Figure 7. Caffeine-induced Ca²⁺ release from the isolated SR vesicles and the effect of DP4.

Ca²⁺ release from the SR vesicles was fluorimetrically determined using fura 2 in a solution containing 0.17 M KCl, 20 mM Mopso, pH 6.8, and 1 mM MgCl₂. The vesicles were actively loaded with Ca²⁺ by addition of 100 μM MgATP and 20 μM CaCl₂, and then Ca²⁺ release was triggered by caffeine. (A) Representative traces of Ca²⁺ release induced by 5 mM caffeine (arrowhead) with (+30 μM DP4) or without added peptide (Control). DP4 was added to the solution just before the start of Ca²⁺ loading. Left, normal SR; right, MHS SR. (B) Caffeine-concentration-dependence of Ca²⁺ release in the presence (●) and absence (○) of 30 μM DP4. Left, normal SR; right, MHS SR. The change in Ca²⁺ concentration was normalized at the maximal values at each condition; 100% denotes 240, 180, 240 and 150 nM for Normal-Control, Normal-DP4, MHS-Control and MHS-DP4 respectively. Results are means±S.E.M. (n=3–6). Note that DP4 sensitized the normal SR to caffeine in releasing Ca²⁺, whereas it produced no sensitizing effect on the MHS SR. DP4-mut (30 μM) produced no effects on either SR (▲).