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Annu. Rev. Physiol. 55:455-72 Quick links to online content
Copyright © 1993 by Annual Reviews Inc. All rights reserved

PACEMAKER MECHANISMS
IN CARDIAC TISSUE

Dario DiFrancesco
Dipartimento di Fisiologia e Biochimica Generali, Elettrofisiologia, Universita di
Italy
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Milano, via Celoria 26,20133 Milano,

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KEY WORDS: cardiac pacemaker, sinoatrial node, pacemaker current, if current, modulation

INTRODUCTION

The heartbeat is a sign of life, and not surprisingly it has attracted much
interest and curiosity since the early stages of scientific investigation. Even
Leonardo da Vinci, in his anatomical studies, realized that rhythmic, restless
activity was an intrinsic property of cardiac muscle (92), "As to the heart: it
moves itself, and doth never stop, except it be for eternity." In fact, a search
for the basis of spontaneous cardiac activity could only be undertaken several
centuries after these primitive observations with the development of techniques
that allowed the study of the electrical properties of excitable tissues and
particularly of cardiac muscle (18, 71,77,23).
Cardiac pacemaker activity originates in specialized myocytes located in
restricted areas of the heart that are characterized by the ability to beat
spontaneously even when separated from the rest of the cardiac muscle (24,
106, 103, 11, 81). Voltage-clamp investigation of pacemaker tissue opened
the way to a better understanding of the ionic mechanisms promoting
rhythmicity in pacemaker tissue (64, 6). In pacemaker cells of the mammalian
sino-atrial (SA) node, spontaneous activity results from a typical phase of
their action potential, the slow diastolic depolarization. The concept that a
slow depolarization is an inherent property of spontaneously active myocar­
dium is an old one that has been actively investigated since the first recordings
of cardiac electrical activity revealed the existence of a slow depolarizing
phase preceding the action potential onset in beating tissue (for a review, see
105). During this phase, corresponding to diastole of the cardiac contraction
cycle, the membrane slowly depolarizes following termination of an action
potential, until threshold for a new action potential is reached. Thus, the
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 456 DIFRANCESCO

diastolic depolarization is responsible for initiating rhythmic behavior and

characterizes action potentials of SA node and other spontaneously active
cardiocytes.
Besides generating rhythmic activity, the diastolic (or pacemaker) depolar­
ization is involved in the control of firing frequency by the autonomic
neurotransmitters. It is known that stimulation of the sympathetic and
parasympathetic nervous systems leads to acceleration and slowing of cardiac
rate, respectively. A main physiological mechanism by which low doses of
autonomic transmitters modulate cardiac rate is the control of the steepness
of pacemaker depolarization. An example of the way this is accomplished is
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illustrated in Figure 1. Here three traces, recorded from a sino-atrial node cell
in normal Tyrode solution and in the presence of either isoprenaline 10 nM
or acetylcholine 3 nM, are superimposed. Clearly, under these conditions
neither drug significantly alters action potential shape or duration, and changes
in the spontaneous frequency are entirely due to effects on the duration of
diastolic depolarization. This means that, at least at these low neurotransmitter
concentrations, autorhythmicity is controlled by the rate of development of
the pacemaker phase. What is the ionic basis of the diastolic depolarization?
As any depolarizing process requires a net inward ionic flow, we expect to
find at least one inward current activated in the pacemaker range of voltages
that is able to drive the diastolic depolarization. In this review we consider
the current experimental evidence for the mechanisms underlying generation
of autorhythmicity in cardiac cells and its control by l3-adrenergic and
muscarinic stimulation. As is discussed below, among the various components
present in SA node cells, the hyperpolarization-activated (ir) current appears
to be most specifically designed to initiate the development of a slow
depolarizing process in the diastolic range and to control its rate of rise in
response to neurotransmitter-induced modulation.

