Intestinal Maturation: Characterization of Mitochondrial Calcium Transport in the Rat

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Abstract
The mitochondria play a major role in the regulation of intracellular calcium. Despite the fact that the enterocytes receive the majority of absorbed calcium, the role of the intestinal mitochondria in calcium transport during maturation is not known. Therefore, the current studies were designed to characterize calcium pump activity of jejunal mitochondria of rats during maturation (suckling, weanling, and adolescent rats). The functional integrity of the intestinal mitochondria of suckling and adolescent rats was determined by oxygen consumption studies demonstrating respiratory control ratios of more than 3 when succinate was used as a test substrate. Ca++ uptake was significantly stimulated by the presence of 3 mM ATP at all age groups studied. Maximal Ca++ uptake in the presence of 3 mM ATP and 2 mM succinate was 31.1 ± 0.4, 50.2 ± 4.2, and 94.3 ± 1.5 nmol/mg protein (mean ± SE) in suckling, weanling, and adolescent rats, respectively. Rates of ATP hydrolysis were 15.5 ± 1.5 and 2.9 ± 0.3 nmol/ATP hydrolyzed/mg protein in adolescent and suckling rats, respectively (p++ uptake was completely inhibited by 0.25 μM ruthenium red, oligomycin (10 μg/mg protein), 0.5 mM dinitrophenol and 1 mM sodium azide at all age groups. Ca++ efflux in the presence of ruthenium red occurred by a Na+-dependent pathway, indicating a Ca++/Na+ exchange mechanism. Kinetic parameters for ATP stimulated Ca++ uptake at 10 s revealed a Km of 0.84 + 0.11, 0.65 ± 0.17, and 0.57 ± 0.03 M and Vmax of 1.83 ± 0.07, 3.62 + 0.26 and 14.15 ± 0.21 nmol/mg protein/10 s in suckling, weanling, and adolescent rats, respectively. These results indicate that the mitochondria of the suckling enterocyte possess a well-characterized calcium transport system; however, the affinity and capacity of the transport system increases with advancing age. The greater capacity seen with increasing age most likely reflects greater rates of ATP hydrolysis in adolescent rats compared to suckling rats, thus providing greater driving force for calcium uptake.