MiniReviews |
From the Laboratory of Cardiovascular Science, Gerontology Research Center, National Institute on Aging, Baltimore, Md.
Correspondence to Rui-Ping Xiao, MD, PhD, Laboratory of Cardiovascular Science, Gerontology Research Center, National Institute on Aging, 5600 Nathan Shock Dr, Baltimore, MD 21224. E-mail XiaoR{at}grc.nia.nih.gov
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Key Words: ß-adrenergic receptor subtype G protein cAMP compartmentalization heart failure
| Overview: Myocardial ß-Adrenergic Receptors |
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(for excitatory) and ß (for
inhibitory) by Ahlquist1 in 1948 on the basis
of their functional behavior in blood vessels, ie, vasoconstriction
versus vasodilation. Ahlquists classification was expanded by Lands
et al,2 who recognized that both
- and ß-ARs could be
further categorized into 2 distinct subtypes based on their relative
potencies for the ligands available at that time. The cloning of a cDNA
for the human ß2-AR and subsequent studies
indicate that the ß-ARs are members of the G proteincoupled
receptor superfamily, which shares a common feature, the
7-transmembrane-spanning domains.3 4 A dogma of cardiac ß-AR signal transduction is that the agonist-bound ß-AR selectively interacts with the stimulatory G protein (Gs), which activates adenylyl cyclase, catalyzing cAMP formation. Subsequently, activation of cAMP-dependent protein kinase A (PKA) leads to phosphorylation of regulatory proteins involved in cardiac excitation-contraction (EC) coupling and energy metabolism, including L-type Ca2+ channels, the sarcoplasmic reticulum (SR) membrane protein phospholamban (PLB), myofilament proteins, and glycogen phosphorylase kinase. PKA also phosphorylates and thereby activates an endogenous protein phosphatase inhibitor I, which further ensures the PKA-mediated protein phosphorylation by inhibiting protein phosphatases.5 6 In addition to these acute effects, chronic ß-AR stimulation affects multiple cellular functions, including gene transcription, cell growth, and death. After its activation, ß-AR signaling is modulated at both the receptor level and downstream of the cascade by the coordinated actions of at least 3 groups of enzymes: G proteincoupled receptor kinases (GRKs), which phosphorylate and thus desensitize the receptor; cyclic nucleotide phosphodiesterases (PDEs), which hydrolyze cAMP; and phosphatases, which dephosphorylate phosphoproteins.
Although several ß-AR subtypes have been cloned and pharmacologically characterized, those expressed in cardiomyocytes were initially thought to be exclusively the ß1-AR subtype.2 However, the traditional notion that only ß1-AR modulates cardiac contractile function has been challenged by recent studies that provide compelling evidence that at least both ß1-AR and ß2-AR functionally coexist in cardiomyocytes of many mammalian species, including humans. Many investigators have struggled to understand whether the coexpression of ß-AR subtypes represents functional redundancy. An alternative, but equally appealing, concept is that these ß-AR subtypes mobilize identical signal transduction pathways but fulfill distinct physiological and pathophysiological roles. Another possibility is that ß-AR subtypes elicit distinct cardiac responses through different signaling pathways. Over the past decade, increasing evidence has demonstrated striking qualitative and quantitative differences in the functions and signaling mechanisms of the cardiac ß1-AR and ß2-AR subtypes. In the present review, we will highlight recent advances in cardiac ß-AR subtypes (particularly ß2-AR signal transduction) that reveal a new level of diversity and specificity of cardiac ß-AR subtype stimulation and provide a conceptual framework for our understanding of the physiological and pathophysiological significance of the coexistence of receptor subtypes.
| Distinct ß-AR Subtype Actions in the Heart |
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ß-AR Subtypes Differentially Regulate Ca2+ Handling
and Contractility
Cardiac EC coupling is initiated by a Ca2+
influx through voltage-dependent sarcolemmal L-type
Ca2+ channels during an action potential. This
Ca2+ influx per se is insufficient to produce a
contraction, but it triggers a large Ca2+ release
from the SR via ryanodine receptors through a
Ca2+-induced Ca2+ release
mechanism.7 The resultant intracellular
Ca2+ (Cai) transient
activates contractile proteins, producing a contraction;
Cai is subsequently removed from the cytoplasm by
the SR Ca2+-ATPase (Ca2+
pump) and the sarcolemmal
Na+-Ca2+ exchanger. ß-AR
stimulation modulates virtually all of these important components of
the cardiac EC coupling cascade and therefore plays a prominent role in
the regulation of cardiac performance.