200 ms control
I

ISO 10 nM
mV

-50

Figure 1 Acceleration and slowing of spontaneous activity induced by isoprenaline 10 nM and

acetylcholine 3 nM, respectively, in a single sino-atrial node. cell. Temperature was 35°C.
 CARDIAC PACEMAKER MECHANISMS 457

GENERATION OF PACEMAKER ACTIVITY

The Rise and Fall of the iK2 Current

First attempts at investigating the mechanisms underlying generation of the

diastolic depolarization were made in Purkinje fibers by Weidmann ( 104),
who measured a decrease in membrane conductance during diastolic depolar­
ization. The simplest interpretation of this result was that the diastolic phase
was associated with a decaying K+ current. On the basis of similar con­
ductance measurements in voltage-clamp conditions, Vassalle (103) proposed
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that the pacemaker depolarization in Purkinje fibers resulted from a decline

of K+ permeability. The view that a decay of a K + current could be responsible
for generation of pacemaker activity later received substantial support from
the work of Noble & Tsien (80) and others (87, 60, 22), who described a
pacemaker current in Purkinje fibers in terms of a pure K + current (iK2)
outward and deactivating on hyperpolarization in the diastolic voltage range.
The iK2 current was regarded for many years as perhaps the best analyzed
cardiac ionic component, especially relevant to electrical behavior of Purkinje
fibers, because it was involved not only in the generation of the pacemaker
depolarization, but also in its control by catecholamines (59, 101). The view
that pacemaker activity of cardiac Purkinje fibers was the result of iK2
deactivation held for over ten years and was, in Weidmann's words,
"universally accepted" (105). However, in the late seventies new evidence
revealed that the iK2 interpretation was deeply incorrect. Rather than an
outward current activated on depolarization, the Purkinje fibers' iK2 was in
fact shown to be an inward current, unselectively carried by K+ and Na+ and
activated on hyperpolarization in the pacemaker range of voltages (27, 28): a
falling iK2 was to be replaced by a rising it.

The iK2 Re-interpretation: Identification with the if Current

The iK2 re-interpretation was made possible by two independent findings: (a)
K+ -depletion processes could distort the time-course of currents during voltage
clamp in Purkinje fibers to the extent of inducing a fake current reversal near
the K+ equilibrium potential (40, 27); (b) a newly found pacemaker current
carried by Na+ and K+, inward and activating on hyperpolarization in the SA
node (if or ih current) (10, 107) shared several properties in common with i K2
(41). The new interpretation (for reviews see 29. 78. 30) accommodated
several results that could not be explained previously, such as the disappear­
ance of the iK2 current in Na+ -free solutions (75), hardly a property of a pure
K+ current, and the fact that the reversal potential was too negative for a K+
current (22). Interestingly, the re-interpretation was not only able to correctly
account for the diastolic depolarization of Purkinje fibers, but also to explain
 458 DIFRANCESCO

the early conductance measurements that had led to the K+ current decay
hypothesis (37).
As in Purkinje fibers, involvement of a K+ current decay mechanism in
the generation of rhythmic activity had bcen proposed in depolarized frog
atrium (7) and in the frog sinus venosus (11). In frog atrium, rhythmic
behavior was not spontaneous, but was evoked by an externally applied
depolarizing current (inward from the cell's point of view), and since no
outward component can generate a depolarization, in this experimental
arrangement, the applied current acted as the proper pacemaker current. In
the frog sinus venosus an inward current activated on hyperpolarization was
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observed, but reported to have an activation range too negative to contribute

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to the pacemaker depolarization (11). This current was later found to have
the same properties as if in the mammalian SA node, and its participation
in pacemaking is still debated (2).

