© 1996 American Heart Association, Inc.
Articles |
Developmental Changes in ß-Adrenergic Modulation of L-Type Ca2+ Channels in Embryonic Mouse Heart
From the Department of Physiology (R.H.A., M.P.D., R.B., R.S.K.), University of Rochester (NY) Medical Center, and the Department of Medicine (P.A.D., S.W.K., K.R.C.), University of California at San Diego.
Correspondence to Dr R.S. Kass, Department of Physiology, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642-8642. E-mail rsks@uhura.cc.rochester.edu.
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and Methods Results Discussion References |
|---|
Abstract In the adult mammalian myocardium, cellular Ca2+ entry is regulated by the sympathetic nervous system. L-type Ca2+ channel currents are markedly increased by ß-adrenergic (ß-A) agonists, which contribute to changes in pacing and contractile activity of the heart. In the developing mammalian heart, the regulation of Ca2+ entry by this enzyme cascade has not been clearly established, because changes in receptor density and coupling to downstream elements of the signaling cascade are known to occur during embryogenesis. In this study, we systematically investigated the regulation of L-type Ca2+ channel currents during development of the murine embryonic heart. We used conventional whole-cell and perforated-patch–clamp procedures to study modulation of L-type Ca2+ channel currents and to assay functional activity of distinct steps in the ß-A signaling cascade in murine embryonic myocytes at different stages of gestation. Our data indicate that L-type Ca2+ channels in early-stage (day-11 to -13) myocytes are unresponsive to either isoproterenol or cAMP. L-type Ca2+ channels in late-stage (day-17 to -19) murine myocytes, however, exhibit responses to isoproterenol and cAMP similar to responses in adult cells, providing evidence that the ß-A cascade becomes functionally active during this period of embryonic development. We found that L-type Ca2+ channel activity in early-stage cells is increased by cell dialysis with the catalytic subunit of cAMP-dependent protein kinase (cA-PK) and that dialysis of early-stage cells with the holoenzyme of cA-PK restores functional responses to forskolin and cAMP, but not to isoproterenol. Our results provide strong evidence that a key factor in the early-stage insensitivity of L-type Ca2+ channels to cAMP is the absence, or low expression level, of the holoenzyme of cA-PK but that in addition, another element in the signaling cascade upstream from adenylate cyclase is expressed at a nonfunctional level or is uncoupled from the cascade and thus contributes to L-type Ca2+ channel insensitivity to ß-A agonists in early stages of the developing murine heart.
Key Words: embryonic mouse heart • development • Ca2+ channels • cAMP • protein kinase A
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and Methods Results Discussion References |
|---|
Voltage-gated L-type Ca2+ channels are physiologically important in many excitable cells but are particularly important in the heart, because Ca2+ entry through these channels not only contributes to impulse generation and conduction but also serves as a second messenger to modulate regulatory protein kinases and the activation of contractile proteins and very likely contributes to the control of gene expression.1 2 In addition, it is well established in the adult mammalian heart that Ca2+ entry via L-type Ca2+ channels is regulated by the ß-A signaling pathway. The binding of agonist to ßAR is coupled to an intracellular cascade by the stimulatory G protein, Gs. Agonist occupancy activates AC to increase intracellular concentrations of cAMP, which, in turn, activates cA-PK by dissociating the inactive holoenzyme into regulatory and catalytic subunits (summarized in Fig 8
). In adult cardiac myocytes, this sequence increases L-type
channel
activity, in part through changes in gating caused by
phosphorylation of the channel or by a closely
associated protein,
CS-cA-PK,3 4 5 6 7
and through direct
membrane-delimited G-protein interactions with the channel
protein.8 The L-type Ca2+ channel is a
multisubunit complex consisting of the primary pore-forming
1 subunit along with additional accessory
2/
, 
, and ß2
subunits.9 It has been reported that L-type channel
modulation by cA-PK is enhanced by coexpression of 1 and
ß2 subunits in heterologous expression
systems10 11 but that the 1 subunit
alone
can be the target of cA-PK.12
|
During development, modulatory responses to catecholamines
can be affected by changes in the relative expression levels of ß-A
signaling cascade components and/or the type and architecture of L-type
channel subunits. There is evidence in the literature for
developmentally mediated alternative splicing of the cardiac L-type
channel 1 subunit13 and considerable
evidence for changes in the cAMP signaling pathway that take place
during early stages of development in the hearts of several
species.14 It has been observed in most species, the mouse
in particular, that the physiological (ie, positive
inotropic and chronotropic) responses to ß-A stimulation of the
developing heart lag behind the expression of ßAR and
AC.15 16 17 Furthermore, in the murine
embryonic heart, total
cA-PK activity increases most markedly during the 6 days before birth,
a period in which the physiological response to
ß-A stimulation also becomes apparent.18 Similarly, in
