Developmental Electrophysiology in the Fetus and Neonate

Introduction

In this chapter, the physiology of both impulse formation and conduction in the developing heart is discussed. Significant developmental or age-related changes in the ionic currents are responsible for the generation of cardiac action potential, as well as changes in the microscopic and macroscopic anatomic and neural substrates that govern the physiology of cardiac depolarization and repolarization during health and disease. Recently, important breakthroughs in clinical fetal electrophysiology have emerged, thanks to new imaging techniques and new genetic studies of arrhythmias. These have furthered our understanding of fetal anatomic substrates, fetal demise, and the response of the fetal conduction system to disease, medication, and other stressors. These new insights emphasize the need for research into diagnostic and management strategies that promote fetal survival. Rhythm and conduction will be integrated into the overall assessment of the high-risk pregnancy in the future, using tools such as fetal magnetocardiography, fetal and neonatal electrocardiography, and fetal magnetic resonance imaging, alongside existing echocardiographic assessment of anatomy and function.

Normal Conduction

Cardiac Action Potential

The cardiac action potential is a transient reversible electromagnetochemical wavefront responsible for the generation of the cardiac impulse. It is based on a complex series of transmembrane ion fluxes that result in a net flow of electric current across the cell membrane. ^, First, there is an unequal distribution of electrically charged sodium, potassium, and calcium ions across the lipid bilayer cell membrane ( Table 46.1 ). These ion gradients are established and maintained largely as a result of energy-expending membrane ion pumps (sodium-potassium adenosine triphosphatase [ATPase]) and ion exchange complexes (the sodium-calcium exchanger). Second, under appropriate biophysical conditions, complex polypeptide pores known as ion channels, embedded within the cell membrane, are opened, thereby allowing the passage of charged ions through the cell membrane. The detailed molecular structure of many of these ion channels is understood, and defects in ion channel transcription lead to significant levels of fetal and neonatal demise ( Fig. 46.1 ).

Table 46.1

Intracellular and Extracellular Ion Distributions in the Cardiac Cell.

	Na ⁺	K ⁺	Cl ⁻	Ca ²⁺
Extracellular fluid	145 mM	4 mM	120 mM	2 mM
	Cell membrane
Intracellular fluid	15 mM	145 mM	5 mM	10 ⁻⁴ mM

Fig. 46.1, (A) Three-dimensional representation of a sodium channel from mammalian brain. The ion channel consists of the α-subunit, with two smaller protein subunits (β 1 and β 2 ) associated with the main channel. The β 1 -subunit is found only in cardiac sodium channels and is thought to stabilize the channel. The treelike (cactuslike) structures are glycosylation sites. (B) The ion channel is composed of four domains of a protein complex that consists of six separate transmembrane segments (S1–S6). The topographic arrangement of one such domain is shown on the left. The peptide loops between S5 and S6 are thought to be the central pore of the channel. Subunit S4 is a highly charged protein segment that is thought to serve as the voltage sensor of the ion channel. (C) The sodium channel, calcium channel, and potassium channel. In sodium and calcium channels, there is covalent linkage of the four domains (I–IV). (Each domain consists of the six transmembrane segments S1–S6 already described.) Potassium channels consist of single domains. There is no covalent linkage between domains. P, Phosphorylation sites; SS, disulfide bond.

At rest, the interior of the typical cardiac cell exhibits a negative electrical potential with respect to the extracellular space. For Purkinje fibers (PFs) and atrial and ventricular myocytes, this transmembrane potential is approximately −90 mV. In the sinoatrial (SA) and atrioventricular (AV) nodes, it is approximately −60 mV ( Table 46.2 ). The energy-requiring ATPase pump maintains a high intracellular concentration of potassium ions and a low intracellular sodium ion concentration relative to the extracellular space. The unequal exchange rate of positive ions (2 K ⁺ ions incorporated, 3 Na ⁺ ions extruded) results in a net negative charge of the intracellular space due to high potassium ion permeability (relative to that for sodium ions and anions).

Table 46.2

Action Potential Characteristics in Cardiac Cells.

Modified from Sperelakis N. Origin of the cardiac resting potential. In: Berne RM, Sperelakis N, eds. Handbook of Physiology, Vol 1. The Cardiovascular System . Bethesda, MD: American Physiological Society; 1979:187–267.

