The role of NADPH- and reduced glutathione-dependent enzymes in the norepinephrine modulation of the ATP-dependent, hepatic microsomal calcium pump: A new pathway for the noradrenergic regulation of cytosolic calcium in the hepatocyte

The authors have recently found that the hepatic, microsomal ATP-dependent Ca⁺⁺ pump activity is decreased through oxidation by cytochrome P-450-generated reactive oxygen species. This inhibition is reversed by reduced glutathione and only partially reversed by reactive oxygen scavengers. In view of these observations, the authors have sought to determine whether norepinephrine could regulate the Ca⁺⁺ pump by differential modulation of the oxidative and reductive pathways. They find that, in the presence of superoxide dismutase (15 μg/ml), catalase (65 μg/ml), reduced glutathione (5 mM) and NADP⁺ (0.39 mM), the pump activity was maximal (142% of no norepinephrine) at 10^-11 to 10^-10 M norepinephrine and decreased with increasing concentrations of norepinephrine. NADPH had no effect on uptake at 10^-11 M norepinephrine, but, between 10^-10 and 10^-8 M norepinephrine, it significantly decreased uptake compared with NADP⁺. At 10^-7 to 10^-6 M norepinephrine, with either NADP⁺ or NADPH, the uptake was significantly lower than at other norepinephrine concentrations. This decrease in the uptake seen at 10^-7 to 10^-6 M norepinephrine disappeared on the addition of 0.25 μM l-(S)-propranolol. The NADPH inhibition of the pump was blocked by imidazole-histidine buffer but not by inhibitors of mitochondrial metabolism. ATP and norepinephrine had little effect on mitochondrial uptake. These studies suggest that norepinephrine may modulate the hepatic, microsomal ATP-dependent Ca⁺⁺ pump through alterations in the balance between oxidative and reductive pathways.