the fetal rat heart during late embryonic stages, there is improved
coupling between ßAR and physiological
responses.19 Thus, it was the aim of the present study
to examine in detail the modulation of L-type Ca2+ channel
activity by ßAR stimulation in the mouse embryonic heart and in order
to test for possible changes in the ß-A signaling cascade that may
occur during embryogenesis and affect the modulation of
Ca2+ entry by circulating catecholamines and,
in turn, contribute to the function of, and control of gene expression
in, the developing heart.20
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and Methods Results Discussion References |
|---|
Embryonic Cardiomyocyte Isolation and Culture
Embryos were removed from pregnant female mice either at 11 to 13 days (early stage) or 17 to 20 days (late stage) after coitus as described by Sturm and Tam.21 Cardiac myocytes were isolated from embryonic hearts as previously described.22 In brief, hearts were dissected from embryos and placed in ADS buffer (mmol/L: NaCl 116, HEPES 20, NaH2PO4 1.0, glucose 5.5, KCl 5, and MgSO4 0.8, pH 7.35). Atrial and ventricular tissues were separated under a dissecting microscope and placed in an Eppendorf tube with 0.5 mL ADS buffer containing 0.5 mg/mL collagenase type II (Worthington) and 1.0 mg/mL pancreatin (GIBCO) for a 15-minute digestion at 37°C. Cells from the enzymatic digestion were placed in culture medium containing 10% fetal bovine serum, plated on gelatin- and laminin-treated glass or untreated plastic coverslips, and cultured in a 10% CO2 incubator.
Electrophysiology
Experimental results described in the
present study were
obtained using patch-clamp procedures in the conventional
whole-cell23 or perforated-patch24
configuration. Intracellular and extracellular solutions and voltage
protocols designed to emphasize L-type Ca2+ channel
currents have been described previously.25 In order to
measure the time course of regulatory responses, we measured L-type
Ca2+ channel currents with the following protocol, which is
referred to in the text as the "train protocol": from a
-40-mV
holding potential, currents were measured during test pulses (40
milliseconds) to +20 mV applied once every 10 seconds. The same train
protocol was used in all time-course experiments. Patch pipettes
(Clay Adams glass) were pulled to resistances of 2.5 to 5.0 M
when
filled with intracellular solutions. Total cell membrane
capacitance was used as a measurement of membrane area and was
determined either by analog capacity compensation or by integration of
current transients in response to 10-mV test pulses.
In dialysis
experiments, cells were patch-clamped with pipettes
filled with internal solution containing CS-cA-PK or holo-cA-PK at
concentrations indicated in the figure legend. Low-resistance
pipettes (0.5 to 2 M) were used to minimize access resistance and
speed diffusion of enzymes throughout the cell. Diffusion times were
estimated from cell capacity, access resistance, and molecular weight
of the enzymes according to the calculations of Pusch and
Neher.26 Cells were thus dialyzed for 10 minutes before
testing for drug effects when the intracellular cA-PK concentration was
calculated to be 
98% of that of the pipette solution.
For perforated-patch recordings, nystatin was dissolved in methanol at a concentration of 50 mg/mL and then added to the standard internal solution to yield a final concentration of 100 µg/mL. Both the nystatin stock solution and the nystatin-containing pipette solution were subjected to 5 to 10 minutes of ultrasonication before use. Capacity transients were monitored as a function of time after attaining a high-resistance seal with the surface membrane. Electrical access to the cell was judged by the time course of the capacity transient, and adequate access was usually attained within 10 minutes of seal formation.
Chemicals were obtained from the following suppliers: isoproterenol and nystatin, from Sigma Chemical Co; 8-CPT-cAMP, from Boehringer Mannheim; and purified catalytic subunit of cAMP-dependent protein kinase, from Promega. Bay K 8644 and nisoldipine were gifts from Miles Pharmaceuticals. Fresh solutions were prepared daily.
Data were collected, stored, and analyzed on IBM (486)-compatible computers interfaced to a Yale Mark IV amplifier constructed in our laboratory or an Axopatch 200A amplifier (Axon Instruments) under the control of PCLAMP software (Axon Instruments). Graphics and statistical data analysis were carried out using ORIGIN software (Microcal). Averaged data are shown as mean±SEM.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and Methods Results Discussion References |
|---|
Rundown of Whole-Cell Currents: Evidence for Regulation
As previously reported by us, L-type Ca2+ channel currents were readily identified and measured using conventional whole-cell patch-clamp procedures in murine cardiac cells at stages as early as days 11 to 13.27 However, currents measured using this procedure rapidly declined in amplitude after rupturing the cell membrane to establish whole-cell recording conditions similar to L-type Ca2+ channel currents in adult cells, in which regulation by the ß-A pathway clearly has been established. Therefore, as expected, we found that rundown of L-type channel current was prevented by using the perforated-patch–clamp technique to avoid unnecessary cell dialysis that might disrupt this signaling pathway (Fig 1
).
