M. Hussain et al., Effects of the protein kinase A inhibitor H-89 on Ca2+ regulation in isolated ferret ventricular myocytes, PFLUG ARCH, 437(4), 1999, pp. 529-537
We investigated the effects of a protein kinase A (PKA) inhibitor, H-89 {N-
[2-(p-bromocinnamylamino)ethyl]-5-iso-quinolinesulphonamide}, on Ca2+ regul
ation in Fura-2-loaded ferret myocytes. H-89 (10 mu mol/l) decreased the am
plitude of the Fura-2 transient to 28.2+/-4.3% (P<0.001) of control and pro
longed its duration, characterized by a decrease in the rate of decline of
Ca2+ to diastolic levels: t(1/2) increased from 311+/-35 ms to 547+/-43 ms
(P<0.001, n=7). Reduced Ca2+ uptake by the sarcoplasmic reticulum (SR) in t
he presence of H-89 was also indicated by a decrease in the SR Ca2+ content
, as assessed with caffeine. The apparent slowing of the SR Ca2+-ATPase was
not caused by changes in phosphorylation of phospholamban (PLB). However,
Ca2+ uptake in microsomal vesicles prepared from canine hearts and fast-twi
tch rat skeletal muscle (which lacks PLB) was decreased by 34.1 and 46.8% (
n=3), respectively, suggest ing that H-89 has a direct inhibitory effect on
the SR Ca2+-ATPase. In electrophysiological experiments, 5.0 mu mol/l H-89
decreased the L-type Ca2+ current (I-Ca) by 39.5% (n=6) and slowed the ups
troke of the action potential and, in some cases, caused loss of excitabili
ty without changes in the resting membrane potential. In summary, data show
that [Ca2+](i) regulation, and hence contraction, is sustained by PKA-medi
ated phosphorylation, even in the absence of beta-agonists. However, the us
e of H-89 as a tool to study the role of this signalling pathway is limited
by the non-specific effects of H-89 on the SR Ca2+-ATPase.