Hypotheses for Pacemaker Generation

The demonstration that iK2 and if were identical and that the Purkinje fibers'
pacemaker depolarization was associated with activation of an inward
current, not with inactivation of an outward current, lent support to the
hypothesis that in the SA node (the natural pacemaker of mammalian heart)
a similar process could be responsible for initiating pacemaker activity
(if-activation hypothesis) (9, 74, 30). Arguments against this view, however,
were based on observations that the range of if activation could be too
negative relative to the range of pacemaker depolarization (l08). Other
possible mechanisms for pacing generation in SA node have been suggested
(6, 78): the activation of the Ca2+ current (l08), and the decay of the
delayed K+ current (iK), which underlies action potential repolarization in
this preparation (iK-decay hypothesis) (12).
It is important to stress that no outward current can generate a depolariza­
tion. Thus it may be misleading to interpret the iK-decay hypothesis in terms
of iK deactivation being the depolarization generating process itself, or iK
being the pacemaker current. Some degree of confusion may arise from the
intuitive but incorrect assumption that changes in voltage are directly related
to changes in current. This is not so. In fact, during activity, the relation Iionic
= - C dV/dt holds, which means that voltage changes are directly proportional
to the absolute value of net ionic current flowing through the membrane. A
pacemaker-generating process thus requires an inward current mechanism able
to supply a constant current flow during development of a depolarization of
constant slope which is what, to a first approximation, the pacemaker
depolarization is. For these reasons, the iK-decay hypothesis above included
the assumption that a background current, inward in the diastolic range, was
unmasked by decay of iK. It should also be noticed that the ir-activation
 CARDIAC PACEMAKER MECHANISMS 459

hypothesis does not exclude that iK decays during the diastolic depolarization;
it simply assumes that the diastolic depolarization is not initiated by the
unmasking of a background inward current (passive mechanism), but by
activation of it (active mechanism). In the following we outline the main
properties of the ionic components of SA node myocytes that are activated
within the range of diastolic potentials and discuss their possible role in
pacemaker depolarization.

CURRENTS INVOLVED IN PACEMAKING

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The Hyperpolarization-activated if Current

The ir current of SA node is an unspecific cation current, normally carried by

Na + and K+, inward and slowly activating upon hyperpolarization in a voltage
range comprising that of the diastolic depolarization. First studied in 1979 in
relation to its role in the catecholamine-induced frequency acceleration (10),
its properties were later described with some detail at the multicellular (9,
107, 74), single-cell (76, 36, 25, 33, 102) and single-channel level (31, 46).
The if features are well suited for generating a depolarizing process in response
to a hyperpolarization that enters the range of it activation (30). In order to
assess the degree of if involvement in the generation of the nodal diastolic
depolarization, it is essential to evaluate this range correctly. Uncertainty over
the role of if may originate from the fact that reported values of it activation
threshold show some variability. Variability of the activation range, however,
appears to be a typical feature of the if current. As discussed below, if is
directly modulated by cAMP, whose action is to shift, under the regulatory
control of neurotransmitters, the if activation curve along the voltage axis.
This affects the degree of current activation and consequently the rate of
development of diastolic depolarization. Thus even in the same type of cell,
variability of the position of the if curve may simply reflect variations in the
intracellular cAMP levels.
In multicellular nodal preparations, it was originally reported to be activated
on hyperpolarization from holding potentials of about -35 mY, well within
the diastolic range (10, 9), although other reports indicated a threshold near
-50mV, which suggests a limited contribution to diastolic depolarization
(107). The it activation range has also been found to vary in single-cell
preparations (36, 39, 25), with estimates of the if activation threshold going
from -35 mV (32) or -40 mV (102) down to -60/-65 mV mV (25). It is possible
that part of this variability is attributable to current run-down resulting from
the wash-out through the patch pipette of a diffusible substance necessary for
channel activity (36), although some variation can also originate from natural
cell heterogeneity. The presence of run-down also suggests that measurement
 460 DIFRANCESCO

of the ir threshold in single cells may yield values more negative than the real
ones. This is because run-down quickly reduces the amount of if measurable
on voltage clamp by first displacing the current activation curve to more
negative voltages, and then by decreasing its fully activated amplitude (36).
Together with run-down, other factors like incomplete series resistance
compensation (33) may contribute to the underestimation of the size of if,
Furthermore, if activation kinetics are slow at depolarized voltages (39) and
require correspondingly long steps for complete activation, or proper JJV curve
protocols. Measurement of the activation curve with steps of fixed 0.5 to 3
sec duration, for example (76, 25, 102), may lead to incomplete current
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activation at depolarized voltages and to a more negative estimate of activation

