Bin-1 Expression in Normal Rat Cardiac Myocytes and in Myocytes with Reduced T-Tubule Density Due to Cell Culture or Heart Failure

Abstract

Bridging Integrator-1 (BIN-1), member of the BAR domain protein superfamily, has been implicated in the formation and maintenance of membrane invaginations in skeletal and cardiac muscle cells. However, it is not clear whether BIN-1 expression and t-tubule density are correlated in all circumstances. The aim of the present study was to investigate the relationship between t-tubule density and BIN-1 expression in (i) fresh atrial and ventricular myocytes (ii) ventricular myocytes cultured for 24-72 hours and (iii) ventricular myocytes from animals with monocrotaline-induced right heart failure (HF). Confocal microscopy was used to assess t-tubule density and BIN1 expression in rat myocytes using di-8-ANEPPS fluorescence or immunofluorescence respectively. In fresh ventricular myocytes, a regular t-tubule network was apparent and BIN-1 exhibited a corresponding striated pattern (n=10). In atrial myocytes t-tubules were typically absent and BIN-1 immunofluorescence was only detectable in the peripheral sarcolemma (n=20). In ventricular myocytes after 24, 48 or 72 hrs cell-culture, t-tubule density was reduced to 61 ± 1.4%, 26 ± 0.6% and 11 ± 0.5% respectively (n=10, 8 and 6, P<0.05), while mean cell area did not change significantly. Correspondingly, BIN1 expression was significantly reduced in ventricular myocytes at 48 (to 17 ± 0.14%, n=7, P<0.05) and 72 hrs (to 16 ± 0.09%; n=5, P<0.05) compared with control. In right ventricular myocytes, from rats with right HF and reduced t-tubule density, BIN1 expression decreased significantly (69 ± 1.3%; n=7, P<0.05) compared with saline-injected rats. These results suggest that where t-tubules are absent (e.g. atrial cells) or in circumstances where t-tubule density is decreased (cell culture, HF) BIN-1 expression is correspondingly reduced. This supports the hypothesis that BIN-1 has an important role in formation and maintenance of t-tubules.