Transverse tubules (t-tubules) are regular invaginations of the surface sarcolemma membrane of ventricular cardiac myocytes. T-tubules enable a rapid and synchronous release of calcium (Ca2+) throughout the cell, thus facilitating contraction. Whilst evidence shows that t-tubules are remodelled in the ventricles of the failing heart, it still remains unclear how t-tubule structure and function changes during cardiac disease. Amphiphysin II (AMPII), a member of the Bin/Amphiphysin/Rvs (BAR) domain family, is known to be involved in membrane invaginations and t-tubule biogenesis (Muller et al, 2003). The aims of this study are to investigate 1) if AMPII is reduced in a tachypacing induced sheep model of heart failure; 2) the role of AMPII in the formation and maintenance of t-tubules in rat ventricular myocytes. Under isoflurane anaesthesia (2-4% v/v in oxygen) sheep were instrumented with a pacemaker and pacing lead. Post-operative analgesia (meloxicam 0.5 mg/kg) and antibiosis (enrofloxacin 2.5 mg/kg) were provided for 24 hours. Heart failure was induced by rapid ventricular pacing (Briston et al. 2011) for 4-5 weeks. Following pentobarbitone euthanasia (200 mg/kg iv) samples of left ventricular and left atrial myocardium were snap frozen. Age-matched non-instrumented animals served as controls. T-tubule density was assessed using the potential sensitive dye di-4-ANEPS and confocal microscopy (Dibb et al. 2009). Data are presented as mean ±S.E.M and statistical significance determined using a students t-test and considered significant when p<0.05. Western blotting performed on protein samples obtained from the left ventricular wall and left auricle showed a significant reduction in AMPII protein in heart failure (ventricle, decreased by 38 ±12.1%, atria, decreased by 43 ±12.1%, n=7, p<0.05). Transfecting ventricular myocytes, enzymatically isolated from rat hearts, with small interfering RNA (siRNA) against AMPII causes t-tubule loss. The distance at which 50% of pixels in target cells are from a membrane (sarcolemma or t-tubule) compared with control cells is increased by 41 ±22.3 %, p<0.05. Western blotting performed on transfected cells shows that AMPII protein is significantly reduced in target cells when compared with control (decreased by 44 ±4.6 %, p<0.05). AMPII protein expression is significantly reduced in the sheep model of tachypacing induced heart failure in both the atria and the ventricle. There is also a loss of t-tubules in cells transfected with siRNA against AMPII, suggesting that AMPII plays a key role in the maintenance of t-tubules. This reduction is likely to cause dysynchrous Ca2+ release, leading to reduced contractility and heart failure.