	SA Node	Atrium	AV Node	His-Purkinje	Ventricle
Resting Potential (mV)
Action potential	250–260	280–290	260–270	290–295	280–290
Amplitude (mV)	60–70	110–120	70–80	120	110–120
Overshoot (mV)	0–10	30	5–15	30	30
Duration (ms)	100–300	100–300	100–300	300–500	100–200
$\dot{V}$ $\dot{V}$ _max (V/s)	1–10	100–200	5–15	500–700	100–200
Conduction velocity (m/s)	<0.05	0.3–0.4	0.1	2–3	0.3–0.4

AV, Atrioventricular; SA, sinoatrial.

When the resting potential of the cardiac cell is made less negative, either as a result of the cell exhibiting spontaneous depolarization (SA or AV node automaticity) or as a result of an advancing wave of electric current (Purkinje cells or cardiac myocytes), the cell reaches a level of depolarization called the activation threshold. The ensuing rapid influx of positively charged sodium ions (phase 0 of the action potential) rapidly depolarizes the cell to potentials close to +20 to 30 mV ( Fig. 46.2 ). The maximal rate of rise of the action potential phase 0, $\dot{V}$ $\dot{V}$ _max , in ventricular and atrial muscle approaches 200 V/s; in Purkinje fibers, even higher rates of change in membrane potential can be observed ( $\dot{V}$ $\dot{V}$ _max of approximately 500 V/s) (see Table 46.2 ). The amplitude and rate of rise of phase 0 of the action potential are key determinants of conduction velocity in the myocardium and specialized conduction system, with greater conduction velocities resulting from action potentials of greater amplitude and greater $\dot{V}$ $\dot{V}$ _max . Phase 0 is triggered by the change in membrane potential that results from the sodium channel changing from a conformationally specific “rest” state to an active or “open” channel configuration ( Fig. 46.3 ).

Fig. 46.2, The atrial, ventricular, and sinoatrial node action potential. (For atrial and ventricular action potentials, the action potentials depict changes in action potential morphology that have been associated with the failing heart.) The major ion currents and ion pumps involved in the generation of the action potentials are depicted. For each of the currents, the direction of current flow is indicated (downward indicating an inward current, upward indicating an outward current), as is the approximate time course of activation and inactivation. Gene products (probable clone) that are associated with each current are shown. Relative current amplitudes are not depicted. INa represents the inward sodium current, ICa-L represents the large, long-lasting slow inward calcium current, ICa-T represents the tiny, transient calcium current, and INaCa represents the current generated by the action of the sodium-calcium exchanger. Outward potassium currents depicted include I K1 (IK1) , the inward rectifier current; I K , the delayed rectifier current; I to , the transient outward current (of which there are two components, represented by Ito1 and Ito2 ); and, limited to the atrium, I Kur (IKur) . IKACh represents the receptor-activated potassium channel (activated by acetylcholine and adenosine). I f represents the “funny” pacemaker current, a current carried by both sodium and potassium. All these currents are discussed further in the text. Not discussed but presented here for completeness are I Cl (ICl), a chloride current; I pump , an ion pump current; and I Na - B (I Na-B), an inward background current thought to be present in sinoatrial node cells.

Fig. 46.3, Three conformational states of the sodium channel. Illustration of the changes in transmembrane action potential (top row), changes in the conformational state of the sodium channel (middle row), and corresponding activation and inactivation of the sodium current (bottom row). At rest, the membrane potential is approximately −90 mV (left). At this membrane potential, the sodium channel exists primarily in the rested state. The m gate of the sodium channel is closed. ( SF denotes the selectivity filter of the ion channel.) As the cell membrane depolarizes (phase 0 of the action potential), the sodium channel changes to the open conformation, associated with opening of the m gate (middle). The sodium current rapidly activates because sodium ions can now pass through the ion channel. As the membrane potential becomes more positive, the sodium channel enters into its third conformational state, the inactivated state. This occurs as a result of closure of the inactivation gate h (right). The sodium current rapidly inactivates, or decays, back towards zero current.

After the completion of phase 0, there is often a short, rapid repolarization phase, which creates a notch in the action potential. This notch, which represents phase 1 of the action potential , and the resulting spike-and-dome appearance of the initial portion of the action potential plateau, are most notable in epicardial ventricular muscle cells and Purkinje fibers ( Fig. 46.4A ). There are two principal charge carriers responsible for this phase 1 notch. One is a transiently activated outward potassium current, referred to as I _to , and the second is an inward chloride current (I _to2 ).

