The delayed rectifier current, IK, responsible for terminating the plateau phase of the cardiac action potential, is composed of two components; the slowly activating component, IKs, and the rapidly activating component, IKr. IKr is the primary target of Class III antiarrhythmic agents, inhibition of which prolongs cardiac action potential and reduces the probability of reentrant arrhythmia induction. However, Class III antiarrhythmic agents can also excessively prolong the QT interval and precipitate a potentially lethal arrhythmia called torsade de pointes (TdP). Furthermore, mutations at the gene coding for IKr can cause congenital long QT syndrome and TdP. Heterologous expression of the human ether-a-go-go -related gene (HERG) elicits currents that closely resemble native IKr. The work presented in this thesis explores in detail heterologously expressed HERG channels, in particular their biophysical and pharmacological properties. We first probed HERG channels with the divalent cation Ba 2+ and the potent Class III antiarrhythmic agent dofetilide to examine the voltage-dependent pore accessibility. Our experimental and mathematical modeling data indicate that Ba2+ blocked wild-type HERG channels in their closed and open states but channel inactivation relieved Ba 2+ block. In contrast to previous reports indicating that dofetilide only blocked the open state of HERG channels, our data clearly show that dofetilide interacted with both the open and inactivated states. We then examined the need for association between the recently cloned MiRP1 (minK-related peptide 1) subunit in imparting properties similar to that of native IKr to HERG. We demonstrated that the pharmacological characteristics of HERG channels expressed in CHO cells is very similar to IKr recorded from guinea pig myocytes. The minor discrepancies between the cloned and native currents (i.e. deactivation kinetics and voltage dependence of activation) were not rectified by MiRP1 co-expression. Thu / In summary, our findings demonstrate that HERG channels are accessible in a state-dependent manner from the extracellular side by Ba2+ ions and the intracellular side by dofetilide. The characteristics of HERG channels that govern its sensitivity to drug blockers do not appear to require an association with its putative MiRP1 beta-subunit. The latter finding has important implications for screening of novel drugs for an early determination of their potential to prolong the QT interval, in particular those agents that are not meant to possess IKr inhibitory properties. Inhibition of HERG channels expressed in a mammalian cell line will provide a very close approximation of the sensitivity of native IKr to these agents.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.38436 |
| Date | January 2002 |
| Creators | Weerapura, Manjula. |
| Contributors | Nattel, Stanley (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Pharmacology & Therapeutics.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001862324, proquestno: NQ78797, Theses scanned by UMI/ProQuest. |
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