Automaticity and conduction properties of bio-artificial pacemakers assessed in an in vitro monolayer model of neonatal rat ventricular myocytes

Abstract

A better understanding of the ionic mechanisms for cardiac automaticity can lead to better strategies for engineering bio-artificial pacemakers. Here, we attempted to better define the relative contribution of I(f) and I(K1) in the generation of spontaneous action potentials (SAPs) in cardiomyocytes (CMs). Monolayers of neonatal rat ventricular myocytes (NRVMs) were transduced with a recombinant adenovirus (Ad) to express a gating-engineered HCN1 construct (HCN1-DeltaDeltaDelta) for patch-clamp and multielectrode array (MEA) recordings. Single NRVMs exhibited a bi-phasic response in the generation of SAPs (62.6 +/- 17.4 b.p.m., Days 1-2; 194.3 +/- 12.3 b.p.m., Days 3-4; 73% quiescent, Days 9-10). Although automaticity time-dependently decreased and subsequently ceased, I(f) remained fairly stable (-5.2 +/- 1.1 pA/pF, Days 1-2; -5.1 +/- 1.4 pA/pF, Days 7-8; -4.3 +/- 1.3 pA/pF, Days 13-14). In contrast, I(K1) declined rapidly (from -16.9 +/- 2.7 pA/pF on Days 1-2 to -4.4 +/- 1.6 pA/pF on Days 5-6). Maximum diastolic potential/resting membrane potential (r = 0.89) and action potential duration at 50% (APD(50), r = 0.73) and 90% (APD(90), r = 0.75) but not the firing rate (r = -0.3) were positively correlated to the I(K1). Similarly, monolayer NRVMs ceased to spontaneously fire after long-term culture. Ad-HCN1-DeltaDeltaDelta transduction restored pacing in silenced individual and monolayer NRVMs but with reduced conduction velocity and field potential amplitude. We conclude that the combination of I(K1) and I(f) primes CMs for bio-artificial pacing by determining the threshold. However, I(f) functions as a membrane potential oscillator to determine the basal firing frequency. Future engineering of automaticity in the multicellular setting needs to have conduction taken into consideration.