P21 activated kinase-1 deficiency in cardiomyocytes increases in susceptibility to ventricular arrhythmogenesis in mice
Y. Wang1, H. Tsui1, W. Liu1,2, Y. Shi3, R. Wang3, Y. Zhang1, R. Solaro4, Y. Ke4, H. Zhang5, E. Cartwright1, X. Wang2, M. Lei1
1. Cardiovascular Research Group, School of Biomedicine,, University of Manchester, Manchester, United Kingdom. 2. Faculty of Life Sciences & Manchester Academic Health Sciences Centre, University of Manchester, Manchester, United Kingdom. 3. Department of Cardiovascular Medicine, Union Hospital, Wuhan, China. 4. Department of Physiology and Biophysics and Centre for Cardiovascular Research,, University of Illinois, Chicago, Illinois, United States. 5. School of Physics and Astronomy,, University of Manchester, Manchester, United Kingdom.
Our recent studies indicate a novel role of P21 activated kinase-1 (Pak1) in regulating cardiac electrical and contractile functions. This following-up study thereby aims at further clarify the mechanisms underlying the critical roles of Pak1 in regulating cardiac electrical function and ventricular arrhythmogenesis under stress conditions in mice with genetic modifications of Pak1. Mice carrying a ventricular cardiomyocyte-restricted deletion of Pak1 (Pak1cko) displayed relative high intrinsic heart rate with normal electrocardiography (ECG) parameters including P-R, QRS and QT intervals at baseline condition compared with control Pak1f/f mice. To evaluate ventricular arrhythmic vulnerability, both Pak1cko and Pak1f/f (control) mice were subjected to either acute treatment of isoprenaline (ISO) at in vivo condition in anesthetised mice (1 mg/kg, i.p) with 1.5-2% isoflurane in oxygen, ex vivo heart (10 -50 nM) or chronic treatment by mini-osmotic pumps (Charles river labs, UK) at concentration 100 mg/ml ISO per gram of mouse for the 14 day pump. For in vivo ECG recordings, three-lead limb ECGs were recorded through subcutaneous 27 gauge needle electrodes using a Powerlab 26T system (AD Instruments, Hastings, UK). The resulting digital recordings (16 bit, 2 kHz/channel) were analyzed using the Chart version 5.0 program (AD Instruments) to obtain the signal-averaged ECG. T-wave durations and QT intervals were measured in lead II attached were placed on three of the four limbs. The ex vivo heart studies involve ECG recorded by ECG electrodes (Harvard Apparatus (UK) Ltd, Cambridge, UK) positioned onto atrial and ventricle with Chart version 5.0 program. Two programmed electrical stimulation (PES) protocols were processed to the hearts, 1). Ventricular effective refractory period was determined by S1S2 pacing that begin with a drive cycle of 100 ms with an S2 coupled at 100 ms and bring in S2 by 1 ms until there is loss of ventricular capture. 2). Ventricular burst pacing (S1S1 pacing) deliver ventricular pacing with three trains of 20 S1 at a cycle length of 100 ms with a pause of one second in between trains. Then repeat the protocol with shortened cycle length from 100ms to 30ms by 10ms step. Followed by programmed electrical stimulation (PES), electrocardiographic analysis of both anesthetized and conscious animals revealed a high occurrence of ventricular arrhythmias such as multiple episodes of ventricular tachycardia (VT) or ventricular fibrillation (VF) in Pak1cko mice but not in control Pak1f/f littermates. Action potentials (APs) recorded from ventricular myocytes isolated from and Pak1cko and Pak1f/f hearts, a delayed after-depolarization (DAD) type AP and spontaneous APs can be observed from PAK1cko myocytes, but not control Pak1f/f myocytes. Thus our study suggests a crucial role of Pak1 signalling in preventing stress associated ventricular arrhythmogenesis.