FIGURE 4.

Functional ryanodine receptors are decreased and showed rapid Ca²⁺-induced inactivation in MetS hearts. Saturation binding curve with [³H]-ryanodine. (A) Aliquots of 50 μg of protein from microsomes (SR-enriched fraction) from control (N = 6) and MetS (N = 5) hearts were incubated with [³H]-ryanodine (concentration range of 0.625–20 nM) in incubation medium (containing in mM: 1000 KCl, 0.1 CaCl₂ 20 HEPES, pH 7.2) for 90 min at 37°C. (B) Specific Ca²⁺-dependent [³H]-ryanodine binding curves of SR-enriched fractions from control (open symbols; N = 9) and MetS (orange symbols; N = 6) hearts. Microsomes were incubated with 7 nM [³H]-ryanodine and the indicated [Ca²⁺]_i at 37°C for 90 min. Values have been normalized and fitted to a modified version of the equation B = B_max([Ca²⁺]^na/([Ca²⁺]^na+K_a^na))(1–[Ca²⁺]ⁿⁱ/([Ca²⁺]ⁿⁱ+K_iⁿⁱ))+C. Non-specific [³H]-ryanodine binding was determined in the presence of 20 μM ryanodine and subtracted from all reported values. Open symbols, control; orange symbols, MetS; dashed line indicates the fitting curve for control and dark solid line the fitting curve for MetS microsomes. *P < 0.05 with respect to corresponding control values.