs,8 even though it is considered an important assay
to distinguish between the two isoforms because of the much
higher Ni2⫹ sensitivity of ␣1H currents (IC50 5 ␮mol/L)
alcium influx through voltage-dependent calcium
channels triggers excitation-contraction coupling and
regulates pacemaking activity in the heart. Two
distinct families of calcium channels have been identified in
cardiac tissue: L-type calcium channels, which are essential
in triggering Ca2⫹ release from internal stores, and lowvoltage-activated (LVA) T-type calcium channels (ICaT),
whose role remains obscure in physiological and pathophysiological conditions. Functional features of ICaT include low
threshold of activation, small unitary conductance, slow
activation, and fast inactivation inducing a typical crisscrossing pattern of current traces for increasing depolarizations, negative steady-state inactivation, and slow deactivation kinetics. In addition, T-type currents compared with
L-type currents are more sensitive to block by mibefradil and
Ni2⫹ ions. Whereas L-type calcium channels have been
extensively characterized at the functional and molecular
levels, classical cloning strategies failed to identify an ␣1
subunit encoding for a T-type channel.
An alternative approach to identifying new members of the
calcium channel family used in silico cloning strategies with
a search of genetic databases for sequences homologous but
not identical to known Ca2⫹ channel ␣1 subunits.1 The
identification of several expressed sequence tags and
genomic sequences corresponding to a subset of distantly
related ␣1 subunits resulted in the identification of full-length
cDNAs encoding three distinct ␣1 subunits: ␣1G in rat,1
mouse,2 and human,3,4 ␣1H in human,5,6 and ␣1I in rat.7
Because ␣1H was obtained by screening a human heart
library,5 it was originally considered the cardiac T-type
channel isoform because of the presence in Northern blot
analysis of a strong signal for ␣1H in the heart. However, an
␣1G transcript is also detected in adult heart, and it has
become an interesting challenge to identify which isoforms
underlie ICaT in cardiac cells.
The article by Satin and Cribbs8 in this issue of Circulation
Research aims to identify the ␣1 isoform encoding the ICaT in
AT-1 cells, an immortalized cell line derived from mouse
atrial tissue. The advantages of using this cell line after a
short period of culture are the absence of sodium current and
the relatively small amplitude of L-type current, allowing for
a better isolation of ICaT. Satin and Cribbs have compared the
The opinions expressed in this editorial are not necessarily those of the
editors or of the American Heart Association.
From the Institut de Genetique Humaine, Montpellier, France.
Correspondence to Joël Nargeot, Institut de Genetique Humaine,
CNRS UPR 1142, 141 rue de la Cardonille, 34396-Montpellier cedex 5,
France. E-mail [email protected]
(Circ Res. 2000;86:613-615.)
© 2000 American Heart Association, Inc.
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March 31, 2000
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versus ␣1G currents (IC50⬎150 ␮mol/L).12,3 Preliminary data
revealed a low Ni2⫹ sensitivity (160 ␮mol/L) of cardiac
T-currents in freshly dissociated atrial myocytes from neonatal rat cells, also suggesting a linkage with the ␣1G isoform.13
However, previous data in rabbit sinoatrial,14 adult guinea pig
cells,15 and rat hypertrophic cells11 indicate that T-currents
are totally blocked by a lower concentration of Ni2⫹ ions
(about 40 ␮mol/L). This would suggest either a differential
expression of ␣1G and ␣1H isoforms among cardiac tissues,
interspecies differences, or a possible developmental switch
between isoforms. The latter hypothesis appears consistent
with the results of Monteil et al,3 who observed in a dot-blot
analysis a developmental regulation of human ␣1G transcripts with a prominent signal in embryonic compared with
the adult heart. It must be emphasized, in agreement with
such a hypothesis, that no ICaT has yet been recorded in adult
human heart.16 Another study worth mentioning used an
antisense strategy, which suggested that the cardiac T-type
current in 3-week-old rats is related to the ␣1E subunit.17 This
result remains puzzling because some properties such as
threshold of activation and deactivation properties differ
markedly between ICaT and ␣1E currents. In addition, ICaT
from freshly dissociated neonatal rat atrial tissue was found13
to be insensitive to the ␣1E-specific toxin SNX 482. It is,
however, worthwhile to note that T-type channel expression
was induced by growth hormone treatment in the experiments
of Piedras-Renteria et al.17
By contrast, with the absence of T-type current in adult
human heart, an LVA calcium current was first reported in
human atria,18 sharing some typical properties with sodium
channels such as TTX sensitivity (designated ICaTTX). Its
description in rat19 and guinea pig20 myocytes indicated that
it is not exclusively observed in diseased human cells. The
study of Heubach et al21 in this issue of Circulation Research
demonstrates for the first time the coexistence of ICaTTX with
ICaT in guinea pig ventricular myocytes. The results show
that adult rat myocytes lack ICaT whereas adult guinea pig
myocytes express both LVA currents that can be isolated by
pharmacological dissection using TTX and Ni2⫹ in the absence of external sodium ions. As mentioned above, T-type
currents are reported here to be blocked by 40 ␮mol/L Ni2⫹
ions (IC50 16 ␮mol/L), a concentration much lower than that
required to block recombinant ␣1G currents but rather close
to that reported to block ␣1H currents. There are major
differences between the biophysical properties of ICaT and
ICaTTX. ICaTTX exhibits a run-up after the rupture of the patch
membrane, a lower voltage for peak current (10 mV), a more
negative steady-state inactivation relationship, faster time
constants for recovery from inactivation, and a faster rate of
deactivation. Interestingly, ICaTTX was blocked by mibefradil,
as is ICaT.
