Characterization of the cyclic nucleotide phosphodiesterase subtypes involved in the regulation of the L-type Ca 2+ current in rat ventricular myocytes

Ignacio Verde (1) , Grégoire Vandecasteele (1) , Frank Lezoualc'H (1) , Rodolphe Fischmeister (1)

1 CARPAT - Signalisation et physiopathologie cardiovasculaire

The effects of several phosphodiesterase (PDE) inhibitors on the L-type Ca current (ICa) and intracellular cyclic AMP concentration ([cAMP]i) were examined in isolated rat ventricular myocytes. The presence of mRNA transcripts encoding for the different cardiac PDE subtypes was confirmed by RT–PCR. IBMX (100 μM), a broad-spectrum PDE inhibitor, increased basal ICa by 120% and [cAMP]i by 70%, similarly to a saturating concentration of the β-adrenoceptor agonist isoprenaline (1 μM). However, MIMX (1 μM), a PDE1 inhibitor, EHNA (10 μM), a PDE2 inhibitor, cilostamide (0.1 μM), a PDE3 inhibitor, or Ro 20–1724 (0.1 μM), a PDE4 inhibitor, had no effect on basal ICa and little stimulatory effects on [cAMP]i (20–30%). Each selective PDE inhibitor was then tested in the presence of another inhibitor to examine whether a concomitant inhibition of two PDE subtypes had any effect on ICa or [cAMP]i. While all combinations tested significantly increased [cAMP]i (40–50%), only cilostamide (0.1 μM)+Ro20-1724 (0.1 μM) produced a significant stimulation of ICa (50%). Addition of EHNA (10 μM) to this mix increased ICa to 110% and [cAMP]i to 70% above basal, i.e. to similar levels as obtained with IBMX (100 μM) or isoprenaline (1 μM). When tested on top of a sub-maximal concentration of isoprenaline (1 nM), which increased ICa by (∼40% and had negligible effect on [cAMP]i, each selective PDE inhibitor induced a clear stimulation of [cAMP]i and an additional increase in ICa. Maximal effects on ICa were ∼8% for MIMX (3 μM), ∼20% for EHNA (1–3 μM), ∼30% for cilostamide (0.3–1 μM) and ∼50% for Ro20-1724 (0.1μM). Our results demonstrate that PDE1-4 subtypes regulate ICa in rat ventricular myocytes. While PDE3 and PDE4 are the dominant PDE subtypes involved in the regulation of basal ICa, all four PDE subtypes determine the response of ICa to a stimulus activating cyclic AMP production, with the rank order of potency PDE4>PDE3>PDE2>PDE1.

Keywords: rat heart; L-type Ca2+ current; intracellular cAMP; phosphodiesterase subtypes; phosphodiesterase inhibitors; MIMX (PDE1 inhibitor); EHNA (PDE2 inhibitor); cilostamide (PDE3 inhibitor); Ro 20-1724 (PDE4 inhibitor); Beta-adrenoceptor agonist

Domains: Cardiology and cardiovascular system; Biochemistry, Molecular Biology; Pharmacology

Ignacio Verde, Grégoire Vandecasteele, Frank Lezoualc'H, Rodolphe Fischmeister. Characterization of the cyclic nucleotide phosphodiesterase subtypes involved in the regulation of the L-type Ca 2+ current in rat ventricular myocytes. British Journal of Pharmacology, 1999, 127 (1), pp.65-74. ⟨10.1038/sj.bjp.0702506⟩. ⟨hal-03612114⟩