Elsevier

Autonomic Neuroscience

Volume 164, Issues 1–2, 28 October 2011, Pages 96-100
Autonomic Neuroscience

Short communication
The muscarinic-activated potassium channel always participates in vagal slowing of the guinea-pig sinoatrial pacemaker

https://doi.org/10.1016/j.autneu.2011.05.009Get rights and content

Abstract

Controversy persists regarding participation of the muscarinic-activated potassium current (cKACh) in small and moderate vagal bradycardia. We investigated this by (i) critical examination of earlier experimental data for mechanisms proposed to operate in modest vagal bradycardia (modulation of If and inhibition of a junctional Na+ current) and (ii) experiments performed on isolated vagally-innervated guinea-pig atria. In 8 superperfused preparations, 10-s trains of vagal stimulation (1 to 20 Hz) produced a bradycardia that ranged from 1 to 80%. Hyperpolarisation of sinoatrial cells accompanied bradycardia in 65/67 observations (linear correlation between bradycardia and increase in maximum diastolic potential (mV) = 0.076x%; R2 = 0.57; P < 0.001). In bath-mounted preparations single supramaximal stimuli to the vagus immediately and briefly increased pacemaker cycle length in 7 of 18 preparations. This response was eliminated by 300 nM tertiapin-Q. Trains of 10 single supramaximal vagal stimuli applied at 1-s intervals caused progressive increase in overall cycle length during the train; immediate and brief increases in cycle length occurred following some stimuli. Immediate brief responses and part of the slower response to the stimulus train were removed by 300 nM tertiapin-Q. Summary: experimental data shows that small and modest vagal bradycardia is accompanied by hyperpolarisation of the pacemaker cell which is severely attenuated by tertiapin-Q. These observations support the idea that activation of IKACh occurs at all levels of vagal bradycardia. Contradictory conclusions from earlier studies may be attributed to the nature of experimental models and experimental design.

Introduction

Controversy still surrounds the contributions made by a variety of ionic mechanisms to slowing of the cardiac pacemaker by the cardiac parasympathetic nerves (PSNS). Following its release from the parasympathetic postganglionic varicosities in the sinoatrial node (SAN), acetylcholine (ACh) binds to muscarinic receptors and the resulting alteration of a number of membrane currents reduces the rate at which the SAN generates action potentials (vagal bradycardia; throughout this paper the term vagal is applied specifically to the efferent cardiac parasympathetic fibres which reach the heart after travelling down the cervical vagus nerve).

Muscarinic stimulation can activate a K+ current, IKACh, that is normally outward and will hyperpolarise the pacemaker cells and lengthen slow diastolic depolarization (SDD) (Pfaffinger et al., 1985, Boyett et al., 1995, Zhang et al., 2002). By reducing the intracellular concentration of cyclic AMP, muscarinic stimulation can also reduce the basal current carried by several types of channels and an exchanger which participate in pacemaking, and which carry net inward current during SDD. Thus, muscarinic stimulation may reduce the slope of SDD by reducing current carried by the hyperpolarisation-activated current (If; DiFrancesco and Tromba, 1987), L-type Ca2+ current (ICaL; Hartzell, 1988, Petit-Jacques et al., 1993, Zaza et al., 1996), Na+/Ca2+ exchange current (NCX, INaCa; Sanders et al., 2006, Maltsev and Lakatta, 2010, van Borren et al., 2010), and the sustained inward current (Ist; Toyoda et al., 2005).

For many years the primary mechanism for the bradycardia was considered to be the activation of IKACh. Then, in 1976, Noma and Irisawa described a novel current in pacemaker cells, the hyperpolarisation-activated current termed Ifunny or Iqueer (If). Subsequently this current was shown to be modulated by autonomic agonists (Brown et al., 1979, DiFrancesco and Tromba, 1988), and in a study published in 1989 DiFrancesco et al. concluded that reduction of If rather than activation of IKACh was the primary mechanism for vagal bradycardia. The model developed by Hirst's group also discounted involvement of activation of IKACh in small to moderate vagal bradycardia (Edwards et al., 1993). Hirst and colleagues have proposed that there are specialised junctional muscarinic receptors in the vicinity of parasympathetic varicosities (Hirst et al., 1996); stimulation of these receptors is considered to produce a bradycardia that involves neither activation of IKACh, nor a reduction in If (Campbell et al., 1989, Bywater et al., 1990), but the inhibition of an inward current carried by Na+ (Edwards et al., 1993). ACh can also bind to non-muscarinic receptors producing a reduction in IK, increasing action potential duration and cycle length (Freeman and Kass, 1995, Lei et al., 1999). However action potential configuration is not modified by vagal stimulation (Campbell et al., 1989), so involvement of IK in vagal bradycardia seems unlikely.

In this paper we present further evidence that IKACh is a significant, and probably the major participant in vagal bradycardia, including even small degrees of bradycardia. As well as providing this additional experimental evidence, we discuss and critique in some detail the evidence for two other mechanisms behind vagal bradycardia — a reduction in If and an inhibition of a background sodium current.

Section snippets

Experiment 1

Experiments were performed on atrial preparations from young guinea-pigs of either sex (150–200 g), and were approved by the animal experimentation ethics committee at the University of Melbourne. Preparations were dissected in the cold from animals that were stunned then desanguinated. Both atria, with vagus nerves attached were pinned out in a shallow dish perfused with Krebs–Henseleit solution at 35 °C, and gassed with 95% O2, 5% CO2. The vagus nerves were passed through an electrode for

Pacemaker hyperpolarisation and vagal bradycardia

Data were obtained for vagal bradycardia that ranged from just detectable bradycardia to over 80% maximum response (Fig. 1; maximum response is arrest). In 65 of 67 responses there was a measureable increase in MDP (≥ 0.5 mV), including 17 observations in which bradycardia was less than 20%. A linear relationship fitted to the data indicated that on average there was a 0.076 mV increase in MDP for each 1% of bradycardia effected by vagal stimulation (y = 0.076 + 0.619; R2 = 0.57; P < 0.0001). We obtained

Discussion

In this paper we present evidence that IKACh in SAN pacemaker cells plays a primary role in vagal bradycardia. Thus we conclude that activation of IKACh will always occur when there is activity in the parasympathetic innervation of the SAN, and that activation of IKACh provides the predominant mechanism for vagal bradycardia over the full range of vagal influence on heart rate. From previous experiments in which tertiapin-Q was used to block IKACh, we concluded that IKACh is probably

Acknowledgement

This work was carried out with financial support from the National Heart Foundation of New Zealand, the University of Otago, and the Australian NH and MRC.

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