There are several striking physiological differences between ß1-AR and ß2-AR subtypes. In rat ventricular myocytes, whereas stimulation of both ß-AR subtypes increases L-type Ca2+ currents (ICa), Cai transients, and contraction amplitude (positive inotropic effect), only ß1-adrenergic stimulation markedly accelerates the Cai transient decay and contractile relaxation (positive lusitropic or relaxant effect).8 The absence of ß2-ARmediated cardiac relaxation has been also observed in many other mammalian species (eg, cat and sheep),9 10 but human and canine hearts are exceptions.11 12 13 14 Since ß-ARinduced cardiac relaxation is mainly mediated by PKA-dependent phosphorylation of PLB, an SR membrane protein,15 16 the different relaxant effects of ß-AR subtypes may largely be attributable to their distinct effects on PLB phosphorylation (see below). Additionally, ß1-AR but not ß2-AR stimulation reduces Cai-myofilament interaction.8 The differential regulation of myofilament sensitivity to Ca2+ by ß-AR subtypes may be also related to the ß1-ARinduced phosphorylation of myofilament proteins (see below), which inhibits the myofilament response to Ca2+. Thus, both an increased SR Ca2+ uptake and a decreased myofilament Ca2+ sensitivity contribute to the relaxant effect after stimulation of ß1-AR but not ß2-AR. Concomitant with the enhanced SR Ca2+ recycling, only ß1-AR stimulation increases the resting cytosolic Ca2+ and the likelihood of spontaneous Ca2+ oscillations in several mammalian species.8 17 18 This observation suggests that ß1-ARs may be more prone than ß2-ARs to elicit Ca2+-dependent arrhythmias. These functional differences between ß1-AR and ß2-AR stimulation suggest that there may be substantial differences in their intracellular signal transduction pathways.
ß2-ARStimulated Cai and Contractile
Responses Can Be Dissociated From Global Increases in cAMP and
PKA-Mediated Phosphorylation of Cytoplasmic
Proteins
In rat ventricular myocyte preparations, the
dose-response curve of total cAMP accumulation induced by
ß1-AR stimulation with
norepinephrine (NE) overlaps that induced by
ß2-AR stimulation with zinterol.19
However, the maximal increase in the particulate cAMP induced by
zinterol is
50% of that caused by
ß1-AR,19 suggesting differential
compartmentalization of cAMP after ß-AR subtype stimulation.
Surprisingly, the ß2-ARstimulated increases
in the particulate and total cAMP levels are apparently dissociated
from the positive inotropic effect or the increase in
Cai transients in adult rat myocytes; in
contrast, ß1-ARstimulated increase in cAMP in
either fraction is closely correlated with the cardiac functional
changes.19 A more striking example of this dissociation
has been observed in canine cardiomyocytes, in which the
ß2-ARmediated augmentation of the
Cai transient or contraction occurs in the
absence of a measurable elevation of cAMP
production.13 14
To elucidate ß-AR subtype signaling downstream from cAMP, the PKA activity in both soluble and particulate fractions and the phosphorylation state of major regulatory proteins involved in cardiac EC coupling or in energy metabolism have been systematically examined in canine hearts. Stimulation of ß2-AR fails to increase PKA activity in either fraction.14 In both rat and canine ventricular myocytes, ß2-AR activation has only a negligible effect on PLB phosphorylation,13 14 19 relative to that after ß1-AR activation. These results obtained from single isolated cardiomyocytes have been recently validated in intact animals. Anesthetized dogs exhibited positive chronotropic, inotropic, and lusitropic effects during ß2-AR stimulation, and these effects occur without a detectable increase in PKA activation or PKA-dependent phosphorylation of nonsarcolemmal target proteins, such as glycogen phosphorylase kinase in the cytoplasm, troponin I and C proteins in myofilaments, and PLB in the SR membrane.14 In contrast, ß1-ARstimulated cardiac effects are well correlated with an increase in phosphorylation of these key regulatory proteins.13 14 19 20 Thus, the lack of PKA-mediated protein phosphorylation during ß2-AR stimulation in rat and dog hearts clearly indicates that, unlike ß1-AR, ß2-AR does not transmit its signal to cytoplasmic regulatory proteins, at least in these species. The reason is unclear for the PLB and troponin I phosphorylation-independent lusitropic effect of ß2-AR stimulation in canine myocytes.