Thus, we used perforated-patch measurements in
most experiments that follow in which external application of an
agonist or a membrane-permeable cAMP analogue was used. Fig 2
shows that by using this recording technique,
clear modulation of early-stage Ca2+ channel current
could be demonstrated for both the dihydropyridine
agonist Bay K 8644 and the blocker nisoldipine (Fig 2),
confirming the
channels as L-type. Bay K 8644 application nearly doubles the current
amplitude, and subsequent exposure to nisoldipine almost completely
inhibits channel activity.
|
, n=6) and perforated-patch (
, n=6) recording
conditions are plotted against time of recording. Time zero was
marked at the time of rupture of the membrane patch for whole-cell
recordings and the time at which the cell capacity transient
was 
1 millisecond for perforated-patch recordings. The
insets show representative current traces at the times
indicated in the figure recorded with conventional whole-cell
(a) and perforated-patch (b) procedures (calibration bars indicate
6 pA/pF and 10 milliseconds). In all cases, the holding potential was
-40 mV, and the test potential was +20 mV.
***P<.001.
|
) is plotted
against test potential (n=3). Bay K 8644 increased peak current in a
voltage-dependent manner.
Response to Isoproterenol in Late- but Not Early-Stage
Cells
Thus, particularly in view of channel rundown using conventional
whole-cell procedures, we were surprised to find that L-type
channels expressed in early-stage cells were unresponsive to
application of the ßAR agonist isoproterenol (3 µmol/L), as shown
in Fig 3a. However, L-type channel currents recorded
in late-stage cells consistently were enhanced by the same
isoproterenol concentration (Fig 3b), indicating that changes
in ß-A
regulation of L-type channels in fact occur between days 11 and 20 in
the embryonic mouse heart. The developmental change in L-type channel
response to isoproterenol could be due to changes in expression levels
and/or coupling of one or more components of the ß-A signaling
pathway. It is also possible that the lack of response to isoproterenol
could be a result of changes in the expression levels and/or assembly
of individual subunits of the
heteromultimeric L-type channel protein. The next
set of experiments was designed to distinguish between these
possibilities and identify which, if any, component of the ß-A
signaling cascade changes during early developmental stages in the
murine embryonic heart.
|
Response to cAMP in Late- but Not Early-Stage Cells
Because
it has been reported that expression of ßAR is low in
the early-stage (day-13) embryonic mouse heart,15 the
contrast between the responsiveness of early- and late-stage L-type
channel currents to isoproterenol may simply reflect a developmental
change in ßAR and/or G-protein expression. The experiments shown in
Figs 4 and 5 were designed to test for
this possibility. In these figures, experiments are summarized in
which the ßAR is bypassed, and the direct effects of cytosolic cAMP
levels are tested on L-type channel activity for both early- and
late-stage embryonic murine myocytes.
|
|
Fig 4
shows that
L-type channel activity in late- but not
early-stage cells is sensitive to changes in intracellular cAMP. In
this experiment, cells were exposed to 8-CPT-cAMP (200 µmol/L), a
membrane-permeable cAMP analogue that has previously been shown to
enhance adult L-type channel activity in guinea pig
myocytes.29 Fig 5 summarizes the rate of onset of
and
recovery from exposure to 8-CPT-cAMP in early- and late-stage cells
for a large number of experiments. L-type channel currents measured in
early-stage (day-11 to -13) cells were not responsive to
8-CPT-cAMP. Neither increasing the 8-CPT-cAMP concentration to 500
µmol/L (2 of 2 cells) nor challenging the cells with the
membrane-permeable cAMP analogue 8-bromo-AMP (300 µmol/L) (2 of 2
cells) enhanced L-type channel currents in early-stage cells (data
not shown). However, L-type Ca2+ currents in late-stage
(day-17 to -20) cells are enhanced by 8-CPT-cAMP (200 µmol/L) (Fig
5). The response in late-stage cells reaches steady state
within 1
to 2 minutes of cell exposure, and full recovery is obtained within 2
minutes of returning to 8-CPT-cAMP–free extracellular solutions. We
found an L-type channel response to cAMP in only 1 (6.25%) of 16
early-stage cells and 3 (37.5%) of 8 intermediate-stage cells
but in 9 (81.82%) of 11 late-stage cells. The results of these
experiments suggest that a limiting step in the developmental change in
L-type channel responsiveness to isoproterenol is downstream from ßAR
or G-protein expression (Fig 8). It could either be a function
of
changes in expression of the L-type channel itself and the regulatory
domains on it or a reflection of changes in the expression of
cA-PK.
