Quantitation of ryanodine receptor of rabbit skeletal muscle, heart and brain |
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Authors: | E Damiani G Tobaldin P Volpe A Margreth |
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Institution: | Centro di Studio per la Biologia e la Fisiopatologia muscolare, Università di Padova, Italy. |
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Abstract: | The total number of high-affinity ryanodine receptor (RyR) binding sites present in skeletal and cardiac muscle and in brain tissue of the rabbit was determined by 3H]ryanodine binding to subfractions obtained by differential centrifugation of homogenates prepared in a low-ionic strength medium, containing 0.5% Chaps. In all three tissues at least 80% of 3H]ryanodine binding was recovered in the total membrane (TM) fraction obtained by centrifuging between 650 g for 10 min and 120,000 x g for 90 min. Skeletal muscle displayed higher contents of high-affinity RyR sites (about 49 pmol/g wet wt) than heart and brain (about 12 pmol and 3.5 pmol/g wet wt, respectively). The affinity for ryanodine, as well as the affinity for Ca2+, in the absence or presence of Ca2(+)-releasing drugs (caffeine and doxorubicin) of TM from skeletal muscle, were found to be identical to those of purified terminal cisternae. As low as 1 g of tissue was sufficient to perform several experiments. |
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