Localized cAMP Signaling During Cardiac ß2-AR
Stimulation
The aforementioned observations demonstrate that
ß2-AR signaling modulates
ICa but cannot phosphorylate
regulatory proteins remote from the cell surface membrane, suggesting
that ß2-AR signaling is tightly localized near
the subsarcolemmal microdomain, in the vicinity of L-type
Ca2+ channels (Figure
).
More direct evidence supporting localized ß2-AR
signaling has emerged from on-cell patch-clamp single L-type
Ca2+ channel recordings. Although the
effect of ß1-AR stimulation is rather
diffusive, the effect of ß2-AR stimulation is
extremely localized: the L-type Ca2+ channel
responds only to local (agonist included in pipette solution) but not
remote (agonist added in bathing solution)
ß2-AR stimulation.21 These
results are in general agreement with the observation that in frog
cardiomyocytes, in which the ß2-AR
subtype predominates,22 a local ß-AR stimulation by
isoproterenol applied to one end of the cell has little stimulatory
effect on remote L-type Ca2+
channels.23
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These results, particularly those in canine hearts, initially evoked
doubts as to whether ß2-AR cardiac response is
mediated by a cAMP-dependent signaling pathway.8 13 14 19
It has been proposed that the cardiac effects of ß-AR (subtype not
specified) might be, in part, mediated by a direct interaction between
Gs
subunits and L-type
Ca2+ channels.24 25 However,
accumulating evidence indicates that the effect of ß-AR stimulation
on cardiac ICa is mediated exclusively by a
cAMP-dependent mechanism.14 26 27 28 To delineate a
role of cAMP-dependent PKA activation in ß-AR subtype signaling,
specific PKA inhibitors, including Rp-cAMP, H-89, and a
peptide PKA inhibitor (PKI), have been used. Most studies, except one
in adult rat myocytes,29 have demonstrated that PKA
inhibitors (Rp-cAMP and H-89) not only block the effect of
ß1-AR stimulation but also completely reverse
the effects of ß2-AR.14 27 28
Similarly, in human and frog cardiac myocytes, the
ß2-ARinduced augmentation of
ICa is totally prevented by
PKI.30 Taken together, several lines of evidence
strongly support the idea that cAMP-dependent PKA activation is
obligatory for ß2-ARmediated cardiac
responses, but this ß2-ARstimulated cAMP/PKA
signaling in some species is highly localized to the surface membrane
and cannot transmit to nonsarcolemmal proteins (Figure
).
Dual Coupling of ß2-AR to Gs and
Gi Proteins
In many biological systems, Gs and
Gi proteins engage in cross talk with each other.
This cross talk is usually mediated through different receptor
families. For example, activation of muscarinic or adenosine
receptors, prototypic Gi-coupled receptors,
markedly antagonizes the positive inotropic effect of ß-ARs.