threshold.
Another consideration relevant to the problem of the if contribution to
normal pacemaking concerns the amount of net inward current required to
drive the diastolic depolarization. Since SA node cells have a mean capacity
of about 30 pF (31, 25), and the rate of diastolic depolarization is normally
around 0. 1 mY/sec, only a tiny net inward current of 3 pA is necessary to
generate the pacemaker depolarization. It is therefore important that in relation
to its contribution to pacemaker activity, if activation is studied on an
adequately expanded current scale. Recent experimental analysis and numer­
ical reconstruction have shown that the it size and kinetics are compatible
with a contribution to diastolic depolarization and that, in fact, the process
of it activation may be considered partly (102) or fully responsible for its
generation (31). Detailed voltage-clamp investigation of the diastolic range
has also shown that in pacing cells the net background (time-independent)
current can be outward down to voltages as negative as -65 m V, thus
indicating that in these cells no pacemaker depolarization would be possible
without if activation (31).
A direct evaluation of the if contribution to diastolic depolarization would
be possible if the current could be blocked by pharmacological means. Cs, a
blocker of ie. slows but does not arrest pacemaker activity in SA node cells
at a concentration of 2 mM (83,26), which may suggest that an if contribution
is present, but not essential to pacemaker generation. It is unlikely, however,
that full block of if is achieved during activity with Cs at the concentration
of 2 mM. For example, even higher concentrations (5 mM) have been shown
to leave a substantial fraction of fully activated ie, ranging from approximately
23% at -65 mV to 38% at -45 mV (data from Reference 36, Figure 10). A
more specific and efficient ie blocker would thus be desirable for a quantitative
analysis of the if contribution. On the other hand, as Cs in the range of 1-2
mM apparently does not affect other currents in SA node cells, the Cs-induced
rate reduction indicates that if normally makes an important contribution to
cardiac pacemaking (26).
 CARDIAC PACEMAKER MECHANISMS 461

If the bulk of evidence now points to if as a major component contributing

to generation of the pacemaker depolarization, its role in mediating autonomic
rate control, as discussed below, strengthens the view that if activation
represents the most relevant mechanism involved in SA node rhythm control
at low neurotransmitter concentrations.

The Slow Inward Currents and the Background Inward

Current
Early measurements in multicellular SA nodes indicated that in this prepara­
tion a Ca2+-dependent current was elicited upon depolarization beyond a
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threshold of about -50 mV (67). It was originally proposed that this current
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could participate in the development of the diastolic depolarization, and a

numerical model for SA node activity was constructed on the assumption that
the Ca2+ current was responsible for pacemaking generation ( l 08, 109).
However, this view was not supported by evidence from current-clamp
experiments, which indicate that Ca2+ entry can give a contribution to only
the last fraction of pacemaker depolarization (14). In a more recent analysis
in single nodal myocytes, two Ca2+ inward components (L-type and T-type)
have been reported (54). The largest, long lasting (L-) component (ica,L)
activates near -30 mY, while the transient (T-) component (b,T) is reported
to activate at about -50 mY. As both these Ca2+ currents are activated on
depolarization with fast activation and, for the T-component, inactivation
kinetics, their properties are appropriate for generation of the rapid action
potential depolarization and upstroke. Because of its more negative threshold,
the T-component has also been proposed to contribute to the diastolic
depolarization, on the basis of evidence that Ni, a blocker of iCa,T, slows the
diastolic depolarization rate (54). This depends on the assumption that Ni acts
specifically on iCa,T. However, Ni also has a moderate depressing action on
if. In three cells, 40 f.LM Ni reversibly decreased if recorded during 2 sec steps
at -65 mV by 18.2 ±3.3% (D. DiFrancesco, unpublished data; see also 54,
Figure 11). In view of the small values of net current flowing during the
pacemaker phase, the slowing action of Ni cannot be taken as conclusive
evidence for a contribution of ica,T to pacemaker depolarization.
Assessment of the degree of involvement of Ca2+ currents in pacemaker
generation can also be done by measuring "window" ci+ components. Since
Ca2+ currents are activated on depolarization and not on hyperpolarization,
the presence of a window is a necessary condition for a current contribution
to initiation of diastolic depolarization. While activation and inactivation
curves for the L-component overlap over a relatively wide range at depolarized
voltages, those for the T-component show either no obvious overlapping (54),
or a substantial overlapping (49). However, evidence that a Ca2+-dependent
background component, as obtained by investigating the sensitivity of the
 462 DIFRANCESCO

