ISSN:
1573-4919
Keywords:
calcium uptake
;
chromaffin granules
;
annexins
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Chemistry and Pharmacology
,
Medicine
Notes:
Abstract Bovine chromaffin secretory granules were purified by isopycnic Metrizamide gradient centrifugation and their Ca2+ sequestration pathways were characterized. The rate of Ca2+ sequestration at 37°C was first order, with a maximal uptake of 26.9 ±0.46 (mean ± S.D., n = 3) nmol Ca2+/mg protein and a first order rate constant (k) of 0.046 ± 0.002 min−1. At 4°C the rate of uptake was substantially attenuated, with only 2.47 ± 0.2 (mean ± S.D, n = 3) nmol Ca2+/mg protein sequestered in 60 min. Ca2+ sequestration was 93% inhibited by 180 mM NaCl [I50% of 78.7 ± 9.3 mM NaCl (mean ± S.D., n = 11)] but only slightly inhibited by KCl or MgCl2. Ca 2+ sequestration was not stimulated by incubation with MgATP but was inhibited by 57% after incubation with 30 μM monensin. Ca 2+ sequestration was dependent on extravesicular Ca 2+ with half-maximal sequestration at pCa2+ 6.81 ± 0.028 (mean ± S.D., n = 3). Sequestered Ca2+ could be exchanged with external 45Ca2+, the exchange rate was first order (k of 0.042 ± 0.004: mean ± S.D., n = 3) and saturated at 27.7 ± 1.1 nmol Ca2+/mg (mean ± S.D., n = 3). The Ca2+/Ca2+ exchange system was totally inhibited by NaCl or KCl but only slightly by MgCl2. About 75% of sequestered 45Ca2+ could be released by incubation with NaCl, but only 8% was released by incubation with KCI. Half-maximal release of sequestered 45Ca2+ required 69.3 ± 12.2 mM NaCl (mean ± S.D., n = 3). The Na+-induced release of sequestered 45Ca2+ was rapid, t0.5 of 2.80 ± 0.63 min (mean ± S.D., n = 3) and inhibited at 4°C. The concurrent incubation of chromaffin granules with 45Ca2+ and either annexin proteins V or VI resulted in attenuated uptake of 45Ca2+. These results suggest that Ca2+ uptake in adrenal chromaffin granules is regulated by Na+ and Ca2+ gradients and also possibly by annexins V and VI.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00229309
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