Fig. 1. Scatchard analysis, pharmacological profile and Ca²⁺-dependence of [³H]-ryanodine binding to Drosophila melanogaster microsomal membrane fractions. Experiments were performed with 100 µg of microsomal protein and the presence of 1-120 nM (A) or 5 nM [³H]-ryanodine (B,C) as described in Materials and Methods. Panel A illustrates a hyperbolic saturation curve as a function of increasing ryanodine concentrations. The inset shows a linear Scatchard plot. This is a representative experiment from a total of 5. Panel B depicts the effects of several activators and inhibitors of RyR in comparison with control conditions (column a). column b, AMP-PCP 2 mM; column c, 10 mM MgCl₂; column d, 5 µM Ruthenium Red; column e, 10 µM xanthine; column f, 2 µM dantrolene; column g, 10 nM free Ca²⁺; column h, the addition of 5 mM caffeine. Panel C represents the [³H]-ryanodine binding to Drosophila microsomal membranes as a function of increasing Ca²⁺ concentrations. The free concentrations of the cation (100 nM to 10 mM) was adjusted using EGTA and according to the Chelator program (Tatusova and Madden, 1999). The results in B and C are expressed as mean±s.e.m. of five independent experimental observations; where not shown, errors bars are smaller than symbols.