Interestingly, early work in lipid vesicles had demonstrated a physical
potential for a single receptor to couple promiscuously to more than
one class of G proteins. ß-AR, for instance, couples to both
Gs and Gi
proteins.31 Using a photoaffinity labeling technique,
recent studies have demonstrated such promiscuous G protein coupling of
ß2-AR in intact cardiomyocytes, as
manifested by a ß2-ARstimulated incorporation
of a photoreactive GTP analogue,
[32P]GTP-azidoanilide, into
subunits of
Gi2 and Gi3 in addition to
Gs.28 The maximal effect of
ß2-AR stimulation on Gi
proteins is comparable to that of carbachol, a muscarinic receptor
agonist.28 Pertussis toxin (PTX) pretreatment or the
ß2-AR antagonist (ICI 118,551)
prevents the ß2-ARmediated
Gi activation. Because
ß1-AR stimulation is not able to increase
Gi activity, the coupling to
Gi is specific for ß2-AR
subtype.28 Thus, ß2-AR signaling
exemplifies a unique mode of receptorG protein interaction; ie, a
given receptor simultaneously activates more than
one class of G proteins routing to functionally opposing pathways.
Furthermore, ß1-AR and
ß2-AR exhibit subtype-selective coupling to
Gs
.32 33 A more recent in vitro study
has revealed that ß2-AR couples to different
split variants of Gs
and that the
ß2-AR coupled to the long splice variant
displays ligand-independent constitutive activity.34
Mechanisms underlying the differential coupling of ß-AR subtypes to G proteins are not well understood. At the molecular level, studies on chimeric or mutated G proteincoupled receptors (including the major subtypes of adrenergic receptors) have shown that the third cytoplasmic loop that connects transmembrane domains V and VI of these receptors is an important determinant for G protein coupling.35 36 37 For example, replacement of the cytoplasmic loop of muscarinic receptor with that of ß2-AR can induce Gs activation in response to muscarinic agonists.37 It has also been shown that a proline-rich region of the third intracellular loop determines the different Gs coupling and sequestration of ß1-AR versus ß2-AR.33 Thus, the differences between ß1-AR and ß2-AR in G protein coupling could be eventually ascribed to some critical differences in the sequences of the cytoplasmic loop of the receptors. Recent evidence also suggests a potential role of posttranslation receptor modification in the receptor/G protein coupling. In HEK293 cells, PKA-mediated phosphorylation of ß2-AR switches the receptor coupling preference from Gs to Gi.38 At the cellular level, it has been suggested that ß-AR subtypes are located in 2 distinct cellular fractions after agonist stimulation: ß1-AR in caveolae and ß2-AR in coated pits.39 40 The difference in the subcellular distribution of ß-AR subtypes may also contribute to their distinct G protein coupling and differences in signaling. In other words, ß2-ARs but not ß1-ARs might be geometrically colocalized with the Gi proteins.
Involvement of ß2-AR/Gi Coupling in Local
Control of ß2-ARStimulated cAMP Signaling
Functional localization of cAMP/PKA signaling could be ascribed to
compartmentalization of cAMP or PKA per se (because of a localized
activation of adenylyl cyclase or PDE)23 41 or to specific
anchoring proteins of PKA.42 43 A close spatial
association of L-type Ca2+ channels with adenylyl
cyclase and PKA42 44 45 would provide a structural basis
for the ß2-ARmediated cAMP-dependent local
regulation of the channel. Nevertheless, in view of the fact that both
cAMP and active PKA catalytic subunits are readily diffusible,
additional constrictive mechanisms must be involved in the local
control of the ß2-AR cAMP/PKA signaling. In
principle, this local control could also arise from countersignal
transduction pathways that locally negate the cAMP/PKA signaling.
