Activating and inhibitory K+-dependent effect of prilocaine on the Ca-ATPase

Objectives: Describe the effect of prilocane (P) on SERCA activity and the modulation of its effect by K+ in the presence and absence of the calcium ionophore calcimycin (C). Methods: Sarcoplasmic reticulum (SR) membranes from rabbit fast-twitch skeletal muscle with calcium accumulation ability were obtained as sealed vesicles by differential centrifugation as described by Champeil et al. (1985). The ATPase activity was measured by the colorimetric method of Baginski et al. (1967). The P half-maximal inhibitory concentration (Ki) was determined (mean±SD, n=6) and evaluated by ANOVA (p<0.05). Results: In the presence and absence of C, P inhibited ATPase activity in the studied range (3-90 mM). With C the activity values were higher than in its absence, even at high [P] p<0.001. Ki values (mM P) increased with C (21.72 ± 0.67; 27.12 ± 1.24), and without C (17.52 ± 0.31; 23.55 ± 0.87) in the range 20-200mM K+, showing a statistically significant difference. In media without P SERCA activity increased linearly as a function of [K+], and as [P] increased, this relationship tends to a constant value. The preexposure of SERCA to P up to 20 min increased the ATPase activity. In addition, preincubation with K+ (50, 100 and 200 mM), decreases such effect. Conclusions: It is concluded that P at concentrations available in pharmaceutical formulations for clinical medical and dental uses inhibit SERCA. The inhibitory effect of P on SERCA activity is modulated by K+ and C. Some authors suggest that at 20 mM [K+] the maximum velocity for the dephosphorylation stage E2P to E2 is reached in the absence of anesthetic. This would probably explain why in the experiments carried out no large differences were found between the activity values for the different [K+]. The increase in activity observed by preexposing the SR membranes to P could be attributed to an increase in the permeability of the SR membranes to Ca2+, avoiding retroinhibition due to Ca2+ accumulation. The results suggest that such decrease in ATPase activity could be consequence of the alteration of the aminoacidic sequence at the phosphorylation and nucleotide site caused by P (SAIO 2015).