instantaneous current to Ca2+ -channel blockers, can only be recorded positive

to -45 mV (33) favors the view that a substantial T-type window component
is lacking in SA node cells.
An unusual, incompletely understood property of the nodal slow inward
current is its dependence upon external Na +. This was clearly observed in
multicellular SA nodes (85, 67) and, for the L-component, in single nodal
myocytes (89). Although the Ca2+ -ionic nature of nodal slow inward currents
is clearly apparent from their Ca 2+ dependence (85, 67) and from the fact
that in single cells both T- and L-type components can be recorded inNa + -free
solutions (54), the problem of the Na+ dependence of these components
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remains to be clarified.
The nature of the net background (time-independent, instantaneous) current
in SA node is still uncertain (26). In the absence of direct experimental
evidence, a background component, inward in the pacemaker range and
Na +-dependent, has often been assumed and included in the reconstruction
of nodal electrical activity, on the basis of indirect evidence extrapolated from
data in other tissues and for consistency with previous models (38, 79, 13).
As mentioned above, the presence of an inward background component is a
necessary requirement of the iK-decay hypothesis.
Recently, aNa + -dependent current has been reported in SA node myocytes
on the basis of ion-substitution experiments (66). According to these results,
theNa + -dependence of the background component would supply some 50 pA
in the diastolic range. This value far exceeds the few pA required for the net
diastolic current (see above). Furthermore, to reconcile these data with
previous evidence that no net inward current is recorded down to about -65
mV in beating cells (33), an additional, nearly equivalent background K+
conductance must also be assumed. This agrees with the fact that the same
background conductance is reported to have a K+ permeability much larger
than that to Na + (55). The balance between inward and outward background
components in the diastolic range might provide the conditions that facilitate
initiation of the diastolic phase by activation of even small amounts of a
time-dependent inward current like if.

CONTROL OF PACEMAKER ACTIVITY BY

NEUROTRANSMITTERS

The modulation of mammalian heart rate by the autonomic nervous system

originates in the SA node, the most densely innervated cardiac region:
sympathetic stimulation leads to acceleration, and parasympathetic stimulation
to slowing of the SA node cell spontaneous activity. We now discuss the
major modifications induced by neurotransmitters on some of the ionic
 CARDIAC PACEMAKER MECHANISMS 463

currents of pacemaker cells and how these modifications contribute to the

nervous control of pacemaking.

Sympathetic Modulation
l3-adrenergic stimulation has several actions on ionic currents participating in
the generation of electrical activity of SA node cells. Stimulation of the Ca2+
current has been reported in multicellular preparations (10, 82) and in single
myocytes, where only the L-component appears to be modulated (54). As in
ventricular myocytes (86, 68), activation of adenylyl-cyclase and cAMP
production are involved in this action. This is indicated, for example, by
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experiments where the L-component is increased by l3-receptor-independent