Involvement of ß2-ARcoupled Gi signaling in the compartmentalization of the Gs-mediated cAMP/PKA signaling has been investigated by using PTX to inhibit Gi functions. In rat ventricular myocytes, PTX treatment, which has no significant effect on baseline parameters, specifically potentiates the ß2-ARmediated positive inotropic effect.46 47 More remarkably, PTX treatment permits ß2-AR stimulation to induce a robust dose-dependent increase in PLB phosphorylation,48 accompanied by a marked relaxant effect.46 48 Thus, inhibition of Gi proteins by PTX causes ß2-AR signaling to closely resemble that of ß1-AR. In contrast, the ß1-ARmediated effects on cardiac EC coupling in rat ventricular myocytes are insensitive to PTX pretreatment,46 48 consistent with the inability of ß1-AR to activate Gi proteins.28 These studies indicate that at least in rat cardiomyocytes, ß2-AR/Gi coupling underlies the functional compartmentalization of the ß2-AR/Gsdirected cAMP/PKA signaling, which may largely account for the qualitative and quantitative differences between ß1-AR and ß2-ARmediated cardiac actions.
It has been noted that in rat ventricular myocytes, PTX
treatment has no significant effect on the
ß2-ARmediated global cAMP
accumulation27 or PKA activation.48 The
simplest explanation for these observations is that the cross talk of
ß2-ARcoupled Gi and
Gs signaling occurs downstream from PKA rather
than at the G protein or adenylyl cyclase level (Figure
). In
this regard, some evidence suggests that typical
Gi-coupled receptors, such as the muscarinic
receptor M2 or adenosine receptor
A1, counteract the effect of PKA, in part, via
activation of protein phosphatases.49 50 The role of
protein phosphatases in
ß2-AR/Gi signaling has
been recently suggested.48 Alternatively, other evidence
indicates that PDE is involved in the compartmentalization of
ß-ARmediated cAMP signaling in canine41 and
frog23 cardiac myocytes. Whether
ß2-AR/Gi coupling
activates protein phosphatases, PDE, or other unknown second
messengers to functionally compartmentalize the
ß2-AR/Gsdirected
cAMP/PKA signaling awaits further study (Figure
).
Possible Role of pHi in Mediating the
ß2-AR Positive Inotropic Response
Recently, a G proteinindependent mechanism underlying a
ß2-ARmediated cellular response has been
demonstrated.51 Specifically, ß-AR agonists can induce a
direct physical association of
Na+-H+ exchange (NHE)
regulatory factor (NHERF), an inhibitor of
Na+-H+ exchanger type 3
(NHE3), to the C-terminus of ß2-AR, relieving
the inhibitory effect of NHERF on NHE3. Thus,
ß2-AR stimulation can exert opposing effects on
NHE3 activity: a stimulatory effect induced by the direct association
of ß2-AR with NHERF and an
inhibitory effect mediated by PKA-dependent
phosphorylation of NHERF, thereby activating NHERF. The
relevance of this phenomenon to ß2-AR signaling
in cardiomyocytes, however, has not yet been explored. It
has been proposed that ß2-AR activation in
adult rat ventricular myocytes increases
pHi, in a PKA-independent and
Na+-H+-independent manner,
resulting in an enhanced contractile response.29 In
contrast, most studies have found that the
ß2-ARmediated positive inotropic effect is
closely related to a proportional increase in Cai
transient or
ICa8 13 14 19 27 28 and
can be completely reversed by specific PKA
inhibitors.14 27 28 48 Quantitative studies
are required to determine the relative contributions of changes of
pHi and Cai to the
ß2-AR contractile response.
Diversity of Cardiac ß2-AR Signaling Among
Mammalian Species
In addition to the differences in the behavior of ß-AR subtypes
within species, there is also a great diversity in
ß2-ARmediated cardiac responses among
species. At one extreme, in murine and guinea pig
cardiomyocytes, ß2-AR stimulation
normally elicits no significant contractile response.28 52
In mouse cardiomyocytes, PTX treatment unmask a de novo
ß2-ARmediated contractile
response.28 At the other extreme, in the chronically
failing human atrium and ventricle, ß2-AR
stimulation induces positive inotropic and lusitropic effects and
phosphorylation of regulatory
proteins.11 12 Between these extremes, in isolated rat and
canine ventricular myocytes,
ß2-ARmediated
ICa, Cai transient,
and contractile responses are present but can be further enhanced
by PTX treatment.46 48 It is possible that this
species-dependent diversity of cardiac ß2-AR
signaling may be largely accounted for by differences in the extent of
ß2-AR/Gi coupling among
species. For example, the
ß2-AR/Gi coupling would
be expected to be extremely robust in mouse heart, but it might be less
efficient in human hearts compared with mouse and rat hearts. It should
be cautioned that most human studies have been conducted in
preparations from chronically failing hearts, and to date, no data are
available on the effect of PTX on ß2-AR
responsiveness in normal human myocardial preparations. Thus, it is not
clear whether these pathophysiological conditions,
such as chronic heart failure, influence the
ß2-AR coupling to Gs
versus to Gi proteins.
