Fast voltage-gated sodium channels of cardiac muscle

This chapter is barely relevant to Section G2(ii) of the 2023 CICM Primary Syllabus, which asks the exam candidate to "describe the normal processes of cardiac excitation and electrical activity".  Specifically, it is focused on fast voltage gated sodium channels, which are an essential element of those processes. One might think that for the specific purposes of the CICM exams it would be enough to know that these channels are ubiquitous, critically important as drug targets, and have a refractory period. But from the comments of the First Part examiners in Question 5 from the second paper of 2022, it would appear that candidates need to submerge into deep sodium channel lore.

Structure of the voltage-gated sodium channel

  • Heteromeric transmembrane protein (Nav1.5)
  • Made up of a larger (260 kDa) α subunit and two smaller (30 kDa) β-subunits
  • Functionally important elements include:
    • Extremely narrow (4.6 Å2) sodium-selective pore
    • Activating gate (gate)
    • Inactivating gate (h gate)

Three states of activity:

  • Resting state: 
    • m gate is closed and h gate is open; the channel is ready to be opened
  • Open state:
    • Opening is triggered ​​​​​​​by voltage (threshold of -60 to -70 mV)
    • Starts with the opening of the m gate ("helical screw" or "sliding helix" mechanism)
    • Permits flow of sodium ions
    • Stops abruptly within a few milliseconds because of the time-triggered closure of the h gate
  • Inactivated state:
    • Closed h gate; flow of sodium ions is impossible
    • This state ends after 100-140 milliseconds
    • A time-triggered opening of the h gate and closing of the  gate returns the channel to the resting state

Refractory periods:

  • The absolute refractory periodh gate is closed, and no depolarisation is possible
  • The relative refractory period  is the short late period of Phase 3 during which a supranormal stimulus can still produce a depolarisation of the myocyte, which will be of a sub-normal amplitude because only some of the fast sodium channels have entered a resting phase, whereas other have not.
  • The effective refractory period is the period during which the cell either is unable to depolarise, or - if it does depolarise - the amplitude of the depolarisation is so depressed that it could not possibly depolarise any surrounding cells.

From the perspective of those frantic last minutes before the exam, this chapter is probably pointless, as that time would be better spent memorising lists of pharmacokinetic information and cramming respiratory equations into their short-term memory. From that, it follows that any reader who has decided not to skip this chapter is probably not experiencing any sort of time-pressured panic, and can peruse the literature in a relaxed manner. For these lucky people, one can recommend Yu & Catterall (2003) or Andavan & Lemmens-Gruber (2011), which were the most helpful in creating the summary that follows.

The voltage-gated sodium channel family

Yu & Catterall (2003) describe this group of proteins to an excellent level of detail, and the reader who needs to know All The Things is referred there instead. For the CICM trainee, it will be sufficient to know that these are everywhere. In general these channels are ubiquitous in essentially everything that has any sort of nervous system or excitable tissue, starting with jellyfish, and in fact excitable tissues are mainly excitable because they are able to express these proteins. They are present on the surface of myocytes in truly vast quantities (Makielski et al counted around 100,000 per cell) and are therefore able to mediate a massive rapid flux of ions. The specific variety seen in the mammalian heart is the Nav1.5 isoform (Zimmer et al, 2014), and there are currently nine individual isoforms identified in humans, named  named Nav1.1 through Nav1.9. 

Structure of the mammalian voltage gated sodium channel

Again, it would be fairly pointless to reproduce diagrams of this thing here, as CICM exam candidates would never be expected to reproduce them in the written paper, and the interested reader is referred to professional resources such as Jiang et al (2020), who do a much better job of this. If the candidate had to produce a brief written description, it would probably contain the following essential elements:

  • All voltage gated sodium channel isoforms share a very similar structure, and subtle variation in his structure are what determines the slight differences in function that determine the activity of the tissue that expresses the channel.
  • In mammals, the channel is mostly made up of a huge "pseudotetrameric" protein, which is the α subunit. This thing is massive (260 kDa) and each isoform of the channel has a slightly different and unique α subunit variant.
  • The α subunit is the thing that contains the pore and the voltage-sensing apparatus
  • The α subunit is also usually bound to one or two smaller (30 kDa) β-subunits which regulate the voltage dependence, manage the gating kinetics, and coordinate the localisation of the channel on the cell surface. The cardiac voltage-gated sodium channel contains two such β-subunits.

To answer the very valid question "what could be the relevance of all this to the intensivist", the reader is reminded that mutations of these channels often cause the kind of disease that lands you in the ICU, and the critical care practitioner must be at least dimly aware of the possibility that these are heritable, which bring about the need to - for example - test the entire family of a patient who, at a young age, had an unexplained cardiac arrest. 

