THE AUSTRALIAN NATIONAL UNIVERSITY

1. THE AUSTRALIAN NATIONAL UNIVERSITY Modulation of the Cardiac Actionpotential and ContractionChristian StrickerAssociate Professor for Systems PhysiologyANUMS/JCSMR - ANUChristian.Stricker@anu.edu.auhttp://stricker.jcsmr.anu.edu.au/Cardiac_Actionpotential.pptx

3. Aims The students should • know the elements involved in EC coupling in heart; • be able to describe Ca2+ clearance from PM; • recognise the receptors involved autonomic control of the heart, incl. which path is normally stronger; • be able to state the different forms of tropism; • understand the signalling cascades and the downstream targets after receptor activation; and • appreciate how Ca2+ channel blockers affect the heart.

4. Contents • EC coupling in the heart and Ca2+ clearance. • Tropisms in the heart. • Innervation of the heart and autonomic control. • Sympathetic effect on SAN and myocyte incl. downstream targets of signalling. • Parasympathetic effect on SAN incl. downstream targets of signalling. • AP under different conditions • Calcium channel blockers. • How hyperkalaemia and hypoxia alter AP.

5. The description of the ionic currents until here “may already have exhausted the reader, but it certainly does not exhaust the mechanisms that have been found.” Denis Noble 1936 - ; Oxford • Why so many different ionic/metabolic currents? • Complexity may result in stability / robustness (?). • Evolutionary artefact (?)

6. I. EC Coupling & Ca2+ Clearance

7. EC-Coupling and Relaxation • Obligatory Ca2+ flow via L-type channels: requires extracellular Ca2+; heart stops beating in Ca2+-free solute. • Ca2+ release from SR stores: electrochemical coupling. • Ca2+ causes shortening of contractile proteins. • Force production regulated by 2nd messenger systems. • Pumps clear sarcoplasma from Ca2+ (ER and plasma membrane).

8. Contractility and ANS • Sympathetic stimulation of β1-receptors via isoproterenol causes a much larger Ca2+ influx and bigger force with a faster rate of shortening and relaxation. • Targets of modulation by 2nd messengers: • Action potential (see next). • Muscle • L-type Ca2+ channels, • Cytosolic Ca2+ concentration, • Store refilling via SERCA/PLB, and • Contractile proteins (troponin I). • When phosphorylated via PKA, stronger inhibition of troponin C → force↑, faster cross-bridge cycling and relaxation. Berne & Levy, 2008

9. Cardiac Tropisms – Neg & Pos • ICS • Chronotropism: in regard to heart rate • Bathmotropism: in regard to AP threshold • Dromotropism: in regard to AVN conduction delay • Myocyte • Inotropism: in regard to contractile force • Lusitropism: in regard to rate of relaxation

10. II. Autonomic Innervation

11. Autonomic Innervation of Heart • VA: restricted innervation (nodes only); right: SA node; left: AV node. • SY: diffuse innervation (whole heart); right: SA node; left: ventricle. • Parasympathetic (X = vagal; VA): via ACh.Effect: inhibitory action (HR↓; SV↓) and fast. • Sympathetic(T1-T5; SY): via noradrenaline.Effect: excitatory action (HR↑; SV↑; TPR↑) and slow. Levick, 5th ed., 2010

12. Autonomic Control of Heart Rate Berne & Levy, 2008 • Both, VA and SY outputs are tonically active. • In modulating HR, in “normal” subjects at rest, VA output is larger and typically predominates over SY output. • Effect directly and mostly on SA node: • VA: via muscarinic receptors (M2-receptor). • SY: via β-adrenergic receptors (β1-adrenoceptor).

13. III. Sympathetic Signalling

14. Sympathetic Stimulation of SAN • Over several beats, HR↑, AV conduction↑, AP shortening, AP amplitude↑, rate of pacemaker decay↑ (positive -tropisms) – no additional depol. • Recovery from stimulation “slow”: terminated by diffusion of NA in ECF and neuronal re-uptake. • Action via β1-R – α- and β2-R are there but outnumbered 4:1. Hutter & Trautwein, J Gen Physiol 39 (1956):715

15. Sympathetic Effect in Myocyte • HR doubes in this case • AP shortens, no change in AP height or RMP, phase 2 larger, phase 3 faster (”spike and dome AP”). • Sarcoplasma: faster Ca2+ rise and decay and larger [Ca2+]-amplitude. • Contraction: both contracti-lity↑ and rate of relaxation↑ (positive ino- and lusotropic). • Consequence: diastolic filling considerably curtailed (later). Levick, 5th ed., 2011

16. Sympathetic Signalling Levick, 5th ed., 2010 • Linked to Gs signalling cascade: • GαINCREASEScAMP by adenylyl cyclase activation (direct). • Mimicked by caffeine, etc., which block cAMP breakdown. • PKA activation (downstream). • ICS: faster pacing → HR ↑. • If ↑ (direct; open probability ↑) → faster decay of PMP → HR ↑. • IK↑ via PKA phosphorylation → faster repolarisation → HR ↑. • ICaL↑ from PKA phosphorylation → faster decay of PMP → HR ↑. • Myocyte: bigger force production • ICaL↑ from PKA phosphorylation (open probability & MOT): plateau current ↑ → bigger force. • PLB phosphorylated: disinhibits SERCA pump → faster clearance. • RyRphosphoryl.: store release ↑. • Signalling turned-off via phosphatase activity (PP2A) around macromolecular complexes (SERCA, RyR). • Chronic stimulation results in CaMK II activ-ation with different downstream signalling.