It is noteworthy that avian species appear to use markedly different signal transduction pathways. For example, in embryonic chick ventricular myocytes, ß2-AR stimulation by zinterol increases the Ca2+ transient and contractility by increasing arachidonic acid, which is negatively regulated by cAMP/PKA signaling pathway.53
Developmental Changes in Cardiac ß-AR Subtype Signaling
In contrast to adult rat myocytes, ß2-AR
stimulation by zinterol in neonatal rat ventricular
myocytes leads to increased intracellular cAMP accumulation and
enhanced phosphorylation of PLB and troponin I,
associated with an increase in the amplitude and an acceleration of the
kinetics of the contraction and Cai
transient.54 All of these responses are similar to those
elicited by ß1-AR stimulation in neonatal rat
myocytes. Interestingly, the dose-response curve of contraction in
response to ß2-AR stimulation by zinterol is
shifted
2 orders of magnitude leftward in neonatal myocytes compared
with adult myocytes. Thus, ß2-ARs may play a
more important role in mediating the response to
catecholamines in the noninnervated neonatal
(or transplanted) heart than in the innervated adult heart.
This developmental change in cardiac ß2-AR
responsiveness is not due to a higher proportion of
ß2-ARs in the neonatal heart, because the ratio
of ß2-ARs to ß1-ARs is
similar in both preparations.54 It is noteworthy that the
contractile dose-response relation to the ß2-AR
agonist zinterol in neonatal rat myocytes54 is similar to
that in PTX-treated adult rat myocytes.46 Thus, it might
be speculated that the ß2-AR coupling to
Gi proteins might be acquired or reinforced
during development, and this could explain the greater sensitivity of
neonatal ß2-AR stimulation in the absence of
PTX. In other words, in neonatal rat myocytes, the
ß2-AR/Gi coupling is
lacking or insufficient to negate the Gs-mediated
relaxant effect and phosphorylation of regulatory
proteins.
| ß2-AR Signal Transduction in Genetically Manipulated Murine Models |
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Spontaneous ß2-AR Activation
In TG4 transgenic mice, cardiac-specific overexpression of
ß2-AR by
200-fold leads to an
agonist-independent enhancement in both the baseline adenylyl cyclase
activity and myocardial contractility in
vivo55 and in isolated atria57 or single
cardiomyocytes.28 This transgenic model opens
a new avenue in the study of ligand-free ß-AR signaling. According to
the prevailing 2-state model, G proteincoupled receptors, like
ß-ARs, exist in an equilibrium between 2 functionally and
conformationally distinct states: an inactive conformation (R) and an
active conformation capable of activating G proteins
(R*).55 57 62 In the absence of a receptor ligand, the
receptor can undergo a spontaneous transition to the activated
state; the equilibrium between R and R* sets the level of basal
receptor activation. Thus, an overexpression of a given receptor would
be expected to proportionally increase the number of R* state
receptors. Receptor agonists have a higher affinity for R*, thereby
shifting the equilibrium to the active conformation R*. Neutral
antagonists bind with equal affinity to R and R*, and
therefore inhibit receptor signaling without altering the equilibrium
between R and R*. A third class of receptor ligands known as inverse
agonists preferentially binds to R, driving the equilibrium toward the
inactive conformation R. As shown in TG4 mice, the enhanced basal
cardiac adenylyl cyclase activity and contraction are reversed by a
ß2-AR inverse agonist, ICI 118,551, both in
vivo and in vitro.28 55 57 At the moment, it is unclear
whether ß1-AR and ß2-AR
have similar abilities to undergo spontaneous activation.