Cardiac voltage gated sodium channels, vs. the others

What's so special about this Nav1.5 variant, that is so structurally or functionally significant? Well. The short answer is, not much. It is not very different from the Nav1.4 isoform that appears in skeletal muscle, and in fact immature skeletal myocytes seem to express  Nav1.5 until it gets replaced by 1.4 in the adult. Moreover Nav1.5 is not a "special" cardiac channel, and all kinds of sub-variations on the theme of Nav1.5 exist in the brain, for example. The most important aspect to explore is the different affinity these channel isoforms have for different drugs which we use to block them. For example, one could point out that variants 1.1, 1.2, 1.3 and 1.7 are all highly sensitive to tetrodotoxin, whereas 1.5, 1.6, 1.8 and 1.9 all have varying degrees of tetrodotoxin resistance.  Sure, we don't tend to use much tetrodotoxin in our clinical practice, but there are other drug selectivity matters to mention:

  •  Local anesthetics, antiarrhythmics and antiepileptic all seem to bind to the same common site inside the pore, and thereby prevent the ion traffic
  • Access to this site for large hydrophilic  molecules is impossible unless the pore is open, which gives rise to the use dependence phenomenon of these drugs (we are mainly talking about Class I antiarrhythmic agents)
  • Access to the pore via fenestrations in the sides of the channel protein is available, but are accessible only from within the lipid bilayer membrane, which means only small lipophilic dryug molecules can affect the receptor in its resting state (we are talking mainly about phenytoin and benzocaine, but also probably other antiepileptics)
  • Subtle molecular variations in the pore binding site confer drug selectivity, and different agents can be highly selective for only one subvariant of the sodium channel (but none of these are currently available for human use). 

Catterall & Swanson (2015) is where most of this information has come from, and the interested reader is redirected there. 

The basis of sodium selectivity

The voltage gated sodium channel is so called because it conducts mainly the traffic of sodium. That seems like an extremely stupid statement to make, but it is important to recognise that other supposedly ion-selective channels (such as, for example, the nicotinic ligand-gated sodium channel at the neuromuscular junction) are barely selective for anything, and allow the passage of whatever cations happen to be around. In contrast to these, sodium is the only thing that passes through Nav1.5. This selectivity is conferred by an extremely narrow (4.6 Å2) pore which is formed by the side chains of four glutamate residues. This molecular site structure is extremely highly conserved and appears to be essential to the functioning of the whole protein, such that minor molecular modifications can completely undermine this selectivity, or control it in a way that permits the conductance of some completely different ions. For example, Heinemann et al (1992) took a rat sodium channel, substituted an alanine for a lysine, and turned it into a calcium channel instead. 

The mechanisms of voltage-gating

Voltage gating occurs by the activity of two gating mechanisms: a voltage-dependent activating gate, conventionally referred to as the "m" gate, and and a time-dependent inactivating gate, usually referred to as the "h" gate.  These names come from the original paper by Hodgkin & Huxley (1952), who were trying to express the sodium currents in the giant squid axon mathematically. In the  equation they devised, describing sodium conductance, represented the concentration of hypothetical "activating" molecules that affect the transfer rate for sodium, and h represented "inactivating" molecules. In brief:

  • The gate opens in response to a depolarisation of the myocyte membrane. The change in the potential difference is sensed by a protein segment made up of some positively charged amino acids which are arranged in a helical pattern and which act as a cuff or throttle of sorts, constricting or releasing the pore depending on the potential difference. The CICM trainee probably does not need to go into too much detail here, and would at most be expected to regurgitate the phrase "helical screw" or "sliding helix".
  • The  gate closes to terminate the flow of sodium, mere milliseconds after the pore has opened. The "h" gate (a loop-shaped intracellular protein segment) closes over the pore like a hinged lid, again by a mechanism that is not completely understood and that has been abundantly reviewed in excellent peer-reviewed works, which spares this author the need to embarrass himself by demonstrating his incomprehension of this subject matter. From an exam perspective, it is only essential to state that the time-dependent inactivation of this gate is responsible for the fast offset of the channels, and contributes to the refractory period, which is discussed below.
  • The h gate remains closed until the cell has partially repolarised, which prevents further depolarisation and therefore protects the myocardium from tetany.
  • The gate closes when the cell has partially repolarised, which means the m and h gates have now returned to their resting state. At this stage, the cell can be depolarised again. There seems to be some heterogeneity as to when this stage is achieved by different voltage-gated sodium channels, which means that, though theoretically you could trigger a depolarisation of a cardiac myocyte late in Phase 3 of the cardiac action potential, the resulting depolarisation would generally be somewhat anaemic, as some of the sodium channels have not yet opened their h gates.  This is best expressed by this ancient diagram from Rosen et al (1974):
    Action potentials are lower in early afterdepolarisation - from Rozen et al, 1974

In summary, these molecular gate acrobatics explain the different states which the voltage-gated sodium channels can occupy:

The three states: resting, open, and inactive

The different configurations of the m and h gates produce three distinct states of being:

  • The resting state, where the  m  gate is closed and the  gate is open, waiting quietly for an action potential to arrive. In this state, the pore is tightly closed, and drugs cannot enter it.
  • The open state,  when both the m gate and the gate are open, and the pore is free to conduct a sodium current. This state is a blink - it lasts only for 1-2 msec, and at the end of this period the sodium current drops to 0.5% of its peak, which makes this a short sharp spike. Over this timeframe, enough sodium enters the cell to depolarise it from -60 mV to something like +40 mV. The abrupt end to the flow of ions is usually referred to as "fast deactivation", and is mediated by the closure of the h gate.
  • The inactivated state, where the channel is not conducting a sodium current, but has not yet returned to its resting state because the h gate is closed.  During this absolute refractory period, the sodium channel cannot be activated again. This is probably the functional characteristic that differs most between different isoforms of the channel; for example cardiac Nav1.5 channels have a relatively short refractory period, lasting perhaps 100-140 msec.

