Physiological and Pharmacological Insights into the Role of Ionic Channels in Cardiac Pacemaker Activity

AbstractThe hyperpolarization-activated cation current If is a key determinant for cardiac pacemaker activity. It is conducted by subunits of the hyperpolarization-activated cyclic nucleotide–gated (HCN) channel family, of which HCN4 is predominant in mammalian heart. Both loss-of-function and gain-of-function mutations of the HCN4 gene are associated with sinus node dysfunction in humans; however, their functional impact is not fully understood yet. Here, we sought to characterize a HCN4 V759I variant detected in a patient with a family history of sick sinus syndrome. The genomic analysis yielded a mono-allelic HCN4 V759I variant in a 49-year-old woman presenting with a family history of sick sinus syndrome. This HCN4 variant was previously classified as putatively pathogenic because genetically linked to sudden infant death syndrome and malignant epilepsy. However, detailed electrophysiological and cell biological characterization of HCN4 V759I in Xenopus laevis oocytes and embryonic rat cardiomyocytes, respectively, did not reveal any obvious abnormality. Voltage dependence and kinetics of mutant channel activation, modulation of cAMP-gating by the neuronal HCN channel auxiliary subunit PEX5R, and cell surface expression were indistinguishable from wild-type HCN4. In good agreement, the clinically likewise affected mother of the patient does not exhibit the reported HCN4 variance. HCN4 V759I resembles an innocuous genetic HCN channel variant, which is not sufficient to disturb cardiac pacemaking. Once more, our work emphasizes the importance of careful functional interpretation of genetic findings not only in the context of hereditary cardiac arrhythmias.

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The effect of temperature on spontaneous action potential discharge of the isolated sinus venosus from winter and summer plaice (Pleuronectes platessa)

Journal of Experimental Biology ◽

10.1242/jeb.198.1.137 ◽

1995 ◽

Vol 198 (1) ◽

pp. 137-140 ◽

Cited By ~ 4

Author(s):

A A Harper ◽

I P Newton ◽

P W Watt

Keyword(s):

Action Potential ◽

Discharge Rate ◽

Cardiac Pacemaker ◽

Intrinsic Rate ◽

Effect Of Temperature ◽

Pacemaker Activity ◽

Duration Of Action ◽

Diastolic Depolarization ◽

Significant Difference

The spontaneous cardiac pacemaker activity and conformation were recorded in vitro, using intracellular recording methods, from heart tissue of summer- and winter-caught plaice. The effects of changing temperature on the pacemaker rate, duration of action potential and diastolic depolarization were investigated by altering the temperature of the superfusing medium. The resting intrinsic rate of discharge was significantly greater in pacemaker cells from winter plaice (P=0.05), but there was no significant difference between winter and summer fish in the apparent Arrhenius activation energies for this process. However, there was a significant difference in the estimated intercept, indicating a thermal shift in the processes underlying the spontaneous pacemaker rhythm. There was no significant difference in the diastolic depolarization duration recorded from winter and summer fish over the temperature range 422 °C. The major effect of previous environmental temperature was on the duration of the action potential (P<0.02), indicating that the observed changes in pacemaker discharge rate were not influenced by the processes that determine the duration of the pacemaker diastolic depolarisation but were modulated by the channel events that give rise to the action potential.

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Cellular and subcellular mechanisms of cardiac pacemaker oscillations

Journal of Experimental Biology ◽

10.1242/jeb.81.1.205 ◽

1979 ◽

Vol 81 (1) ◽

pp. 205-215

Author(s):

R. W. Tsien ◽

R. S. Kass ◽

R. Weingart

Keyword(s):