stimulation of adenylyl-cyclase by forskolin (D. DiFrancesco, unpublished
observation).
Epinephrine increases the fully-activated Ca2+ current of SA node without
apparently affecting its kinetics (82), in agreement with evidence in other
cardiac preparations (91). Also in agreement with results in different cardiac
cells (90, 20, 17), single-channel studies in nodal myocytes have shown that
epinephrine increases the probability of L-channe1 opening without modifica­
tion of its conductance (54). As the T-component is not modulated by
[3-adrenergic stimulation, and the L-component is activated at voltages more
positive than - 30 mV (54), neither current is likely to play a major role in
sympathetic rhythm modulation in pacemaker tissue. This does not exclude
the possibility that modification of the Ca2+ current will affect the spontaneous
rate, as implied by the finding that Ca2+ current activation occurs during the
last fraction of the pacemaker depolarization (13).
A catecholamine-activated, cAMP-regulated current, inward near resting
voltages, has been described in ventricular myocytes (47, 48) and attributed
to chloride ions (58, 1, 73). Although a cr background current has been
described in the SA node (94), evidence for a catecholamine-activated
component is lacking, which rules against a contribution of background
components to the chronotropic action of sympathetic stimulation in SA node
cells.
Catecholamines have also been reported to activate the iK current in
multicellular SA node preparations, although single-cell studies are still
lacking (10, 82). Dependence upon [3-adrenergic stimulation has been
characterized for delayed K+ currents in other cardiac cell types (16, 51, 113),
where it appears to involve phosphorylation by protein kinases A and C (99,
110).
The iK increase caused by l3-stimulation provides a further, stringent
element of discrimination between different hypotheses for pacing generation.
Indeed, if decay of iK and unmasking of a background inward current were
 464 DIFRANCESCO

the main mechanisms promoting pacemaker depolarization, l3-adrenergic

stimulation would be accompanied by a larger K+ conductance and would
thus give rise to slowing, rather than acceleration. This apparent paradox can
only be resolved by assuming that the inward current promoting diastolic
depolarization is itself stimulated by catecholamines (15). As discussed above,
an increase of iCa,L can only affect the last fraction of pacemaker depolariza­
tion. This consideration supports the view that the event leading to generation
of diastolic depolarization under normal conditions is activation of the if
current. By accelerating repolarization and thus shortening action potential
duration, catecholamine-induced iK increase may help keep the ratio of action
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potential duration to cycle length within an appropriate range during adren­

ergic stimulation.
Modulation of the hyperpolarization-activated current by epinephrine and
the relevance of this mechanism to the sympathetic control of rhythm have
been known since the current was first studied in the SA node (10). In the
SA node (36) and Purkinje fibers (101), if activation by l3-adrenergic
stimulation occurs via a positive shift of the voltage dependence of the current
activation curve and kinetics. This causes acceleration of spontaneous activity
by increasing the slope of diastolic depolarization, which shortens the time
required to reach the threshold for action potential generation (see Figure 1).
Although a detailed quantitative analysis of the action of catecholamines
on if in SA node myocytes is still lacking, an if increase and a simultaneous
frequency acceleration can be recorded at fairly low isoprenaline doses
(0.001-0.003 fLM) (A. Zaza et aI, unpublished data). The comparison in Figure
1 of the activities recorded in a single SA node myocyte in control conditions
and during perfusion with isoprenaline 0. 01 fLM shows that l3-stimulation
modifies the diastolic depolarization without significant change in action
potential shape and duration, which rules against large variations of either
2+ +
Ca or K currents. This evidence supports the notion that the main process
mediating chronotropic response of moderate l3-activation is if modulation.

Parasympathetic Modulation
One of the first phenomena investigated by microelectrode recording in cardiac
tissue, the action of vagal stimulation on heart rate, was shown in early studies
to be associated with membrane hyperpolarization and an increased K+
permeability (62, 19, 24, 63). A K+ current activated by ACh (iK,Ach) has
been described in both amphibian and mammalian heart, and its properties
have been characterized in detail in intact cardiac tissue (52, 98, 5 0 84),
,

single-cell (88, 4, 96), and single-channel experiments (93, 97, 70).

