The hyperpolarization-activated, cyclic nucleotide-gated (HCN) ion channels are responsible for generating the pacemaker current (funny current, I_f) which is the inward current that contributes to the early stage diastolic depolarization in the SAN (DiFrancesco, 1993). At diastolic potentials, I_f is predominantly carried by Na⁺ (DiFrancesco, 1993). I_f is activated by hyperpolarizing voltage steps, with the threshold potential varies from -40 to -60 mV. The voltage dependence of the I_f activation is influenced by intracellular cyclic adenosine monophosphate (cAMP); a direct binding of cAMP to this channel increases the open probability via a depolarizing shift in the midpoint activation voltage (V_1/2) (DiFrancesco and Tortora, 1991).

While HCN4 is the most predominant isoform expressed in rodent SANCs (Marionneau et al., 2005), recent reports showed that all three cardiac HCN isoforms (HCN1, HCN2, and HCN4) are highly expressed in the human SAN (Li et al., 2015). Several studies demonstrated that HCN channels localize to caveolae based on the presence of HCN4 in low-density membrane fractions along with Cav-3 as well as the specific interaction of HCN4 with Cav-3 (Barbuti et al., 2007; Ye et al., 2008). Barbuti et al. (2012) reported that all HCN isoforms have a conserved caveolin-binding domain which impact both channel function and trafficking. Disruption of caveolae by reducing membrane cholesterol using methyl-β-cyclodextrin (MβCD) or by expression of dominant negative caveolin mutants alters the gating of HCN channels by shifting the voltage dependence of the activation by approximately 10 mV in the positive direction (Ye et al., 2008). In addition, β₂-adrenergic receptor (β₂AR) modulation of HCN4 channel is lost when caveolae are disrupted, which is supported by co-localization of β₂ARs and HCN4 channels in caveolae (Barbuti et al., 2007).

Recent studies indicate that caveolae-associated β₂AR-dependent stimulation of HCN4 channels relies on subcellular compartmentalization of cAMP signaling. First, it was found that HCN4 can be phosphorylated by cAMP-dependent protein kinase A (PKA) at the distal C-terminus, in addition to the well-studied cAMP binding to a conserved cyclic nucleotide binding domain in the proximal C-terminus (Liao et al., 2010). Moreover, PKA activity is necessary for cAMP-dependent signaling between β₂ARs and HCN channels in SANCs; inhibition of PKA with an inhibitory peptide, PKI, significantly reduced the shift in V_1/2 produced by β₂ARs stimulation while does not affect a direct stimulation of the channels by exogenous cAMP and Rp-cAMP (an analog than cannot activate PKA) (St Clair et al., 2013). St Clair and colleagues showed that PKA modulation of HCN4 channels depends on distinct cAMP signaling domains created by subcellular localization of cyclic nucleotide phosphodiesterases 3 and 4 (PDE3 and 4) which are responsible for cAMP degradation in the SANCs. The authors demonstrated that PDE4 inhibition in mouse SANCs produced a PKA-independent depolarizing shift in the V_1/2 of I_f at rest, likely via a direct binding of elevated cAMP to the channel, but did not remove the requirement for PKA in β₂AR-to-HCN signaling. In contrast, PDE3 inhibition produced PKA-dependent changes in I_f both at rest and in response to β₂AR stimulation (St Clair et al., 2017). Microdomain-specific localization and activity of PDEs in SANCs have been recently reviewed by Vinogradova et al. (2018) and highlight functional importance of local cAMP microdomains with high and low cAMP levels which are involved in local regulation of coupled-clock system components.

In the center of the SAN, where the AP upstroke is slow, and little or no Na⁺ current is expressed, I_Ca,L is principally responsible for the upstroke. Though Ca_v1.2 represents the major isoform of the L-type Ca²⁺ channel (LTCC) central pore subunit expressed in the heart, SANCs also express Ca_v1.3 isoform (Zhang et al., 2002). In comparison to Ca_v1.2 channels which are activated at -40 mV and mainly contribute to AP upstroke, channels formed by Ca_v1.3 are activated ∼20 mV more negatively than Ca_v1.2 and thus play an important role in the generation of diastolic depolarization. Knockout of Ca_v1.3 in mice decreased I_Ca,L density by 69% and resulted in severe bradycardia and highly erratic pacing rate in the SAN (Mangoni et al., 2003).