Channels in Early-Stage Cells Respond to the Catalytic Subunit
of cA-PK
In order to distinguish between these possibilities, we
carried
out experiments in which we dialyzed cells with solutions containing
CS-cA-PK. Because cell dialysis was necessary, we could not use the
perforated-patch procedure but instead used the conventional
whole-cell arrangement of the patch clamp. Channel rundown, as
illustrated in Fig 2, was apparent, and challenging cells with
membrane-permeable 8-CPT-cAMP did not alter the time course or
magnitude of L-type channel rundown (Fig 6a), as
expected from the results of our perforated-patch experiments. This
is in contrast to the effects of dialysis with CS-cA-PK, which clearly
slows and almost eliminates channel rundown (Fig 6b). After a
5-minute
dialysis period, there is a fourfold increase in the amplitude of
currents measured in the presence of CS-cA-PK compared with control
currents measured at the same time after establishing whole-cell
mode (arrows in figure). Thus, the L-type channel can be modulated by
CS-cA-PK, but not 8-CPT-cAMP, at this early embryonic stage (days 11 to
13). This indicates that the channel protein is expressed with cA-PK
phosphorylation sites intact at this period of
development and that the lack of responsiveness to 8-CPT-cAMP and
isoproterenol is probably partially due to a deficiency of cA-PK in the
early developmental stage. We reasoned that if this were the case, it
should be possible to reconstruct the signaling cascade by dialysis
with holoenzyme.
|
Early-Stage Cell Dialysis With Holo-cA-PK Restores Early-Stage
L-Type Channel Sensitivity to cAMP but Not Isoproterenol
We attempted
to reconstruct the ß-A signaling pathway by
dialyzing early-stage cells with holo-cA-PK and then
challenging cells at different steps of the cascade. Fig 7
summarizes the results of these experiments and shows
responses of L-type channel current in these cells to 8-CPT-cAMP,
forskolin, and isoproterenol after cell dialysis with holo-cA-PK
("Materials and Methods"). After holo-cA-PK dialysis, both
8-CPT-cAMP (Fig 7a, 4 of 4 cells) and forskolin (Fig
7b, 3 of 3 cells)
enhance early-stage cell L-type channel currents. These results
confirm that cA-PK is a limiting factor in the
phosphorylation cascade of early-stage cells. In
addition, the L-channel response to forskolin, a direct AC
activator, indicates that there is detectable activity of
AC in early-stage cells, although this effect is submaximal, as
evidenced by the further enhancement of current by subsequent exposure
to 8-CPT-cAMP (Fig 7b). Despite this evidence for functional
activity
of AC and cAMP after dialysis with cA-PK, L-type channel currents were
still insensitive to isoproterenol (Fig 7c, 4 of 4 cells).
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and Methods Results Discussion References |
|---|
To our knowledge, this is the first systematic investigation of developmental changes in ß-A regulation of L-type Ca2+ channels in the embryonic mouse heart. In the present study, modulation of L-type Ca2+ channel activity was used as a functional assay to test the physiological activity of different steps of the ß-A enzymatic cascade in embryonic murine heart. Our finding that there is a marked change in the responsiveness of L-type channels to ßAR stimulation during development of the mouse heart was at first surprising because of previous reports of robust responsiveness of L-type Ca2+ channels in neonatal mouse myocytes30 and stem cell–derived cardiocytes31 32 to isoproterenol. However, the absence of a modulatory response of L-type Ca2+ channel current to isoproterenol in early-stage cells clearly suggests that one or more steps in the signal transduction pathway is not functionally active at this stage of development of the mouse heart.
We took advantage of cell dialysis of conventional whole-cell
patch-clamp procedures to identify nonfunctional cascade
components. Fig 8 summarizes the well-established
signaling cascade in order to aid in the discussion that follows. The
fact that in the absence of cA-PK dialysis, membrane-permeable cAMP
analogues 8-CPT-cAMP and 8-bromo-cAMP did not increase L-type
Ca2+ channel current suggests that one of the limiting
factors might be either cA-PK or the structure of the channel protein
itself. Because the Ca2+ channel 1 subunit
alone can be a substrate for protein kinase A12 but L-type
channel sensitivity to cA-PK is likely to be enhanced by the ß
subunit,10 11 the developmental change in L-type
channel
modulation by isoproterenol could be due to changes in expression
levels and/or assembly of individual subunits of the
heteromultimeric channel protein. However, our
subsequent experiments clearly rule out the possibility that channel
assembly and/or channel expression accounts for the insensitivity to
cAMP. Our results clearly show that cell dialysis with cA-PK restores
L-type channel sensitivity to cAMP and thus provide strong evidence
that channel subunit expression and/or assembly does not underlie the
absence of modulatory responses in early-stage cells. The data,
instead, suggest that a limiting factor in early-stage cells is
most likely the level of cA-PK expression during early stages of
embryogenesis.