The remaining question concerns the molecular basis of
ICaTTX. Is this current related to modified sodium channels in
the absence of external sodium ions20 or is it due to a new
population of channels? Answering this will require the
molecular identification of the pore-forming subunit using
various strategies, including antisense, expression cloning,
and other conventional molecular and biochemical isolation
techniques. There is a lack of evidence for a correlation
between ICaTTX and the controversial slip-mode conductance
that has been described for sodium channels in the presence
of protein kinase A (PKA) and cardiotonic steroids.22 The fact
that ICaTTX does not require activation of the PKA pathway
argues for a distinct population of channels. Additional
experiments to test the effect of PKA inhibitors on ICaTTX
would be of interest. Heubach et al21 underline some discrepancies between the TTX concentration required to block
modified sodium channels and ICaTTX. In addition, the work
of Lemaire et al18 on human atrial cells showed that ICaTTX
similarly conducts calcium and barium ions. Permeation to
barium ions was not tested on the modified sodium channels,
but most of the conclusions were based on calcium transient
measurements. This property seems atypical for sodium
channels but might indicate either a strong alteration of the
sodium channel permeability in the absence of extracellular
sodium ions or molecular similarities between ICaT and
ICaTTX.
From the results presented in this issue of Circulation
Research and other studies, it is now clear that two LVA
channels (ICaT and ICaTTX) can coexist in cardiac myocytes.
What is the physiological role of these LVA channels in the
heart? T-type channels are assumed to play a role in pacemaking activity because of their presence in sinoatrial node
and the negative chronotropic effect of Ni2⫹ ions.14 Whether
ICaTTX is also expressed in sinoatrial cells is unknown, and it
would also seem important to reinvestigate the role of LVA
versus L-type calcium channels in the pacemaking activity.
An interesting feature of T-type calcium currents is related to
their slow deactivation kinetics and the existence of a window
current in the range of the cell membrane resting potential.
Slow deactivation can mediate larger calcium influx than
high-voltage-activated channels during short depolarizations,
as shown by the application of a neuronal-type action
potential such as a voltage-clamp command on recombinant
␣1G channels,3 the calcium transients in response to different
spike shapes or frequencies also being isoform dependent.23
In spite of their rapid kinetics of inactivation, the existence of
a window current would confer a role of T-type channels in
maintaining intracellular Ca2⫹concentration. T-type currents
do not seem to play an important role in cardiac excitationcontraction coupling,24 but they could contribute to finetuning of basal calcium levels and control physiological
processes such as hormone secretion, as previously suggested
in adrenal cells.25 Their expression is also cell cycle dependent26 and their involvement in cell growth and proliferation
was suggested in cardiac9 and smooth muscle cells27,28
respectively. In contrast to ICaT, ICaTTX is observed in human
heart and is likely to be sensitive to mibefradil as found in
guinea pig myocytes.
Future studies will probably look for an overexpression of
ICaT and ICaTTX in pathological conditions, which would be
expected to generate cardiac arrhythmias. Interestingly, it was
recently reported that mibefradil prevents tachycardiainduced electrophysiological remodeling in dogs.29 Thus,
inhibition of both LVA currents by mibefradil might be
considered in terms of pathophysiology. Combining specific
functional assays and molecular tools should allow a better
Nargeot
understanding of the physiological and pathological roles of
LVA channel isoforms in cardiovascular cells.
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KEY WORDS: Ca2⫹ channels
䡲 T-type channel 䡲 cardiac muscle
A Tale of Two (Calcium) Channels
Joël Nargeot
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Circ Res. 2000;86:613-615
doi: 10.1161/01.RES.86.6.613
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Copyright © 2000 American Heart Association, Inc. All rights reserved.
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