A number of important Ca_v1.2 subpopulations have been identified in cardiomyocytes that associate with unique macromolecular signaling complexes and scaffolding proteins (Figures 1, 2) (Best and Kamp, 2012). These include channels localized to dyadic junctions (i.e., T-tubules) as well as extradyadic channels that reside in biochemically distinct regions of surface membrane, including caveolae, lipid rafts, and plasmalemma. Most of the studies performed on isolated mouse SANCs (Le Scouarnec et al., 2008; Christel et al., 2012) demonstrated a predominant plasma membrane localization of Ca_v1.2 channels; however, a ratio between caveolar and non-caveolar LTCCs in SANCs is unknown and requires additional studies. In contrast, immunofluorescence analysis of mouse SANCs showed Ca_v1.3 channels localized both on the plasma membrane (Le Scouarnec et al., 2008) and in sarcomeric structures (presumably, within a sparse T-tubule network observed in subsidiary pacemakers (Ayettey and Navaratnam, 1978) where they co-localize with RyRs (Christel et al., 2012). STimulated Emission Depletion (STED) co-immunofluorescence labeling of mouse atrial myocytes showed that Ca_v1.3 clusters are located close to Cav-3 clusters in axial tubule junctions where highly PKA phosphorylated clusters of RyR-pS2808 are identified (Brandenburg et al., 2018). Such highly phosphorylated, both at PKA- (P-Ser²⁸⁰⁹) and Ca²⁺/calmodulin-dependent protein kinase II (CaMKII)-dependent (P-Ser²⁸¹⁵), clusters of RyRs have been found in rabbit SAN and proposed to contribute to spontaneous LCRs. Along with a heterogeneous distribution of T-tubules in the SAN (compare primary vs. subsidiary pacemakers in Figure 1), this may indicate a variable contribution of Ca_v1.3 to I_Ca,L in different SAN pacemaker clusters, their functional interaction with RyRs and thus their impact on heart rate. Indeed, Christel and colleagues demonstrated a significantly stronger inactivation of Ca_v1.2 versus Ca_v1.3 channels measured in mouse SANCs during strong (+80 mV) depolarization. The authors used a numerical model of mouse SAN automaticity and found that Ca_v1.3 voltage-dependent facilitation enhanced recovery of pacemaker activity after pauses and positively regulated pacemaking during slow heart rate (Christel et al., 2012). In a subsequent study from the same group, Torrente et al. (2016) showed that Ca_v1.3 deficiency strongly impaired [Ca²⁺]_i dynamics, reducing the frequency of local [Ca²⁺]_i release events and preventing their synchronization. These data highlight an importance of microdomain-specific localization of Ca_v1.3 channels in calcium-voltage clock coupling and in opposing abnormal slowing of heart rate.

Along with distinct functional roles that LTCCs play in cellular microdomains, emerging evidence indicate that subpopulations of LTCCs may possess different biophysical properties resulted from the difference in channel’s structure and/or subcellular microenvironment influence, and therefore experience diverse pathological remodeling. For example, extratubular LTCCs measured in rat and human atrial myocytes, which express both Ca_v1.2 and Ca_v1.3, demonstrate low single channel activity (Glukhov et al., 2015a) indicating either a different LTCC structure (both on pore-forming Ca_v1.2 and Ca_v1.3 or regulatory Ca_vβ_1-4 and α₂δ (Foell et al., 2004) subunits) or microdomain-specific regulation, including the association with various scaffolding proteins. β-subunits enhance channel trafficking to the distinct plasma membrane microdomains, increase the channel’s open probability, cause a hyperpolarization shift in the voltage-dependence of activation, and affect voltage-dependent inactivation (Foell et al., 2004). Though Ca_vβ₂ is the most abundant β subunit expressed in the heart, both Ca_vβ₁ and Ca_vβ₃ mRNA expression have been found in mouse (Marionneau et al., 2005) and rat (Yanni et al., 2011) SAN, and their mRNA levels were higher than those measured in atrial and ventricular myocardium. Similar to Ca_vβ, α₂δ subunits are also involved in channel’s trafficking via interactions with other proteins, such as extracellular matrix and other membrane proteins involved in cell adhesion (Whittaker and Hynes, 2002).

