We have developed a long-term culture of adult rat ventricular myocytes and investigated the cellular and subcellular Ca2+ signalling along with the effects of chronic neurohormonal stimulation. First, we assessed the loading state of internal Ca2+ stores and the functional activity of pumping/extrusion systems in Fura-2 loaded myocytes by caffeine (10 mM) applications and electrical pacing. Spatially resolved Ca2+ imaging (realtime confocal microscopy) was undertaken to characterise the spatio- temporal features of spontaneous Ca2+ sparks and waves. At days3 and 6, we observed decreased amplitudes of caffeine-induced Ca2+ transients, whereas the frequency of spontaneous waves is augmented from day 6 onwards. We measured a 2-fold increase in the size of the spark spread from day 1 onwards. The spark distribution changed from a homogeneous cytoplasmic spread into a sub-plasma membrane-localised ring at day 6. Secondly, chronic stimulation with phenylephrine (10-100 mM) or endothelin-1 (100 nM) led to a rapid (within 3 days) dedifferentiation of the myocytes into so-called myoballs, which formed spreading complexes from day 6 onwards. From this we conclude that neurohumoral stimulation of adult myocytes is an important part of cardiac remodelling in-vivo that can be mimicked in an in-vitro model of adult myocytes. Fundings: HOMFOR and the DFG: SFB 530, and Graduiertenkolleg

"Cellular Regulation and Growth".