However, the well established notion that vagal stimulation slows heart rate
by activating a K+ current has been recently challenged by the finding that
 CARDIAC PACEMAKER MECHANISMS 465

in SA node cells, ACh has an additional, strong inhibitory action on the if

current (43). ACh inhibits if by a mechanism, opposite to that of catechola­
mines, that involves reduced cAMP production and a consequent shift of the
current activation curve to more negative voltages (44, 45). The experimental
evidence indicates that the if inhibition is the main process by which moderate
ACh concentrations control cardiac rate (35). Although both actions are
mediated by muscarinic receptors, activation of K+ channels requires 20-fold
higher doses of ACh than if inhibition. Thus, whereas half inhibition of if
occurs at 0.013 f.l-M ACh, a concentration of 0.26 fLM ACh is required to
half activate iK,Ach. Measurement of the dose-response relationships for if
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inhibition and iK,Ach activation indicates that in the concentration range up to

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0.01-0.03 fLM, ACh acts selectively on if. Since at these doses ACh is indeed
effective at slowing spontaneous rate, the ir inhibition appears to play a key
role in the vagal control of rhythm. The notion that slowing is caused by
inhibition of the inward if current, rather than activation of a K+ conductance,
agrees with the evidence shown in Figure 1 that slowing caused by low ACh
concentrations or moderate vagal stimulation occurs without membrane
hyperpolarization (61).
There is substantial evidence that the Ca2+ current is inhibited by ACh in
various cardiac preparations (57). The ci+ current of intact SA node has
been reported to be "remarkably insensitive" to ACh (84), although a
significant reduction of basal current can sometimes be observed in single
cells (45, 8). In experiments where ir and i Ca,L were recorded simultaneously,
we have repeatedly observed that 0.03 fLM ACh substantially inhibits if, but
does not modify ica.L (34). In a study done in the range 0.01 to 300 fLM, we
have observed that the threshold ACh concentration, which reduces ica,L by
a measurable amount, is 0.1 fLM in single nodal myocytes. Even at 300 fLM
ACh, iCa.L can be maximally reduced by about only 30% «D. DiFrancesco
et aI, unpublished data). These data rule against a major role for Ca2+ currents
in ACh-induced slowing, even at relatively high ACh doses.

MECHANISMS THAT REGULATE CURRENTS

INVOLVED IN PACEMAKER ACTIVITY

Significant progress in understanding the cellular processes that regulate

cardiac ion channels has come from the use of patch-clamp techniques (56)
in isolated myocytes. Whereas the bulk of information concerning Ca2+ and
K+ currents comes from experimentation in ventricular and atrial tissues (for
review, see 57, 6, 100, 65), relevant evidence on the if and iK,Ach currents
has been derived from experiments on nodal cells.
Direct G protein-mediated channel activation was first demonstrated for the
 466 DIFRANCESCO

iK,Ach current in atrial cells (88, 4, 70, 110, 72). This mechanism, in which
channel regulation occurs across the membrane without the involvement of a
diffusible intracellular second messenger, may be especially relevant to
beat-to-beat regulation of cardiac performance. There seems to be general
agreement that a subunits of the G protein controlling the ACh-activated
channel (GK) are directly involved in channel opening, and !3'Y subunits may
control channel performance via activation of phospholipase A2 (5, 69, 3).
+
The presence of a fast, direct, G protein-mediated modulation of the Ca2
current reported in other cardiac preparations (111, 95) has not been
investigated in the SA node.
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Activation by catecholamines and inhibition by ACh of the ircurrent involve

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adenylyl-cyclase activity and cAMP, which is the second messenger in if

