Research Report 2010 - MDC

Figure 3. Membrane resealing assay performed on wild-type mousesingle skeletal muscle fibres. Membrane damage (5x5µm) wasinduced with a two-photon confocal laser-scanning microscope (LSM510 META, Zeiss) coupled to a 488-nm Argon Ti:Sapphire Laser in thepresence of FM 1-43FX fluorescence dye. Top: Plot of fluorescenceintensity (n=10) against time in the presence (red line) and absence(black line) of Ca 2+ . Data are means +-SEM. Bottom: fluorescenceobtained in the presence (right) and absence (left) of Ca 2+ .Ahnak1 – a novel, prominent modulator of cardiacL-type Ca 2+ channelsHannelore Haase, Ines Pankonien, Karin Karczewski,Steffen Lutter (In collaboration with Nathan Dascal,University Tel Aviv, Israel)Ahnak1 is located at the sarcolemma and T-tubuli ofcardiomyocytes indirectly associated with the voltagedependentL-type Ca 2+ channel (L-VDCC) via its β2-subunit.The goal of this study will be understanding theinteractions of ahnak1 with L-VDCC, i.e. α1C and β2, andtheir modulation by β-adrenergic stimulation, mutations,and PKC activation (Figure 2). This will be achievedby heterologous expression of ahnak1 fragments inXenopus oocytes and HEK cells and the characterizationof their effects on cardiac and smooth muscle L-VDCC. Furthermore, reconstitution of β-adrenergicmodulation of L-VDCC in heterologous expression systems,and elucidation of the role of ahnak1, Cavβ andother signaling components in PKA and PKC modulationwill be studied.The functional role of the ahnak protein familyin adult skeletal muscle fibersAndreas Marg, Petra DomaingThe aim of the project is the elucidation of the role ofahnak protein family in skeletal muscle fibers. Thetransmembrane protein dysferlin seems to anchorahnak1 and ahnak2 to the sarcolemma, thus providing amembrane-stabilizing dysferlin-ahnak-actin complex.We investigate whether the ahnak protein family isimportant for membrane stability and Ca 2+ handling ofskeletal muscle fibers. A laser-assisted membraneresealing (Figure 3) and a Fura2-based fluorescenceassay of electrically stimulated enzymatically isolatedsingle skeletal muscle fibers from mouse Flexor digitorumbrevis will be applied.Identification of new adipocyte-derivedcardiodepressant factorsChristiane Look, Lena Martin (In collaboration withValeria Lamounier-Zepter, University Dresden)Recently, we showed that cultivated human adipocytesecrete factors which depress contraction of cardiomyocytes(Figure 4). The aim of this project is to identifyadipocyte-derived cardiodepressant factors. We willassess the role of different adipose tissue depots in producingthese cardiodepressant factors in our in vitroadipocyte/cardiomyocyte and adipocyte/isolated perfusedheart models. Finally, the adipocyte-derived cardiodepressantfactors will be analysed in vivo in microdialysissamples of subcutaneous adipose tissue ofobese patients as well as in serum of our obese patientcohort. The resulting data will allow the developmentof risk profiling and novel therapeutic strategies forheart dysfunction in obese patients.14 Cardiovascular and Metabolic Disease Research