Spontaneous and Agonist-Induced Active ß2-ARs Exhibit
Different Signaling
Although a 2-state model for the adrenergic receptor is sufficient
to explain many aspects of ß2-AR activation,
multiple active functional states of the receptor have been
demonstrated.63 64 65 This idea is reinforced by several
important differences between spontaneously activated
ß2-ARs and agonist-stimulated
ß2-ARs in TG4 cardiomyocytes.
First, whereas spontaneously active ß2-ARs
significantly increase the baseline contractility of
the TG4 heart, ß2-AR agonists, at maximal
concentrations, are unable to further increase contraction amplitude,
even though the contractility is not yet saturated.
Second, whereas ß2-AR agonists induce a marked
increase in ICa, ligand-independent
constitutive ß2-AR activation increases cardiac
contractility but cannot modulate
ICa.66 Finally, spontaneously
activated ß2-ARs and agonist-stimulated
ß2-ARs may differentially couple to
Gi proteins, because PTX robustly enhances the
ß2-AR agonistmediated contractile response
but only slightly enhances the effect of spontaneous
ß2-AR signaling on basal
contractility in TG4 heart cells.28 These
occurrences suggest that spontaneously active
ß2-ARs and agonist-activated
ß2-ARs may represent functionally
distinct conformational states of the receptor.
Compensatory Changes in Genetically Manipulated Animal
Models
A peculiar feature of TG4 mice is that
ß2-AR overexpression causes a marked reduction
in PLB expression, which contributes to the accelerated basal cardiac
relaxation.67 Additionally, in transgenic mice
overexpressing Gs
, there is a significant
upregulation of ß-adrenergic receptor kinase (ßARK).68
In fact, in many transgenic models, a specific or even a global
remodeling process such as hypertrophy is accompanied by
the upregulation or downregulation of a gene or set of
genes69 70 (for a review, see Reference 7171 ). Although
these compensatory changes might be beneficial for the animals
survival, they introduce complications in understanding their
phenotypes. Thus, caution should be exercised when drawing a
direct mechanistic link between genetic manipulations and
phenotypes.
| Implications of ß2-AR Signaling in Heart Failure |
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Distinct Phenotypes Induced by Chronic ß1-AR
Versus ß2-AR Stimulation
The ß-AR subtypes have markedly different chronic effects on
cardiac hypertrophy, as manifested by the distinct
phenotypes of transgenic mice overexpressing cardiac
ß1-AR versus ß2-AR. For
example, vast overexpression of cardiac ß2-AR
by
200-fold does not induce detectable cellular
hypertrophy or heart failure, at least in the short
term.28 55 57 In contrast, low level (5- to 15-fold)
overexpression of ß1-AR results in cardiac
hypertrophy and heart failure.58 This
phenotype is similar to that obtained by transgenic
overexpression of cardiac Gs
in
mice76 or by chronic ß-AR stimulation by agonist
infusion.77 78 Furthermore, heart-specific overexpression
of ß2-AR at
30-fold not only rescues
ventricular function but also reverses cardiac
hypertrophy induced by transgenic overexpression of
Gq
.79 Emerging evidence also
indicates that these ß-AR subtypes even have opposing effects on
apoptosis in cultured rat cardiomyocytes:
ß1-AR stimulation induces
apoptosis,80 81 whereas
ß2-AR stimulation inhibits
apoptosis.81 82 These studies shed new light on
whether long-term ß-AR stimulation is beneficial. Answering this
question may require discriminating ß-AR subtypes, because
ß1-AR and ß2-AR have
distinct, even opposite, chronic effects.