modulation. This has been shown in Purkinje fibers (101,21) and in SA node
myocytes (36, 45). The' chain of events leading to rhythm modulation by
neurotransmitters through the if current may therefore be summarized as
follows: by activating adenylyl-cyclase, catecholamines increase cAMP levels
and induce a consequent rightward shift of the if activation curve, which leads
to cardiac acceleration; the opposite mechanism, i.e. inhibition of adenylyl­
cyclase, decrease of cAMP levels, and generation of a leftward shift of if
activation curve, characterizes the slowing action of ACh. Modifications of
it kinetics occur without effect on the fully activated IN curve (36, 44). In
agreement with these whole cell results, direct recording of single if-channels
(31) has shown that /3-stimulation increases the probability of it channel
opening on hyperpolarization, but does not affect the single-channel conduc­
tance. A recent investigation of the interaction between cAMP and irchannels
has led to the unexpected result that cAMP controls it channels directly, rather
than by phosphorylation through cAMP-dependent protein kinase, as occurs
for all other cardiac cAMP-regulated channels (42). In this respect, if channels
are similar to cyclic-nucleotide-dependent channels of sensory neurons.
In addition to cAMP, if channels have been reported to be directly controlled
by G proteins (112). These results indicate that if channels can be activated
by Gs and inhibited by Go with a mechanism that would provide fast irchannel
control during beat-to-beat modulation. Control of if by cAMP and partial
control by G protein are not mutually incompatible. However, several
observations point to cAMP-mediated control as the main mechanism for if
modulation. These include evidence of h modulation by maneuvers that alter
the intracellular cAMP level independently of the interaction with membrane
receptors (like direct stimulation of adenylyl-cyclase or inhibition of phos­
phodiesterase) (45), as well as evidence that if channels in cell-attached patches
can be modulated by externally perfused epinephrine (31). Finally, a suggested
2+
direct role for intracellular Ca ions in modulation of it' channels (53) has
not been confirmed in inside-out macro-patch experiments (114).
 CARDIAC PACEMAKER MECHANISMS 467

CONCLUSIONS

Elucidation of the mechanisms underlying cardiac pacemaking requires a

detailed knowledge of the properties of ionic currents flowing during the
diastolic depolarization. An ionic current apt to generate and control the slow
depolarization of pacemaker cells should possess a few specific properties:
(a) the current should activate on hyperpolarization at the termination of an
action potential and should be substantially activated when the membrane
potential approaches the MDP in the range -65-70 mY; (b) its kinetics should
allow the generation of a slow voltage depolarization of approximately
Annu. Rev. Physiol. 1993.55:455-472. Downloaded from www.annualreviews.org

constant slope; (c) in order to mediate rhythm modulation by neurotransmit­

by Queens University - Kingston (Canada) on 07/25/13. For personal use only.

ters, the inward current should increase during l3-adrenergic stimulation and
decrease during cholinergic stimulation.
The it current is the only current system displaying all the above features
in SA node myocytes. Indeed, its peculiar property of being an inward current
activated on hyperpolarization in the diastolic range is by itself clearly capable
of generating a depolarizing process following action potential repolarization.
Furthermore, the ability to generate a slow depolarizing process is best
achieved by a slow time-dependent activation process, which allows the
development of a constant current flow during a de olarization. Notice that
r
inward currents activated on depolarization (e.g. Ca + and Na + currents) are
apt to underlie regenerative, all-or-nothing depolarizing processes in a
positive-feedback fashion, rather than slow, controlled depolarizations. For
this reason these latter components are responsible for generation of the fast
depolarizing phase of the action potential. Finally, if is modulated in opposite
ways by l3-adrenergic and cholinergic stimuli via a cAMP-dependent,
phosphorylation-independent pathway that allows a fine rhythm control by
the autonomic nervous system. The key element in the pacemaking process
and its modulation by moderate neurotransmitter concentrations thus appears
to be the position of the if activation curve on the voltage axis: the more this
curve is shifted to the right, the more if current will be available for a faster
diastolic depolarization. Since the position of the if activation curve is
controlled by cAMP, the diastolic rate can be finely tuned by the opposite
actions of l3-adrenergic and cholinergic transmitters to achieve the desired
autorhythmic rate.

ACKNOWLEDGMENTS
This work was supported by the Consiglio Nazionale delle Ricerche
(CT90.03303) and the Ministero dell' Universita e della Ricerca Scientifica
e Tecnologica. I thank Dr. Richard B. Robinson for comments on the
manuscript and·Prof. Denis Noble for translating Leonardo into Shakespearean
English.
 468 DIFRANCESCO
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