Cryoultramicrotomy, electron probe microanalysis and STEM of myocardial tissue

Heart muscle preparations (papillary muscles and trabeculae) were frozen at 4.2 K on metal plate under vaccum after their length-tension relationships showed that no damage had occurred during dissecting and mounting the strips on the holder. Freeze substitution of some preparations directly after freezing demonstrated that no important cell damage due to ice crystals occurred in superficial cell layers during freezing. Ultrathin cryosections obtained at -130 degrees C were freeze dried and analyzed, in the STEM mode. The better the freezing procedure, the poorer was the contrast of the sections under electron microscopy. Preliminary approaches to increasing contrast after sublimation of tissue water show that a small increase in contrast is generally obtained at the cost of the peak/background ratio due to pronounced mass loss. The results of our analyses show that C1 content in heart muscle cells is high and distributed throughout the cytoplasm. Ca is detectable and quantitable in resting muscle in SR cisternae. The Ca amount in cytoplasm is low and just at the limit of detectability under our current analysis conditions. Preliminary experiments on papillary muscles subjected to caffeine contracture showed that calcium is not detectable in the cisternae as is the case in control experiments. Only a moderate amount of Ca is detectable in mitochondria, whereas the concentrations in cytoplasm, as in resting cardiac muscle, is too low to be quantitated.