Biochemical studies have provided evidence that the activity of cA-PK changes during the same developmental period for which we here report changes in channel regulation based on electrophysiological measurements. cA-PK activity has been measured in an age-dependent manner in embryonic and adult mice.18 The results of this study showed that in the mouse heart, total cardiac protein kinase activity increases very steeply during the 6 days before birth. The maximum kinase level is achieved in the 7-day-old neonatal mouse. The earliest developmental stage investigated in this previous work was an intermediate stage (day 14) of gestation, in which cA-PK activity was found to be <20% of maximum adult levels. Thus, it is very likely that in the early-stage (day-11 to -13) cells investigated in the present study, the cA-PK activity is <20% of that in adult cells.
Our finding that cell dialysis with cA-PK restores functional modulation of L-type channels by cAMP and forskolin, a direct stimulator of AC, provides clear evidence that a contributing factor in the early-stage insensitivity of L-type channels to the ß-A agonist isoproterenol is the level of cA-PK expression at this stage of embryogenesis. However, the persistent insensitivity of L-type channels to isoproterenol even after cA-PK dialysis provides evidence for incomplete development of another step in the signaling cascade that precedes AC: either expression of the ßAR or the Gs protein, which couples the receptor to the signaling cascade in adult cells.
Chen et al15 used agonist-displaceable [3H](-)dihydroalprenolol binding to measure ßAR density and identified receptor expression in embryonic mouse heart before detection of an agonist-induced positive chronotropic effect. This suggested that some additional step in the ß-A signaling cascade was not yet fully developed at early stages of embryogenesis. Subsequent studies by the same group using the same technology in combination with measurements of adenylate cyclase activity confirmed expression of ßAR and AC in the mouse heart as early as gestation day 13, again at a stage that precedes the positive chronotropic effect of ßAR agonists.16 Similar results have been reported in chick14 and rat17 embryonic heart. More recently, Slotkin et al,19 working with fetal rat heart, have reported developmental changes in the coupling between ßAR and AC activity via G proteins during embryogenesis. Functional coupling is not fully established at gestational day 12 but is intact by gestational day 18. Taken together, this previous work, in combination with our results, suggests Gs-protein uncoupling of ß-A from downstream elements in the signaling cascade in early-stage murine embryonic cardiac cells.
The results that we report are directly relevant to regulation of L-type Ca2+ channels in the developing mammalian heart, but they are also relevant to at least one form of human heart disease. In chronic heart failure, there is a reduced responsiveness to ßAR agonists, which may contribute to a reduction in contractile activity due in part to downregulation and sequestration of receptors33 and in part to uncoupling of receptors from downstream steps in the signaling cascade, which is mediated by ßAR kinase and ß-arrestin.34 35 36 37 38 In addition, dilated cardiac hypertrophy, a hallmark of heart failure, is accompanied by reactivation of genes that are expressed in fetal heart development,39 40 suggesting that understanding fetal programming of the ß-A cascade, including interactions with L-type Ca2+ channels, is likely to provide valuable insight into processes contributing to mortality due to these diseases. Coupled with recent advances in genetically modified mice, in which specific steps in the ßAR system are specifically targeted in the heart,41 42 the present data should provide an important baseline that can be used to test directly the functional consequences of disruption of specific steps in the ß-A pathway in the fetal heart.
|
Selected Abbreviations and Acronyms |
|---|
|
|
Acknowledgments |
|---|
This study was supported by USPHS grant 1RO1-HL21922-18.
Received July 11, 1995; accepted November 27, 1995.
|
References |
|---|
|
TopAbstractIntroductionMaterials and Methods Results Discussion References |
|---|
1. Kass RS. Ionic basis of electrical activity in the heart. In: Sperelakis N, ed. Physiology and Pathophysiology of the Heart. 3rd ed. Norwell, Mass: Kluwer Academic Publishers; 1994.
2. Tsien RW, Ellinor PT, Horne WA. Molecular diversity of voltage-dependent Ca channels. Trends Pharmacol Sci. 1991;12:349-354. [Medline] [Order article via Infotrieve]
3. Kameyama M, Hescheler J, Hofmann F, Trautwein W. Modulation of Ca current during the phosphorylation cycle in guinea-pig heart. Pflugers Arch. 1986;407:123-128. [Medline] [Order article via Infotrieve]
4.
McDonald TF, Pelzer S, Trautwein W, Pelzer DJ.
Regulation and modulation of calcium channels in cardiac,
skeletal, and smooth muscle cells. Physiol Rev. 1994;74:365-507. Review.
5. Herzig S, Patil P, Neumann J, Staschen CM, Yue DT. Mechanisms of ß-adrenergic stimulation of cardiac Ca2+ channels revealed by discrete-time Markov analysis of slow gating. Biophys J. 1993;65:1599-1612. [Medline] [Order article via Infotrieve]
6.
Yue DT, Herzig S, Marban E. ß-adrenergic
stimulation of calcium channels occurs by potentiation of
high-activity gating modes. Proc Natl Acad Sci
U S A. 1990;87:753-757.
7.