Along with spatial compartmentalization, LTCCs show a microdomain-specific cAMP- mediated and PKA-dependent regulation, both at baseline and during autonomic stimulation [reviewed in details elsewhere (Best and Kamp, 2012; Vinogradova et al., 2018) and discussed below in ‘Ca²⁺ clock’ section]. The specificity of cAMP signaling is modulated by the binding of PKA to A kinase-anchoring proteins (AKAPs) (Figure 2), which target the kinase to specific intracellular locations and LTCC pools, and provide spatiotemporal control over cAMP signaling events. Various AKAPs serve as a scaffold to assemble proteins involved in Ca²⁺ signaling regulation via microdomain-specific phosphorylation of different subpopulations of LTCCs thus adding another level of complexity to I_Ca,L and Ca²⁺ signaling regulation (see section “SND and Scaffolding Proteins”).

Hagiwara et al. (1988) reported the existence of both long-lasing (I_Ca,L) and transient (I_Ca,T) Ca²⁺ currents in isolated rabbit SANCs. The authors described their distinct biophysical properties, including voltage dependencies for activation/inactivation and pharmacology. Threshold for activation of I_Ca,T was around -60 mV, and I_Ca,T was fully activated at about -10 mV. Because activation range of I_Ca,T overlapped the pacemaker potential of SAN, the potential involvement of I_Ca,T in the generation of the diastolic depolarization in SANCs was proposed. Indeed, selective inhibition of I_Ca,T by Ni²⁺ (Hagiwara et al., 1988) or mibefradil (Protas and Robinson, 2000) resulted in significant slowing of spontaneous activity of isolated rabbit SANCs by selective suppression of the later phase of pacemaker depolarization in a concentration-dependent manner, with no effects on AP amplitude and duration.

Two out of three T-type Ca²⁺ channel α-subunits, namely Ca_v3.1 (α_1H) and Ca_v3.2 (α_1I), were detected in the heart. In contrast to Ca_v1, Ca_v3 channels produce large currents in the absence of co-expressed accessory β or α₂δ subunits, and therefore these proteins are not obligate auxiliary subunits for Ca_v3 channels. Genetic inactivation of Ca_v3.1 channels significantly slowed the intrinsic in vivo heart rate, prolonged the SAN recovery time, and slowed pacemaker activity of mouse SANCs through a reduction of the slope of the diastolic depolarization (Mangoni et al., 2006). In contrast, Ca_v3.2-deficient mice displayed normal sinus rhythm without arrhythmias (Chen et al., 2003).

To date, experimental data on T-type Ca²⁺ channels subcellular distribution, including SAN, is limited. In mice with cardiac-specific, conditional expression of the Ca_v3.1 channels, immunocytochemical labeling revealed their presence primarily on the surface sarcolemma of ventricular myocytes, with less staining within the T-tubules (Jaleel et al., 2008). Osmotic shock, which selectively eliminates T-tubules, induced a greater reduction in L- versus T-type Ca²⁺ currents. Ca²⁺ influx through T-type Ca²⁺ channels also did not induce normal SR Ca²⁺ release which suggests that T- and L-type Ca²⁺ channels are located in different portions of the sarcolemma (Jaleel et al., 2008). Similarly, in wild type mouse atrial myocytes, Curran et al. (2015) showed both Ca_v3.1 and Ca_v3.2 expressed on the plasma membrane and, surprisingly, over the atrial Z-line while their association with T-tubules is questionable. Co-immunoprecipitation analysis suggested an association of Ca_v3.1 and Ca_v3.2 channel isoforms with caveolae scaffolding protein Cav-3 in neonatal mouse ventricular myocytes (Markandeya et al., 2011). Interestingly, co-expression of Cav-3 significantly decreased the peak Ca_v3.2 current density in HEK293 cells, whereas co-expression of Cav-3 did not alter peak Ca_v3.1 current density. It is possible that Ca_v3.1 and Ca_v3.2 channels are located on the surface sarcolemma, in association with different structural microdomains (Figure 1).

NCX plays a critical role in SAN pacemaking by producing a depolarizing current and boosting depolarization rate in late diastole when local Ca²⁺ released by RyRs beneath the cell surface membrane (Figure 1, primary pacemaker) (Lakatta et al., 2010). NCX1 is the predominant isoform expressed in the heart. Atrial-specific NCX knock-out in mice resulted in severe pacemaker abnormalities manifested by “tachy-brady” arrhythmias and associated with quiescent isolated SANCs where, however, copious intracellular Ca²⁺ waves were present but failed to trigger APs (Torrente et al., 2015). These highlight a key role of NCX in synchronizing Ca²⁺ and membrane clocks.

