The hyperpolarization-activated current I_f in ventricular myocytes of non-transgenic and β₂-adrenoceptor overexpressing mice

Abstract.

In transgenic mice (TG4) overexpressing the human β₂-adrenoceptor (β₂-AR), unoccupied receptors are supposed to activate spontaneously the signalling cascade, leading to enhanced levels of cAMP. This second messenger shifts activation curves of the hyperpolarization-activated current I _f towards less negative potentials. Here, we characterize I _f of ventricular myocytes from non-transgenic littermate (LM) and TG4 mice and investigate whether I _f is modulated by spontaneous β₂-AR signalling.

I_f was activated in whole-cell voltage-clamp experiments during test steps ranging from –65 mV to –135 mV (holding potential: –55 mV; 36°C). In TG4 the maximum amplitude was fivefold larger than in LM myocytes (–1.10±0.11 pA/pF vs. –0.22±0.04 pA/pF at –135 mV), and the potential for half-maximum I_f current (V_I0.5) was less negative (–100.5±1.0 mV in TG4 vs. –108.4±2.6 mV in LM). (-)-Isoproterenol (1 µM) shifted V_I0.5 of LM myocytes by 10.4 mV towards less negative potentials but had no significant effect in TG4. However, the inverse β₂-AR agonist ICI 118,551 (300 nM) shifted V_I0.5 of TG4 myocytes to values observed in LM under control conditions, suggesting a relation to spontaneously active β₂-ARs. Enhanced expression of hyperpolarization-activated and cyclic nucleotide gated channels (HCN) could contribute to increased maximum I_f amplitude in TG4 myocytes. Semi-quantitative RT-PCR analysis demonstrated a 1.8-fold elevation of HCN4 mRNA and no significant change for HCN2 mRNA in TG4 ventricle. Cardiac hypertrophy was not detected in TG4 mice investigated here.

We conclude that spontaneous β₂-AR signalling in hearts of TG4 mice shifts I _f current-voltage relation towards less negative potentials. Increased maximum I _f amplitude in TG4 myocytes is in line with enhanced expression of HCN channels. Both mechanisms could contribute to larger inward current at physiological diastolic potentials.