The role in the heart of the cardiac isoform of the cystic fibrosis transmembrane conductance regulator (CFTR), which underlies a protein kinase A-dependent CL current (I-CLPKA) in cardiomyocytes, remains unclear. The identification of a CFTR-selective inhibitor would provide an important tool for the investigation of the contribution of CFTR to cardiac electrophysiology. GlyH-101 is a glycine hydrazide that has recently been shown to block CFTR channels but its effects on cardiomyocytes are unknown. Here the action of GlyH-101 on cardiac I-CLPKA and on other ion currents has been established. Whole-cell patch-clamp recordings were made from rabbit isolated ventricular myocytes. GlyH-101 blocked I-CLPKA in a concentration- and voltage-dependent fashion (IC50 at +100 mV = 0.3 + 1.5 mu M and at -100 mV = 5.1 +/- 1.3 mu M). Woodhull analysis suggested that GlyH-101 blocks the open pore of cardiac CFTR channels at an electrical distance of 0.15 +/- 0.03 from the external membrane surface. A concentration of GlyH-101 maximally effective against I-CLPKA (30 mu M) was tested on other cardiac ion currents. Inward current at -120 mV, comprised predominantly of the inward-rectifier background K+ current, I-K1, was reduced by similar to 43% (n = 5). Under selective recording conditions, the Na+ current (I-Na) was markedly inhibited by GlyH-101 over the entire voltage range (with a fractional block at -40 mV of similar to 82%; n = 8). GlyH-101 also produced a voltage-dependent inhibition of L-type Ca2+ channel current (I-Ca.L); fractional block at +10 mV of similar to 49% and of similar to 28% at -10 mV; n = 11, with a similar to-3 mV shift in the voltage-dependence of I-Ca.L activation. Thus, this study demonstrates for the first time that GlyH-101 blocks cardiac I-CLPKA channels in a similar fashion to that reported for recombinant CFTR. However, inhibition of other cardiac conductances may limit its use as a CFTR-selective blocker in the heart. (C) 2011 Elsevier Inc. All rights reserved.