Reuter H, Kokubun S, Prod'hom B. Properties
and modulation of cardiac calcium channels. J
Exp Biol. 1986;124:191-201.
8. Brown AM. Membrane-delimited cell signaling complexes: direct ion channel regulation by G proteins. J Membr Biol. 1993;131:93-104. Review.[Medline] [Order article via Infotrieve]
9.
Catterall WA. Functional subunit structure of
voltage-gated calcium channels. Science. 1991;253:1499-1500.
10. Klockner U, Itagaki K, Bodi I, Schwartz A. Beta-subunit expression is required for cAMP-dependent increase of cloned cardiac and vascular calcium channel currents. Pflugers Arch. 1992;420:413-415. [Medline] [Order article via Infotrieve]
11. Haase H, Karczewski P, Beckert R, Krause EG. Phosphorylation of the L-type calcium channel beta subunit is involved in beta-adrenergic signal transduction in canine myocardium. FEBS Lett. 1993;335:217-222. [Medline] [Order article via Infotrieve]
12.
Sculptoreanu A, Rotman E, Takahashi M, Scheuer T,
Catterall WA. Voltage-dependent potentiation of the activity
of cardiac L-type calcium channel alpha 1 subunits due to
phosphorylation by cAMP-dependent protein
kinase. Proc Natl Acad Sci U S A. 1993;90:10135-10139.
13.
Diebold RJ, Koch WJ, Ellinor PT, Wang JJ, Muthuchamy M,
Wieczorek DF, Schwartz A. Mutually exclusive exon splicing of
the cardiac calcium channel alpha 1 subunit gene generates
developmentally regulated isoforms in the rat heart. Proc
Natl Acad Sci U S A. 1992;89:1497-1501.
14. Sperelakis N, Haddad GE. Developmental changes in membrane electrical properties of the heart. In: Sperelakis N, ed. Physiology and Pathophysiology of the Heart. Norwell, Mass: Kluwer Academic Publishers; 1995:669-700.
15. Chen FM, Yamamura HI, Roeske WR. Ontogeny of mammalian myocardial beta-adrenergic receptors. Eur J Pharmacol. 1979;58:255-264. [Medline] [Order article via Infotrieve]
16.
Chen FC, Yamamura HI, Roeske WR.
Adenylate cyclase and beta adrenergic receptor
development in the mouse heart. J Pharmacol Exp
Ther. 1982;222:7-13.
17. Kojima M, Ishima T, Taniguchi N, Kimura K, Sada H, Sperelakis N. Developmental changes in beta-adrenoceptors, muscarinic cholinoceptors and Ca2+ channels in rat ventricular muscles. Br J Pharmacol. 1990;99:334-339. [Medline] [Order article via Infotrieve]
18. Haddox MK, Roeske WR, Russell DH. Independent expression of cardiac type I and II cyclic AMP-dependent protein kinase during murine embryogenesis and postnatal development. Biochim Biophys Acta. 1979;585:527-534. [Medline] [Order article via Infotrieve]
19. Slotkin TA, Lau C, Seidler FJ. Beta-adrenergic receptor overexpression in the fetal rat: distribution, receptor subtypes, and coupling to adenylate cyclase activity via G-proteins. Toxicol Appl Pharmacol. 1994;129:223-234. [Medline] [Order article via Infotrieve]
20.
Misra RP, Bonni A, Miranti CK, Rivera VM, Sheng M,
Greenberg ME. L-type voltage-sensitive calcium channel
activation stimulates gene expression by a serum response
factor-dependent pathway. J Biol Chem. 1994;269:25483-25493.
21. Sturm K, Tam PP. Isolation and culture of whole postimplantation embryos and germ layer derivatives. Methods Enzymol. 1993;225:164-190. [Medline] [Order article via Infotrieve]
22. Kubalak S, Doevendans PA, Rockman H, Hunter JJ, Tanaka N, Ross J, Chien KR. Molecular analysis of cardiac muscle diseases via mouse genetics. Methods Mol Genet. In press.
23. Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 1981;391:85-100.[Medline] [Order article via Infotrieve]
24.
Horn R, Marty A. Muscarinic activation of ionic
currents measured by a new whole-cell recording
method. J Gen Physiol. 1988;92:145-159.
25. Bangalore R, Baindur N, Ruttledge A, Triggle DJ, Kass RS. L-type calcium channels: asymmetrical intramembrane binding domain revealed by variable length, permanently charged 1,4-dihydropyridines. Mol Pharmacol. 1994;46:660-666. [Abstract]
26. Pusch M, Neher E. Rates of diffusional exchange between small cells and a measuring patch pipette. Pflugers Arch. 1988;411:204-211. [Medline] [Order article via Infotrieve]
27. Doevendans PA, Davies M, An RH, Bangalore R, Rubin L, Kass RS, Chien KR. Cellular electrophysiology and immunocytochemistry of single isolated murine embryonic cardiac myocytes: a new method to study cardiogenesis in vivo and in vitro. Biophys J. 1995;68:A251. Abstract.