Fig. 46.4, (A) Representative action potentials from ventricular endocardium and epicardium. In the epicardium, there is a prominent spike-and-dome appearance of the action potential plateau (arrow). This spike-and-dome appearance is the result of a prominent transient outward current, carried by potassium, which is present in the epicardium and separates phases 1 and 2 of the action potential. This is much less pronounced in action potentials recorded from the endocardium. (B) Demonstration of the rate-dependent behavior of the transient outward current in canine ventricular myocardium. In epicardium, decreasing basic cycle length (BCL) (i.e., increasing heart rate) from 2000 to 300 msec markedly diminishes the spike-and-dome appearance of the action potential plateau and causes marked shortening of action potential duration. This is due to rate-dependent reduction of the transient outward current at faster heart rates (due to incomplete recovery from inactivation). This is not seen in endocardium, where the transiently activated outward potassium current (I to ) is negligible, resulting in little or no change in action potential duration in response to increases in heart rate. (Note the effects of the rate-dependent decreases in I to on action potential duration are variable. In some species and preparations, action potential duration increases; in others, it decreases.) APD 50 , Action potential duration to 50% repolarization; APD 90 , action potential duration to 90% repolarization.

The cardiac action potential is unique in its long duration when compared with ion channels in skeletal muscle and neurons. The plateau of the action potential, phase 2, which accounts for this length (see Fig. 46.2 ), can persist for hundreds of milliseconds and reflects the delicate balance of both inward and outward currents on a high-resistance membrane. The most important of the inward currents during the action potential plateau is the inward calcium current. L-type ( L for long-lasting , large ) and T-type ( T for tiny, transient ) calcium channels predominate in the heart. The L-type calcium current (I _Ca-L ) is the main charge carrier responsible for maintaining the action potential plateau during phase 2. T-type calcium channels, which activate at potentials more negative than those of the L-type calcium channels, may primarily contribute to pacemaker activity in the heart. The slow inward current, I _Ca-L , is also the major current responsible for phase 0 of the action potential in the sinoatrial and AV nodes, which lack a rapid inward sodium current (see Fig. 46.2 , right side ), and is characterized by a very slow $\dot{V}$ $\dot{V}$ _max of approximately 10 V/s. The action potentials recorded from these regions are therefore referred to as slow response action potentials .

Balancing the inward depolarizing plateau currents is a family of repolarization currents, largely outward potassium currents that drive the membrane potential back towards the resting potential (see Fig. 46.2 ). I _K begins to activate relatively late during the action potential plateau, as the time-dependent slow calcium current I _Ca-L begins to inactivate. In humans, there are two principal components of the delayed rectifier potassium current, a fast (I _Kr ) and a slow (I _Ks ) component.

Phase 3 of the action potential represents the phase of rapid repolarization of the cell towards the resting membrane potential. Phase 3 occurs as the result of decay in the inward calcium current and the activation of (several) outward potassium currents. The ion current responsible for the terminal portion of this phase of the action potential, as well as the current responsible for maintaining the resting potential, is an outward potassium current activated at negative membrane potentials, referred to as the inward rectifier, or I _K1 . An additional outward potassium repolarization current called I _Kur ( ur for ultrarapid ) is identified in the human atrium. This atrial current is a rapidly activating, noninactivating potassium current.

Other outward potassium currents have been described that can become important under specific conditions. For example, the ATP-dependent potassium current, I _K(ATP) , activates strongly under conditions of depleted intracellular stores of ATP. This outward current may become important under hypoxic conditions.

Under normal conditions, spontaneous automaticity is generally confined to the pacemaker cells of the sinoatrial node, which entrain all subsidiary pacemakers with each systole. In these cells, microelectrode recordings of atrial tissue have revealed a slow spontaneous depolarization of the membrane potential during phase 4 of the action potential, from the maximal diastolic potential towards the activation, or threshold, potential ( Fig. 46.5 ). Atrial myocytes, AV junctional cells, and His-Purkinje cells can all exhibit spontaneous diastolic depolarization, at slower rates than the sinoatrial node. These subsidiary natural pacemaker cells have an important function in maintaining viability in the fetus that develops complete heart block due to AV nodal damage from certain maternal antibodies associated with systemic lupus erythematosus (SSA and SSB), or in complex congenital heart disease. The rates of these “escape” pacemakers are determined by their location, with atrium greater than ventricle. It is now believed that spontaneous automaticity may result from a combination of factors, including a decline in an outward potassium current (possibly a decline in I _K or in AV node cells, I _K1 ), increases in both L-type and T-type calcium currents, and a transient background Na ⁺ current. ^, Spontaneous automaticity contributes to “parasystolic foci,” which are premature ventricular contractions (PVCs) that can maintain a regular rhythmic pattern despite the SA node influence, and may also explain many ectopic beats. When rhythmic, abnormal spontaneous automaticity can result in tachyarrhythmia, such as sinus tachycardia and atrial ectopic tachycardia. Another inward current, carried partly by potassium and partly by sodium and termed I _f ( f for funny ), has been described and is believed to be an important pacemaker current in relatively hyperpolarized cells.