Though data on NCX distribution in SANCs is limited, studies on atrial and ventricular myocytes indicate that NCX membrane localization might be associated with different targeting components. Immunohistological evidence from rabbit SANCs indicate significant submembrane co-localization of NCX1 and RyRs which exceeds that measured in atrial and ventricular myocytes (Lyashkov et al., 2007) where NCX1 labeling is present throughout the cell (attribute to an extensive T-tubular network) (compare ‘caveolae’ vs. ‘axial tubule junction’ compartments in Figure 1). Such localization allows crosstalk between RyR LCRs and NCX (Figure 1, ‘surface sarcolemma’ compartment). Several groups have identified NCX associated with Cav-3 by both co-immunoprecipitation and immunolabeling (Scriven et al., 2005; Camors et al., 2006) with the latter finding those complexes enriched on the cell surface and little in the T-tubules (Scriven et al., 2005). It was also reported that NCX1 has several caveolin binding motifs and that the NCX1 protein associates with Cav-3 (Bossuyt et al., 2002). Others, using the same techniques, have found little, if any, association (Cavalli et al., 2007).

Besides the ion channels discussed above, there are other ion channels that participate in the SAN pacemaking. These include (1) ion channels involved in store-operated Ca²⁺ entry (SOCE); (2) SR-Ca²⁺ activated non-selective Na⁺ transient receptor potential melastatin 4 (TRPM4) ion channel; (3) sodium channels Na_v1.5 which are likely expressed the periphery but not in the center of the SAN; and (4) chloride channels. All these channels provide an additional inward current during spontaneous diastolic depolarization and thus may contribute to SAN pacemaking. Though these channels may not play a prominent role in SAN activity, the mutations of the genes that encode them are reported to associate with SND observed clinically, suggesting their contributions to the SAN pacemaking. Importantly, most of those channels are associated with different structural proteins and have been linked to distinct microdomains, including caveolae/lipid rafts, intercalated disks, luminal SR, etc., and thus may require a precise spatial arrangement to support their functional coupling and integrity. Disruption in one of the protein localization could affect other proteins involved in a complex protein–protein interaction and thus disturb pacemaker automaticity. Finally, potassium ion channels, including a rapidly recovering transient outward current I_to (K_v4.2), a rapidly, I_Kr (ERG), and slowly activating, I_Ks (K_vLQT1), delayed rectifier currents, acetylcholine-activated K⁺ current I_K,ACh (K_ir3.1 and K_ir3.4), and ATP-sensitive K⁺ current I_K,ATP (K_ir6.2), are expressed in the SAN and also demonstrate microdomain-specific distribution and regulation, as reviewed in details elsewhere (Balycheva et al., 2015).

β₁- and β₂ARs are the primary sympathetic receptors in the heart and play different roles in the regulation of cardiac rhythm. In early 2000s, researchers already found that both ARs show a compartmentalized distribution in cardiomyocytes: β₂ARs are predominantly concentrated in T-tubules and caveolar structures, whereas β₁ARs are mainly localized in the non-caveolar membrane and non-lipid raft heavy fractions of plasma membrane (Rybin et al., 2000) (Figure 1). Distinct functional domains then were found in cardiomyocytes to conduct β₁- and β₂-adrenergic signaling (Shcherbakova et al., 2007). The cardiomyocyte membrane was reported to develop into specialized zones associated with scaffold proteins SAP97 and AKAP79/150, where β₁ARs are found enriched, whereas, β₂ARs are excluded from such microdomains (Shcherbakova et al., 2007) (Figure 2). Disrupted compartmentalized pattern of βARs was shown to be involved in ventricular myocyte remodeling associated with heart failure (Nikolaev et al., 2010).

In SAN myocytes, β₂/β₁ expression ratio is higher comparing to atrial and ventricular myocytes (Brodde et al., 1982), and a higher efficiency of β₂ARs was observed (Brodde et al., 2001). Co-immunoprecipitation and immunocytochemistry studies found in rabbit SANCs that β₂ARs co-localized with HCN4 channels in the caveolar microdomain (Figure 1), and their disruption via cholesterol depletion by MβCD completely abolished the effect of β₂-adrenergic stimulation, while the effect of β₁-adrenergic stimulation still maintained (Barbuti et al., 2007). Altogether, these suggest that the β-adrenergic stimulation in the SAN is presented in a compartmentalized pattern, in which, β₁- and β₂ARs localize and function in distinct microdomains.