28.
Sanguinetti MC, Krafte DS, Kass RS. Bay K8644:
voltage-dependent modulation of Ca channel current in heart
cells. J Gen Physiol. 1986;88:369-392.
29.
Walsh KB, Begenisich TB, Kass RS. B-adrenergic
modulation of cardiac ion channels: differential
temperature-sensitivity of potassium and calcium currents.
J Gen Physiol. 1989;93:841-854.
30.
Nuss HB, Marban E.
Electrophysiological properties of
neonatal mouse cardiac myocytes in primary culture.
J Physiol (Lond). 1994;479:265-279.
31. Rohwedel J, Maltsev V, Bober E, Arnold HH, Hescheler J, Wobus AM. Muscle cell differentiation of embryonic stem cells reflects myogenesis in vivo: developmentally regulated expression of myogenic determination genes and functional expression of ionic currents. Dev Biol. 1994;164:87-101. [Medline] [Order article via Infotrieve]
32. Wobus AM, Kleppisch T, Maltsev V, Hescheler J. Cardiomyocyte-like cells differentiated in vitro from embryonic carcinoma cells P19 are characterized by functional expression of adrenoceptors and Ca2+ channels. In Vitro Cell Dev Biol. 1994;30A:425-434.
33.
Yu SS, Lefkowitz RJ, Hausdorff WP.
Beta-adrenergic receptor sequestration: a potential
mechanism of receptor resensitization. J Biol
Chem. 1993;268:337-341.
34.
Ungerer M, Parruti G, Böhm M, Puzicha M, DeBlasi
A, Erdmann E, Lohse MJ. Expression of ß-arrestins and
ß-adrenergic receptor kinases in the failing human heart.
Circ Res. 1994;74:206-213.
35. Gilbert EM, Olsen SL, Renlund DG, Bristow MR. Beta-adrenergic receptor regulation and left ventricular function in idiopathic dilated cardiomyopathy. Am J Cardiol. 1993;71:23C-29C. Review. [Medline] [Order article via Infotrieve]
36.
Ungerer M, Bohm M, Elce JS, Erdmann E, Lohse MJ.
Altered expression of ß-adrenergic receptor kinase and
ß1-adrenergic receptors in the failing human
heart. Circulation. 1993;87:454-463. Comments.
37. Bristow MR, Feldman AM. Changes in the receptor-G protein-adenylyl cyclase system in heart failure from various types of heart muscle disease. Basic Res Cardiol. 1992;87(suppl 1):15-35. Review.
38. Dhalla NS, Dixon IM, Suzuki S, Kaneko M, Kobayashi A, Beamish RE. Changes in adrenergic receptors during the development of heart failure. Mol Cell Biochem. 1992;114:91-95. Review. [Medline] [Order article via Infotrieve]
39.
Pennica D, King KL, Shaw KJ, Luis E, Rullamas J, Luoh
SM, Darbonne WC, Knutzon DS, Yen R, Chien KR, et al. Expression
cloning of cardiotrophin 1, a cytokine that induces cardiac
myocyte hypertrophy. Proc Natl Acad Sci
U S A. 1995;92:1142-1146.
40. Chien KR, Knowlton KU, Zhu H, Chien S. Regulation of cardiac gene expression during myocardial growth and hypertrophy: molecular studies of an adaptive physiologic response. FASEB J. 1991;5:3037-3046. Review. [Abstract]
41.
Koch WJ, Ellinor PT, Schwartz A. cDNA cloning of
a dihydropyridine-sensitive calcium channel
from rat aorta. J Biol Chem. 1990;265:17786-17791.
42. Gaudin C, Ishikawa Y, Wight DC, Mahdavi V, Nadal-Ginard B, Wagner TE, Vatner DE, Homcy CJ. Overexpression of Gs alpha protein in the hearts of transgenic mice. J Clin Invest. 1995;95:1676-1683.