Fig. 46.5, Mechanisms of modulation of cardiac pacemaker rate. Top panel , Two sinoatrial node action potentials are illustrated. The threshold potential (TP) for both action potentials is −40 mV. The maximal diastolic potential (MDP) is approximately −60 mV. If the slope of spontaneous depolarization decreases (b), the pacemaker firing rate is delayed. Bottom panel , Three sinoatrial node action potentials are shown. A decrease in the TP (i.e., less negative, TP-1 to TP-2) results in a delay in firing rate ( c vs. b ). Pacemaker firing rate can also be delayed ( e vs. c ) by an increase in the MDP (i.e., a more negative diastolic potential) ( a vs. d ), with the slope of spontaneous depolarization and the TP remaining constant.

Developmental Changes in Action Potential Morphology and Transmembrane Ion Currents

Changes in action potential morphology and corresponding changes in ion current physiology occur throughout the course of development.

Resting Membrane Potential

Increase in sodium-potassium ATPase activity noted during development may in part result from expression of different isoforms of the sodium-potassium ATPase pump, resulting in a more negative resting membrane potential.

An increase in membrane permeability to potassium with development also contributes to the more negative resting potential observed with maturation. Specific increases in the current density of the inward rectifier current, I _K1 , the main outward potassium current responsible for maintenance of the resting membrane potential, have been reported in animal studies.

In the human fetus at mid-gestation (about 20 weeks), and in young infants undergoing heart surgery, resting membrane potentials are similar to values reported in adults. ^,

Action Potential Upstroke, Phase 0

Increases in action potential phase 0 amplitude and $\dot{V}$ $\dot{V}$ _max with maturation have been documented and are only partly the result of the increases in the resting membrane potential already described. Developmental changes in the structure or function of the ion channel or channels responsible for the upstroke of the action potential appear to contribute. ^,

Phase 1 of the Action Potential

Maturation of the spike-and-dome appearance in Phase 1 correlates with the messenger RNA expression of genes associated with the transient outward current I _to , specifically, KCNIP2 and KCND3 (which encodes Kv4.3). The maturation of the transient outward current is also associated with the development of the phenomenon of “cardiac memory,” which refers to changes in repolarization that occur with, and that persist after, a period of chronic pacing. In human atrial cells, an absence of the typical spike-and-dome shape of the action potential has been reported in infant tissue, and is believed to reflect a paucity of the transient outward current. However, I _to current was subsequently identified in some cells isolated from young infants undergoing heart surgery, ^, significant changes in current density and in recovery kinetics have been reported with maturation to adulthood. A larger current density, faster inactivation, and slower recovery from inactivation of I _to in the neonate has been described and are believed to be a function of the differences in the relative expression of KCNIP2 and KCND3 . Thus there appear to be consistent age-related increases in I _to , the transient outward current ( Fig. 46.6 ). The functional significance of the increase in the transient outward current is not fully understood, nor are the mechanisms that lead to the increased expression of the transient outward current in postnatal life, although augmentation of I _to may be linked to postnatal increases in oxygen tension.

Fig. 46.6, (A) Comparison of human atrial action potentials. Note that in the adult atrial action potential, there is a prominent spike-and-dome appearance to the plateau phase of the action potential. This is presumed to reflect a prominent transient outward current (I to ) in the adult. This spike-and-dome shape of the atrial action potential is absent in the recording from a young infant. (B) Relationship between age and I to amplitude in human atrial myocytes. Note that the I to amplitude (pA/pF) is significantly less ( asterisk, P < .05) in atrial cells isolated from infants younger than 2 years.

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