M₂ muscarinic receptors are the primary type of muscarinic receptors expressed in cardiac myocytes. It was shown that M₂ receptors are located outside of caveolar fractions of plasma membrane (Feron et al., 1997). Upon parasympathetic stimulation, the translocation of M₂ receptors to caveolar fractions was detected using a PKA FRET-based biosensor (Warrier et al., 2005). It is reported that the M₂ receptor-mediated cAMP response is associated with distinct AC isoforms expressed in different membrane microdomains (Iancu et al., 2007).

Ankyrin-B (AnkB) is an important multifunctional scaffolding protein that is essential for membrane structure organization as well as trafficking and localization of various pacemaker proteins. AnkB syndrome, i.e., a type 4 long QT syndrome, is a rare cardiac arrhythmia syndrome, which is associated with a loss-of-function mutation of AnkB in the heart (Mohler et al., 2004). Dysfunctions in AnkB result in SND in human (Le Scouarnec et al., 2008) and mice (Le Scouarnec et al., 2008; Glukhov et al., 2010) and are associated with atrial fibrillation (Le Scouarnec et al., 2008; Cunha et al., 2011).

Heterozygous knocking out AnkB in mice results in significantly reduced expression of NCX, NKA and IP₃Rs in the SAN (Figure 2); furthermore, AnkB^+/- SANCs show abnormal localization of Ca_v1.3 and NCX proteins. In contrast to the homogenous membrane distribution of Ca_v1.3 channels in wild type SANCs, Ca_v1.3 expression in AnkB^+/- cells was limited to an internal perinuclear distribution (Le Scouarnec et al., 2008). This was associated with a concomitant decrease in a whole-cell I_Ca,L density. Cunha et al. (2011) demonstrated that AnkB directly associates with Ca_v1.3, and this interaction is regulated by a short, highly-conserved motif on the C4 region of the C-terminus of Ca_v1.3. Importantly, no changes in Ca_v1.2 expression and localization were found in AnkB^+/- SAN and atrial myocytes. Similar, AnkB^+/- mice showed significant reduction of I_NCX current density in SANCs (Le Scouarnec et al., 2008). Altogether, these resulted in SAN pacemaking abnormalities observed both in single SANCs (Le Scouarnec et al., 2008) and isolated SAN preparations (Glukhov et al., 2010).

Cav-3, the integral membrane protein that is essential in formation of caveolae (Figures 1, 2), is also linked with SND (Lang et al., 2016; Campostrini et al., 2017). As discussed in the previous sections, Cav-3 is involved in the regulation of multiple ion channels and transporters, including those involved in pacemaking. Cav-3 F97C and S141R mutations have been linked to the long QT type 9 inherited arrhythmia syndrome (LQT9) causing AP duration prolongation which had been attributed to an increase in late Na⁺ current (Vatta et al., 2006). In spontaneously beating neonatal cardiomyocytes, the expression of the T78M mutation in CAV3 gene significantly increased AP peak-to-peak variability without altering neither the mean rate nor the maximum diastolic potential (Campostrini et al., 2017) and was associated with a positive shift of activation of HCN4 channels, in a dominant way. The authors also identified a small cohort of patients with supraventricular arrhythmias including sinus tachycardia, bradycardia, and atrial fibrillation where the T78M Cav-3 variant is more frequent than in the general population. Recent preliminary findings from out laboratory indicate that cardiac-specific conditional knock-out of Cav-3 in mice results in significant beat-to-beat heart rate lability linked with suppressed SAN function, enhanced atrial ectopy and paroxysms of alternating periods of tachycardia-bradycardia rhythm (Lang et al., 2016). These results highlight the importance of Cav-3 in supporting functional integrity of the SAN pacemaker complex.

Another scaffolding protein involved in the SAN pacemaking and dysfunction is spinophilin. Spinophilin is ubiquitously expressed (Figure 2) and interacts with a variety of target proteins essential for Ca²⁺ homeostasis and cellular contraction in adult ventricular myocytes (Petzhold et al., 2011). Spinophilin mediates the targeting of protein phosphatase 1 to RyR (Ragusa et al., 2010). Single RyR channel’s open probability was observed to increase in spinophilin knockout mice (Chiang et al., 2014). A recent study reported a bradycardic response to α₂-adrenergic stimulation found in spinophilin null mice; however, no significant heart rate changes comparing to wild type mice were observed at baseline condition (Lu et al., 2010).

