Munir Hussain

Greater antiarrhythmic activity of acute 17β-estradiol in female than male anaesthetized rats: correlation with Ca2+ channel blockade

Female sex hormones may protect pre‐menopausal women from sudden cardiac death. We therefore investigated the effects of the main female sex hormone, 17β‐estradiol, on ischaemia‐induced cardiac arrhythmias and on the L‐type Ca2+ current (ICaL).

The role of constitutive PKA-mediated phosphorylation in the regulation of basal ICa in isolated rat cardiac myocytes

1 Pharmacological inhibitors of protein kinase A (PKA) and protein phosphatases 1/2A were used to determine whether basal L‐type Ca2+ current (ICa) observed in the absence of exogenous β‐adrenergic receptor stimulation is sustained by PKA‐mediated phosphorylation. Amphotericin B was used to record whole‐cell ICa in the perforated patch‐clamp configuration. 2 Calyculin A and isoprenaline (both 1 μmol l−1) increased basal ICa (P<0.05), whereas H‐89 inhibited ICa in a concentration‐dependent manner with an IC50 ∼5 μmol l−1. H‐89 also inhibited the response to 1.0 μmol l−1 isoprenaline, although relatively high concentrations (30 μmol l−1) were required to achieve complete suppression of the response. 3 Double‐pulse protocols were used to study the effects of 10 μmol l−1 H‐89 on time‐dependent recovery of ICa from voltage‐dependent inactivation as well as the steady‐state gating of ICa. T0.5 (time for ICa to recover to 50% of the preinactivation amplitude) increased in the presence of H‐89 (P<0.05) but was unaffected by calyculin A or isoprenaline. 4 Steady‐state activation/inactivation properties of ICa were unaffected by 10 μmol l−1 H‐89 or 1 μmol l−1 calyculin A, whereas isoprenaline caused a leftward shift in both curves so that V0.5 for activation and inactivation became more negative. 5 Data show that basal ICa is regulated by cAMP‐PKA‐mediated phosphorylation in the absence of externally applied β‐receptor agonists and that relatively high concentrations of H‐89 are required to fully suppress the response to β‐adrenergic receptor stimulation, thereby limiting the value of H‐89 as a useful tool in dissecting signalling pathways in intact myocytes.

Effect of Streptozotocin-Induced Type 1 Diabetes Mellitus on Contraction, Calcium Transient, and Cation Contents in the Isolated Rat Heart

Abstract:  Cations play major physiological and biochemical roles in the excitation–contraction coupling processes in the heart. This study investigated the effect of streptozotocin (STZ)‐induced type I diabetes mellitus (DM) on contraction, calcium transient [Ca2+]i, and cation contents in the isolated rat heart compared to age‐matched control. Diabetes rats weighed significantly (P < 0.05) less compared to control. They also had significantly (P < 0.05) elevated blood glucose compared to control. The whole heart, as well as the atria, right and left ventricles of the diabetic heart weighed significantly (P < 0.05) less compared to hearts from age control rats. The force of contraction and time to peak (t‐pk) contraction in diabetic ventricular myocytes increased significantly (P < 0.05) compared to control. By contrast, these parameters did not change for the Ca2+ transient except for the time to half (t½) relaxation. The levels of sodium (Na+), potassium (K+), calcium (Ca2+), magnesium (Mg2+), iron (Fe2+), copper (Cu2+), and zinc (Zn2+) in the hearts varied from diabetic compared to control animals. The results indicate that 6–8 weeks of STZ‐induced DM is associated with marked changes in contraction and in cation contents of the heart. The delay in the t½ relaxation of the Ca2+ transient may be responsible for the elevated contraction seen in the diabetic heart. Moreover, the changes in cation contents in the heart may be responsible for abnormal cardiac rhythms and activity during DM.

H-89 inhibits transient outward and inward rectifier potassium currents in isolated rat ventricular myocytes.

1 Voltage clamp was used to investigate the effects of N‐[2‐p‐bromo‐cinnamylamino)ethyl]‐5‐isoquinolinesulfonamide (H‐89), a potent inhibitor of PKA, on transient outward K+ current (Ito) and inward rectifying K+ current (IK1) in rat cardiac muscle. 2 Initial experiments, performed using descending voltage ramps, showed that H‐89 inhibited both the outward and inward ramp currents in a concentration‐dependent manner at concentrations between 5 and 60 μmol l−1. A similar degree of inhibition was observed when Ito and IK1 were recorded using square wave depolarising and hyperpolarising voltage steps, respectively. 3 The IC50 was 35.8 μmol l−1 for Ito and 27.8 μmol l−1 for IK1 compared to 5.4 μmol l−1 for L‐type Ca2+ current (ICa). The Hill coefficients for Ito, IK1 and ICa were −1.97, −1.60 and −1.21, respectively. In addition to inhibiting Ito amplitude, H‐89 also accelerated the time to peak and the rate of voltage‐dependent inactivation so that the time course of Ito was abbreviated. 4 Paired‐pulse protocols were performed to study the effects of H‐89 on steady‐state activation and inactivation as well as recovery from voltage‐dependent inactivation. H‐89 produced a concentration‐dependent rightward shift in voltage‐dependent activation but had no significant effect on steady‐state inactivation. Recovery from voltage‐dependent inactivation was delayed, although this was only visible at the highest concentration (60 μmol l−1) used. 5 In experiments investigating the effects of elevated cyclic AMP, the β‐adrenergic agonist isoprenaline and the phosphatase inhibitor calyculin A had no major effects on Ito or IK1. 6 Data suggest that the effects of H‐89 on K+ currents are more complex than simple inhibition of PKA‐mediated phosphorylation.