Differential Regulation of ß1-AR Versus
ß2-AR in Failing Hearts
Except for mitral valve disorderinduced heart
failure,83 most studies involving end-stage human heart
failure73 74 75 84 and some heart failure animal
models72 85 have shown a selective decrease in
ß1-AR number with little or no loss of
ß2-AR. The selective reduction in
ß1-AR density may be attributed to elevated
plasma levels of NE, which has a higher affinity for
ß1-AR.2 In the line of the
distinct phenotypes of transgenic overexpression of
ß1-AR versus ß2-AR, the
selective downregulation of ß1-AR may reflect a
protective mechanism in the failing heart. However, the
ß2-ARmediated cardiac response, similar to
that of ß1-AR, is also significantly
diminished.73 74 75 84 85 Next, we will discuss possible
role of ß2-AR/Gi coupling
in the loss of ß2-AR contractile response in
heart failure.
Upregulation of Gi Proteins in Heart Failure
Studies in rat and guinea pig have shown that chronic infusion of
catecholamines increases the expression of
Gi.86 87 In transgenic mice, in
which the human ß2-AR is overexpressed, the
Gi protein abundance is also significantly
enhanced.28 These results suggest that chronic ß-AR
stimulation elevates Gi expression. Increasing
evidence has demonstrated that heart failure is associated with
elevated plasma catecholamine levels, which could result in
an increase in Gi protein abundance or function.
Indeed, in chronic heart failure in both humans88 89 and
animal models,72 marked increases in
Gi mRNA levels, PTX-induced ribosylation, and
Gi abundance have been reported. Because the
coupling of ß2-AR to Gi
proteins negatively regulates the Gs-mediated
contractile response in the heart of many mammalian
species,27 28 46 47 48 the enhanced Gi
signaling might serve as a mechanism underlying the diminution of the
ß2-ARstimulated positive inotropic effect. In
human chronic heart failure, some evidence suggests that
Gi proteins may be also involved in the
diminution of ß1-AR contractile
response.90
G ProteinCoupled Receptor Kinases in Failing Hearts
Agonist-dependent desensitization can be initiated by
phosphorylation of activated receptors by
members of the GRK family.91 ßARK1 is a prototypic GRK
that has been shown to phosphorylate activated
ß1-AR and ß2-AR
subtypes in vitro.92 93 Phosphorylated
receptors become binding substrates for a class of
inhibitor proteins, ß-arrestins, which inhibit further G
protein coupling.94 Recently, it has been demonstrated
that overexpression of a second myocardial GRK, GRK5, in transgenic
mice also leads to significant cardiac ß-AR
uncoupling.95 In chronic human heart failure, the levels
and enzymatic activity of ßARK1 are significantly
elevated.96 In heart failure animal models, ßARK1
activity is also correlated with ß-AR responsiveness.97
Transgenic mice with myocardial overexpression of ßARK1 (3- to
5-fold) have a blunted contractile response to ß-AR stimulation by
isoproterenol56 98 or NE.99 More strikingly,
a genetic model of murine heart failure (MLP-/-) can be largely
reversed by an overexpression of ßARK1
inhibitor.100 Taken together, chronic heart
failure is associated with a marked increase in
Gi proteins, a selective downregulation of
ß1-AR (higher
ß2/ß1), and an
increased expression and activity of GRK2 (ßARK1). The exaggerated
ß2-AR/Gi signaling and
the enhanced ßARK1 activity may contribute to the heart
failureassociated dysfunction of ß-AR stimulation and play a
critical role in the pathogenesis of heart failure.
| Summary and Perspectives |
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Many important questions remain to be answered. Further studies are required to determine the molecular, structural, and microarchitectural bases underlying the differential G protein coupling of ß2-AR versus ß1-AR. Information on downstream signaling of ß2-ARcoupled Gi proteins only begins to emerge. In addition, studies of the chronic noncontractile effects of cardiac ß-AR subtype stimulation, eg, on cardiac cell growth and death, will broaden and deepen our understanding of the differential regulation and functionality of ß-AR subtypes in health and disease. Finally, the possible role of ß2-AR/Gi coupling in the pathogenesis of chronic heart failure merits future investigation.
Received March 2, 1999; accepted September 15, 1999.
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