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Actions of RSD1015 and RSD1000 on macroscopic currents in rat ventricular myocytes

University of British Columbia

Two novel agents RSD1015 and RSD1000, compounds with potent antiarrhythmic effects in vivo, have been studied in rat ventricular myocytes using the whole-cell patch clamp technique. Both drugs exhibit potent mixed blockade effects on potassium (l[sub to]) and sodium current (l[sub Na]), with little or no effects on other currents including l[sub K1] and l[sub Ca]. The effects of these drugs on different kinetic states of; potassium and sodium channels were investigated. The ischemic-selectivity of RSD1015 and RSD1000 was also studied using solutions of varying pH. The rationale for this being that physicochemical properties of compounds, especially their pKa, act as one of the important determinants of ischemia-selective antiarrhythmic activity. At bath solution of pH 7.4, with extracellular concentrations from 1 to 30 μM, both RSD1015 and RSD1000 reduced the inactivation time constant (τ) of the transient outward potassium current (l[sub to]) with an EC₅₀ values 1.3 ± 0.4 μM and 5.8 ±0.7 μM respectively. The voltage dependence of both activation and inactivation of lto and the time course of recovery from current inactivation were not significantly changed by the RSD compounds at concentrations up to 10 μM. The inhibition of l[sub to] increased with time during depolarizing pulses, indicating that RSD1015 and RSD1000 interacted with the open state of the channel which can be characterized by k1 (onset) and k-1 (offset) rate constants. From the study, the k₁ and k₋₁ for RSD1015 on l[sub to] blockade are 16,3±3.4x 10⁶ M⁻¹ s⁻¹ and 27.6 + 4.3 s⁻¹, respectively and the k[sub d] ([sub d]=k₋₁/k₁) is 1.7 μM. The values of the k₁, and k₋₁ of RSD1000 were 8.1 ± 1.9 x 10⁶ M⁻¹ s⁻¹, 52.3 ± 3.3 s⁻¹ respectively, the k[sub d] is 6.5 μM. The decreased peak amplitudes of l[sub to] by agents are also consistent with rapid open channel blockade. In addition, RSD1015 and RSD1000 decreased the peak amplitudes of the inward sodium current (l[sub Na]) with an EC₅₀ of 4.1 ± 0 . 9 μM and 1.5 + 0.3 μM, respectively. This action was not accompanied by any change in the voltage dependence of activation, but both agents cause hyperpolarizing shifts in the V[sub1/2] of l[sub Na] inactivation curve. Use-dependence of l[sub Na] was evident in the presence of RSD1015 and RSD1000 with a stimulating frequency of 5 Hz or higher; the onset of the use-dependent block is rapid. The blockade of l[sub Na] by these agents indicated that they interacted mainly with the inactivated and activated states of sodium channel. In an acid solution (pH 6.4), the potency of RSD1015 and RSD1000 on l[sub to] blockade did not significantly change; however, the potency for RSD1000 on l[sub Na] blockade was significantly enhanced from the EC₅₀ of 1.5 ± 0.3 μM ( pH 7.4) to 0 .5 ± 0.1 μM (pH 6.4). Therefore, RSD1000 has higher ischemia-selectivity compared to RSD1015. Use-dependent blockade of l[sub Na] by RSD1000 also significantly increased under acidic conditions (pH 6.4) compared with the effects in normal solution of pH 7.4. This ischemia-selectivity of RSD1000 in acidic conditions is consistent with in vivo studies and can provide clinical usefulness, especially in the mechanisms of cardiac arrhythmogenesis. The ischemic-selectivity of RSD1000 appears to be the function of its pKa value, which causing different percentage of molecule protonated in the solution of pH 7.4 or 6.4. The pH-dependent interaction of the tertiary amine sodium channel blockers has been suggested to be a mechanism of pH modulated actions of RSD1000. Overall, this study showed that RSD1015 and RSD1000 are potent mixed blockers of l[sub to] and l[sub Na] in rat ventricular myocytes. These results demonstrated the important mechanisms of antiarrhythmic effects of these two agents.

Thesis/Dissertation

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2009-04-29

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http://hdl.handle.net/2429/7689

Pharmacology

University of British Columbia

UBCV

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