Compartmentation of the β-adrenergic Signal by phosphodiesterases in Adult Rat Ventricular Myocytes
Schwartz, Jesse Milo
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Previous studies have suggested that phosphodiesterase (PDE) hydrolysis of cyclic adenosine monophosphate (cAMP) is important in the generation of specific and segregated cAMP signals within cells. The purpose of this study was to determine if PDE compartmentation was important in cardiac ventricular myocytes. Therefore, we investigated the effects of β-adrenergic (β-AD) stimulation with isoproterenol in the presence of cilostamide, a PDE3 inhibitor, or Ro 20-1724, a PDE4 inhibitor, on unloaded cell shortening, L-type calcium currents and intracellular calcium levels in freshly dissociated adult rat ventricular myocytes. PDE3 inhibition resulted in a 216 ± 17 % (n=8) increase in unloaded cell shortening after ten minutes of isoproterenol exposure, whereas isoproterenol produced a statistically smaller increase of 155 ± 12 % (n=8) in the presence of PDE4 inhibition. There was a non-significant trend for PDE4 inhibition to produce larger increases in calcium currents (179 ± 17 % (n=4) of controls) than PDE3 inhibition (155 ± 10 % (n=6) of controls). Both PDE3 and PDE4 inhibitors had similar effects on isoproterenol-stimulated increases of calcium transient amplitude with values of 209 ± 14 % (n=8) and 185 ± 12 % (n=8), respectively. Determination of sarcoplasmic reticulum (SR) calcium load using caffeine pulse experiments demonstrated that PDE4 inhibition and isoproterenol superfusion produced a statistically larger increase in SR-calcium loading (139 ± 9 % (n=6)) than PDE3 inhibition and isoproterenol superfusion (113 ± 9 % (n=6)). These results suggest that PDE3 may be active in proximity to the contractile apparatus of cardiac myocytes, whereas PDE4 may be localized in a domain consisting of the L-type calcium channel and junctional SR. Consequently, our study provides functional evidence for differential localization of PDE isoforms in cardiac myocytes.