This article has been cited by other articles:
 |
|
 |
|
  P. Weissgerber, B. Held, W. Bloch, L. Kaestner, K. R. Chien, B. K. Fleischmann, P. Lipp, V. Flockerzi, and M. Freichel Reduced Cardiac L-Type Ca2+ Current in Cav{beta}2-/- Embryos Impairs Cardiac Development and Contraction With Secondary Defects in Vascular Maturation Circ. Res., September 29, 2006; 99(7): 749 - 757. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. C. Knollmann, M. C. Casimiro, A. N. Katchman, S. G. Sirenko, T. Schober, Q. Rong, K. Pfeifer, and S. N. Ebert Isoproterenol Exacerbates a Long QT Phenotype in Kcnq1-Deficient Neonatal Mice: Possible Roles for Human-Like Kcnq1 Isoform 1 and Slow Delayed Rectifier K+ Current J. Pharmacol. Exp. Ther., July 1, 2004; 310(1): 311 - 318. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Stieber, S. Herrmann, S. Feil, J. Loster, R. Feil, M. Biel, F. Hofmann, and A. Ludwig The hyperpolarization-activated channel HCN4 is required for the generation of pacemaker action potentials in the embryonic heart PNAS, December 9, 2003; 100(25): 15235 - 15240. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Xu, A. Welling, S. Paparisto, F. Hofmann, and N. Klugbauer Enhanced Expression of L-type Cav1.3 Calcium Channels in Murine Embryonic Hearts from Cav1.2-deficient Mice J. Biol. Chem., October 17, 2003; 278(42): 40837 - 40841. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Mummery, D. Ward-van Oostwaard, P. Doevendans, R. Spijker, S. van den Brink, R. Hassink, M. van der Heyden, T. Opthof, M. Pera, A. B. de la Riviere, et al. Differentiation of Human Embryonic Stem Cells to Cardiomyocytes: Role of Coculture With Visceral Endoderm-Like Cells Circulation, June 3, 2003; 107(21): 2733 - 2740. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Banach, M. D. Halbach, P. Hu, J. Hescheler, and U. Egert Development of electrical activity in cardiac myocyte aggregates derived from mouse embryonic stem cells Am J Physiol Heart Circ Physiol, June 1, 2003; 284(6): H2114 - H2123. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Qian, M. C. Colbert, D. Witte, C.-Y. Kuan, E. Gruenstein, H. Osinska, B. Lanske, H. M. Kronenberg, and T. L. Clemens Midgestational Lethality in Mice Lacking the Parathyroid Hormone (PTH)/PTH-Related Peptide Receptor Is Associated with Abrupt Cardiomyocyte Death Endocrinology, March 1, 2003; 144(3): 1053 - 1061. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N Abi-Gerges, G J Ji, Z J Lu, R Fischmeister, J Hescheler, and B K Fleischmann Functional expression and regulation of the hyperpolarization activated non-selective cation current in embryonic stem cell-derived cardiomyocytes J. Physiol., March 1, 2000; 523(2): 377 - 389. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R-H An, B M Heath, J P Higgins, W J Koch, R J Lefkowitz, and R S Kass {beta}2-Adrenergic receptor overexpression in the developing mouse heart: evidence for targeted modulation of ion channels J. Physiol., April 1, 1999; 516(1): 19 - 30. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. A. Maltsev, G. J. Ji, A. M. Wobus, B. K. Fleischmann, and J. Hescheler Establishment of ß-Adrenergic Modulation of L-Type Ca2+ Current in the Early Stages of Cardiomyocyte Development Circ. Res., February 5, 1999; 84(2): 136 - 145. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. J. JI, B. K. FLEISCHMANN, W. BLOCH, M. FEELISCH, C. ANDRESSEN, K. ADDICKS, and J. HESCHELER Regulation of the L-type Ca2+ channel during cardiomyogenesis: switch from NO to adenylyl cyclase-mediated inhibition FASEB J, February 1, 1999; 13(2): 313 - 324. [Abstract] [Full Text]
|
|
|
|
|
|
W. Liu, K. Yasui, A. Arai, K. Kamiya, J. Cheng, I. Kodama, and J. Toyama beta -Adrenergic modulation of L-type Ca2+-channel currents in early-stage embryonic mouse heart Am J Physiol Heart Circ Physiol, February 1, 1999; 276(2): H608 - H613. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Kolossov, B.K. Fleischmann, Q. Liu, W. Bloch, S. Viatchenko-Karpinski, O. Manzke, G.J. Ji, H. Bohlen, K. Addicks, and J. Hescheler Functional Characteristics of ES Cell-derived Cardiac Precursor Cells Identified by Tissue-specific Expression of the Green Fluorescent Protein J. Cell Biol., December 28, 1998; 143(7): 2045 - 2056. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Kupershmidt, T. Yang, and D. M. Roden Modulation of Cardiac Na+ Current Phenotype by ß1-Subunit Expression Circ. Res., August 24, 1998; 83(4): 441 - 447. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J Hescheler, B.K Fleischmann, S Lentini, V.A Maltsev, J Rohwedel, A.M Wobus, and K Addicks Embryonic stem cells: a model to study structural and functional properties in cardiomyogenesis Cardiovasc Res, November 1, 1997; 36(2): 149 - 162. [Full Text] [PDF]
|
|
|
|
|
|
C. Seisenberger, V. Specht, A. Welling, J. Platzer, A. Pfeifer, S. Kuhbandner, J. Striessnig, N. Klugbauer, R. Feil, and F. Hofmann Functional Embryonic Cardiomyocytes after Disruption of the L-type alpha 1C (Cav1.2) Calcium Channel Gene in the Mouse J. Biol. Chem., December 8, 2000; 275(50): 39193 - 39199. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||