Why do we care? Well: sodium channel blockers used as Class I antiarrhythmic bind only to the channel in the open or inactivated state, and dissociate from the binding site when the channel is resting during diastole. The kinetics of this relationship are the thing that determines which class of agent the drug falls into:

  • Class Ia: intermediate dissociation kinetics,
  • Class Ib: fast dissociation kinetics, and
  • Class Ic: slow dissociation kinetics

Absolute, effective and relative refractory periods

These are explained to a much better level of detail in the chapter on the ionic basis of spontaneous electrical activity in cardiac muscle. It is relevant to fast voltage gated sodium channels because they are mostly responsible for the "absoluteness" of the absolute period: while the  h gate is closed, there is no movement of sodium ions, and no additional stimulus to the cardiac myocyte will promote a depolarisation. In short:

  • The absolute refractory period lasts through Phase 0, 1, 2 and well into Phase 3; this is the period during which sodium channels cannot be activated again.
  • The relative refractory period  is the short late period of Phase 3 during which a supranormal stimulus can still produce a depolarisation of the myocyte, which will be of a sub-normal amplitude because only some of the fast sodium channels have entered a resting phase. whereas other have not.
  • The effective refractory period is the period during which the cell either is unable to depolarise, or - if it does depolarise - the amplitude of the depolarisation is so depressed that it could not possibly depolarise any surrounding cells.

refractory periods of the Purkinje cells from Weidmann (1955)

References

Yu, Frank H., and William A. Catterall. "Overview of the voltage-gated sodium channel family." Genome biology 4.3 (2003): 1-7.

SB Andavan, G., and R. Lemmens-Gruber. "Voltage-gated sodium channels: mutations, channelopathies and targets." Current medicinal chemistry 18.3 (2011): 377-397.

Jiang, Daohua, et al. "Structure of the cardiac sodium channel." Cell 180.1 (2020): 122-134.

Goldin, Alan L. "Resurgence of sodium channel research." Annual review of physiology 63.1 (2001): 871-894.

Wang, Jun, et al. "Multiple Nav1. 5 isoforms are functionally expressed in the brain and present distinct expression patterns compared with cardiac Nav1. 5." Molecular medicine reports 16.1 (2017): 719-729.

Catterall, William A., and Teresa M. Swanson. "Structural basis for pharmacology of voltage-gated sodium and calcium channels." Molecular pharmacology 88.1 (2015): 141-150.

Payandeh, Jian, et al. "The crystal structure of a voltage-gated sodium channel." Nature 475.7356 (2011): 353-358.

Priest, Birgit T., and Gregory J. Kaczorowski. "Subtype-selective sodium channel blockers promise a new era of pain research." Proceedings of the National Academy of Sciences 104.20 (2007): 8205-8206.

Heinemann, Stefan H., et al. "Calcium channel characteristics conferred on the sodium channel by single mutations." Nature 356.6368 (1992): 441-443.

Ulbricht, Werner. "Sodium channel inactivation: molecular determinants and modulation." Physiological reviews 85.4 (2005): 1271-1301.

Makielski, Jonathan C., and Amanda L. Farley. "Na+ current in human ventricle: implications for sodium loading and homeostasis." Journal of cardiovascular electrophysiology 17 (2006): S15-S20.

Richmond, J. E., et al. "Slow inactivation in human cardiac sodium channels." Biophysical journal 74.6 (1998): 2945-2952.

Shander, Gregg S., Zheng Fan, and Jonathan C. Makielski. "Slowly recovering cardiac sodium current in rat ventricular myocytes: effects of conditioning duration and recovery potential." Journal of cardiovascular electrophysiology 6.10 (1995): 786-795.

Hodgkin, Alan L., Andrew F. Huxley, and Bernard Katz. "Measurement of current-voltage relations in the membrane of the giant axon of Loligo.The Journal of physiology 116.4 (1952): 424.

Hodgkin, Alan L., and Andrew F. Huxley. "A quantitative description of membrane current and its application to conduction and excitation in nerve." The Journal of physiology 117.4 (1952): 500.

Rosen, Michael R., Andrew L. Wit, and Brian F. Hoffman. "Electrophysiology and pharmacology of cardiac arrhythmias. I. Cellular electrophysiology of the mammalian heart." American Heart Journal 88.3 (1974): 380-385.