Membrane Potential ◽

Voltage Clamp ◽

Surface Membrane ◽

Cardiac Pacemaker ◽

Oscillatory Activity ◽

Purkinje Fibre ◽

Pacemaker Activity ◽

Heart Cells ◽

Control Loop ◽

Internal Oscillation

Rhythmic oscillations in the membrane potential of heart cells are important in normal cardiac pacemaker activity as well as cardiac arrhythmias. Two fundamentally different mechanisms of oscillatory activity can be distinguished at the cellular and subcellular level. The first mechanism, referred to as a surface membrane oscillator, can be represented by a control loop in which membrane potential changes evoke delayed conductance changes and vice versa. Since the surface membrane potential is a key variable within the control loop, the oscillation can be interrupted at any time by holding the membrane potential constant with a voltage clamp. This mode of oscillation seems to describe spontaneous pacemaker activity in the primary cardiac pacemaker (sinoatrial node) as well as other regions (Purkinje fibre, atrial or ventricular muscle). In all tissues studied so far, the pacemaker depolarization is dominated by the slow shutting-off of an outward current, largely carried by potassium ions. The second mechanism can be called an internal oscillator since it depends upon a subcellular rhythm generator which is largely independent from the surface membrane. Under voltage clamp, the existence of the internal oscillation is revealed by the presence of oscillations in membrane conductance or contractile force which occur even though the membrane potential is held fixed. The two oscillatory mechanisms are not mutually exclusive; the subcellular mechanism can be preferentially enhanced in any given cardiac cell by conditions which elevate intracellular calcium. Such conditions include digitalis intoxication, high Cao, low Nao, low or high Ko, cooling, or rapid stimulation. Several lines of evidence suggest that the subcellular mechanism involves oscillatory variations in myoplasmic calcium, probably due to cycles of Ca uptake and release by the sarcoplasmic reticulum. The detailed nature of the Cai oscillator and its interaction with the surface membrane await further investigation.

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Abstract 11423: Mitochondrial Calcium Flux Modulates Pacemaker Activity in Mouse Stem Cell Derived Cardiomyocytes

Circulation ◽

10.1161/circ.130.suppl_2.11423 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

An Xie ◽

Anyu Zhou ◽

Hong Liu ◽

Guangbin Shi ◽

Kenneth R Boheler ◽

...

Keyword(s):

Ryanodine Receptor ◽

Mrna Level ◽

Embryonic Stem ◽

Calcium Flux ◽

Pacemaker Activity ◽

Receptor Subtypes ◽

Ip3 Receptors ◽

Cell Current ◽

Beating Rate

INTRODUCTION: Ca2+ release from sarcoplasmic reticulum (SR) is known to contribute to the pacemaker activity in embryonic stem cells (ESC) derived cardiomyocytes (CMs). Mitochondria are known to participate in Ca2+ cycling. Nevertheless, the role of mitochondria in pacemaker activity is unclear. We studied the role of mitochondrial Ca2+ flux in spontaneously activity of ESC derived CMs. METHODS: CMs were derived from Wt and ryanodine receptor type 2 knockout (RYR2-/-) mouse ESC. Action potentials (APs) were recorded by perforated whole-cell current-clamp. Cytoplasmic and mitochondrial Ca2+ transients were determined by Fluo-4 and Rhod-2 respectively. Mitochondrial Ca2+ uniporter (MCU) siRNA was used. The mRNA level was evaluated by qPCR. RESULTS: As predicted, SR Ca2+ handling inhibitors, 10 μM ryanodine and 2 μM 2-APB, reduced spontaneous beating rate to 56% and 73% respectively in Wt CMs. Inhibition of mitochondrial Ca2+ flux by 10 μM Ru360 showed a similar inhibition effect on the pacemaker activity as 2 μM 2-APB in Wt CMs. To isolate the mitochondrial component, we used RYR2-/- CMs. In these cells, MCU inhibition by pharmacological or molecular biological means reduced beating rate. The MCU mRNA decreased by 96% after MCU siRNA silence 72 hrs (p<0.01). AP and mitochondrial Ca2+ transient synchronous recording revealed that the reduction of spontaneous beating rate accompanied with the depressed mitochondrial Ca2+ uptaking and releasing. In RyR2-/- CMs, 2 μM 2-APB could significantly lower the spontaneous beating rate. While 2 μM 2-APB was applied to MCU silenced RyR2-/- CMs, the beating rate couldn’t be slowed down further. This indicated IP3 receptors reduced spontaneous beating rate via MCU. Thapsigargin could substantially slow down beating rate like 2-APB. Caffeine depletion experiments showed other ryanodine receptor subtypes didn’t contribute Ca2+ release in RyR2-/- CMs. A L-type Ca2+ channel block, 10 μM nifedipine, couldn’t reduce beating frequency. This indicated spontaneous beating rate is Ca2+ influx independent in RyR2-/- CMs. CONCLUSIONS: Mitochondrial Ca2+ handling plays an important role in decreasing spontaneous beating rate. IP3R reduced spontaneous beating rate through MCU.

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