17. IV. Parasympathetic Signalling

18. Parasympath. Stimulation of SAN • ± Instantaneous drop in HR: release of ACh – broken down by AChE. • Fast – beat by beat… • In humans, effect restricted ONLY to ICS cells – NOT on myocyte. • Nerve stimulation generates • PMP decay↓ (large) with AP amplit.↓: M2-R. • Nature of hyperpolarisation: controversial. • In most textbooks: AChsuperfusion activates IKACh→ hyperpolarisation due to GIRK activation via Gβγ(fast): M2-AChR (likely extrasynaptic ). • Wrong as not experim. blocked by Cs+ (GIRK). • IJPs, hyperpolarising MP (a few mV). • Mediated by synaptic M2-AChR (IbNa↓). • Nerve stimulation ≠ AChperfusion ! (in most textbooks not differentiated) Bolter et al., AutonNeurosci 94 (2001):93

19. Parasympathetic Signalling • Mostly exerted via M2 activation. • Linked to Gisignalling cascade: • GαDECREASEScAMP (↓) by adenylyl cyclaseinhibition (direct). • “Converse” of β1 activation. • Downstream signalling: PKA activity↓. • ICS: slower pacing • If↓ (direct; open probability↓ → slower decay of PMP. • IK↓ via PKA activity↓ → slower repolarisation, longer AP. • ICaL↓ from PKA activity↓ → slower decay of PMP. • Only at high stimulus rates, IK ACh↓ via direct Gβγ activation → hyperpolarisation (not physiol.). Levick, 5th ed., 2011

20. Summary of Signalling in ICS • Yin & yan around AC (“agonist / antagonist”) • Sympathetic: cAMP↑ • Global effect on heart • Faster decay of PMP • Faster and larger Ca2+ influx • Shorter interval: IK↑ • No additional depolarisation • Parasympathetic: cAMP↓ • Effect only on ICS • Slower decay of PMP • Slower Ca2+ influx • Longer interval: IK↓ • Small hyperpolarisation (IJP) Raff & Levitzky, 1st ed., 2011

21. V. AP under different conditions

22. L-Type Ca2+ Channel Blockers • Verapamil, nifedipine& …ipine • Dose-dependent block of ICaL in clinically relevant doses. • Verapamil better on cardiac tissue. • …ipine(s) better on vessels (vary). • SAN • Prolongs PMP decay and amplitude↓ (small): HR↓ - neg. chronotropic. • AVN • Reduces amplitude and shortens AP: currents↓ for depolarisation of surroun-ding cells: neg. dromotropic. • Cardiac myocyte • Shoulder↓and amplitude↓ (small): ICaL↓ • Force↓: neg. inotropic. Hirthet al., J Mol Cell Cardiol 15(1983):799

23. AP in Hyperkalaemia and Hypoxia • Hyperkalaemia: RMP↑ (EK↑) → • Inactivation of INa → • APs start to resemble those in ICS. • Loss of phase 0 and 1. • Hyp-/anoxia causes • due ATP↓ drop in Na+/K+-ATPase → • IKATP↑ → shorter AP → less Ca2+ influx → contractility↓ → sympathetic activation: HR↑. • store overload: delayed afterdepolarisation • Mechanisms causing overload • Sympathetic reflex activation: ICaL↑ (see later). • ATP↓ → transporter activity↓ (Na/K-ATPase activity↓ and Na/H-exchanger↓) → [Na+]↑ → NCX in reverse → Ca2+ influx: • Depolarisation (arrhythmia). • RyR activation (spontaneous discharge). • Ca2+ clearance into stores↑: overload. Berne & Levy, 2008 Levick (2010), 5th Ed.

24. Take-Home Messages • Ca2+ influx via L-type channels triggers EC coupling. • Contractility and clearance of plasma modulated by 2nd messenger systems. • Autonomic innervation is diffuse (SYM) and specific (VA). • Normally, parasympathetic activity predominates in heart. • β1-AR signal via cAMP↑ to upregulateIf, IK and ICaL. • M2-AChR signal via cAMP↓ to downregulateIKand ICaL. • There is a difference between action of synaptic & extrasynapticAChR. • Ca2+ channel blockers are neg. ino-, dromo- and chronotrop.

25. MCQ Victor Helms, a 26 year-old is admitted to the Emergency Department (ED) after being stabbed in his neck. The ED physician diagnoses that his left vagal nerve may likely have been damaged? Which of the following statements is most consistent with this diagnosis? • Sinus tachycardia. • Increased cardiac contractility. • Tachycardia with shortened PR interval. • Ventricular bradycardiawith large P waves. • Sinus bradycardia with prolonged QRS complex.

26. That’s it folks…

27. MCQ Victor Helms, a 26 year-old is admitted to the Emergency Department (ED) after being stabbed in his neck. The ED physician diagnoses that his left vagal nerve may likely have been damaged? Which of the following statements is most consistent with this diagnosis? • Sinus tachycardia. • Increased cardiac contractility. • Tachycardia with shortened PR interval. • Ventricular bradycardiawith large P waves. • Sinus bradycardia with prolonged QRS complex.