The Popeye domain containing (Popdc, POPDC1-3) gene family displays preferential expression in skeletal muscle and the heart, and encode membrane proteins harboring an evolutionarily conserved Popeye domain, which functions as a binding domain for cAMP. Popdc proteins are abundantly present in intercalated disks, lateral membranes and T-tubules (Figure 2). In the heart, atrial expression of Popdc1 is higher than in the ventricle, and the entire cardiac conduction system, including the SAN, displays the most intense expression levels (Froese et al., 2012). Null mutations of members of the Popdc gene family in mice are associated with a stress-induced sinus bradycardia and prominent sinus pauses. Moreover, the phenotype develops in an age-dependent manner, being absent in the young animal and becoming increasingly severe, as the animals grow older. In addition to cAMP binding site, Popeye domain of POPDC1 has the binding sites of KCNK2 (TREK-1, a member of two-pore K⁺ channels family) (Froese et al., 2012) and Cav-3 (Alcalay et al., 2013) proteins. It was found that TREK-1 current was increased twofold in the presence of Popdc1-3 proteins. Along with a prevalent SAN phenotype characterized by bradycardia with frequent episodes of sinus pause following stress in cardiac-specific TREK-1-deficient mice (Unudurthi et al., 2016), this may contribute to SND observed in Popdc-null mice. Finally, Popdc1-null cardiomyocytes showed a statistically significant 70% reduction in caveolae number (Alcalay et al., 2013) which can additionally contribute to SND phenotype via modulation of various pacemaker ion channels in transporters discussed in previous sections.

Another scaffolding protein family linked to SND, is AKAPs. AKAPs localize PKA to a different subcellular compartment, permitting a higher degree of selectivity and specificity of phosphorylation for different downstream PKA substrates. More than 14 different AKAPs have been shown to be expressed in both rodent and human heart tissue including: AKAP5 (AKAP150/79), AKAP7 (AKAP 15/18), gravin (AKAP12), AKAP9 (yotiao) and mAKAP. Though AKAPs’ function in the SAN is not clear, those proteins are critically important for [Ca²⁺]_i regulation and thus may contribute to Ca-clock regulation and SAN pacemaking.

Throughout all AKAPs, AKAP150/79 (AKAP5) is probably the most studied in the heart. AKAP150/79 targets PKA and phosphatases to regions near Ca_v1.2 (Figure 2) increasing the probability of long openings and coupled gating events between channels. Enzymes known to associate with AKAP150/79 include PKA, protein kinase C, CaM, AC5/6, and PP2B. Sympathetic stimulation of adult cardiomyocytes requires association of AKAP150/79 with a subpopulation of LTCCs coupled with Cav-3 (Nichols et al., 2010). Ablation of AKAP150 in mice with long QT syndrome 8 (LQT8), a disease also known as Timothy syndrome characterized by sinus bradycardia, prolonged QT interval and lethal arrhythmias, restores normal gating in Ca_v1.2-LQT8 channels and protects the heart from arrhythmias (Cheng et al., 2011). Dysfunction in AKAP10 has been associated with SAN pacemaking and SND in both mice and humans by effecting vagal regulation of the heart (Tingley et al., 2007).

A rare variant in MYH6, which encodes protein myosin heavy chain α isoform (MHC-α) is also reported associated with high risk of sick sinus syndrome (Holm et al., 2011). There are also some nuclear structural proteins that have been reported to be involved in the SND. A type II integral membrane protein that anchored to the inner nuclear membrane, emerin (Figure 2), whose mutation is known to develop Emery-Dreifuss Muscular Dystrophy (Emery and Dreifuss, 1966), is reported to associate with SND with underlying mechanism yet to illustrate (Karst et al., 2008). Mutations in the lamin A/C gene LMNA are reported to cause a variety of heart diseases including SND (Figure 2) (Hoorntje et al., 2017). Though the direct mechanisms of nucleus-associated scaffolding protein-induced modulation of SAN activity are unknown, it might be linked to pacemaker protein expression and/or trafficking.