Inositol 1,4,5-Trisphosphate-induced Increase in Ca2+-ATPase Activity in the Microsomes of Tracheal Epithelial Cells

• 영문명: Inositol 1,4,5-Trisphosphate-induced Increase in Ca²⁺-ATPase Activity in the Microsomes of Tracheal Epithelial Cells
• 발행기관: 대한생리학회
• 저자명: Cho, Hyoung-Jin Park, Sung-Shin Kim, Young-Kee
• 간행물 정보: 『대한생리학회지』제29권 제2호, 269~278쪽, 전체 10쪽
• 주제분류: 의약학 > 의학일반
• 파일형태: PDF
• 발행일자: 1995.12.31

영문 초록

Membrane vesicles were prepared by differential centrifugation from epithelial cells of porcine trachea. Total activity of microsomal ATPases was measured spectrophotometrically by a coupled enzyme assay. The steady-state activity of the enzyme was 329±10 nmol/min mg protein. Thapsigargin, a specific antagonist of intracellular Ca²⁺-ATPase, inhibited about 50% of the activity, leaving 178±18 nmol/min .mg protein (n=6), indicating that the Ca²⁺-ATPase is one of the major microsomal ATPases. The microsomes used in this study appeared to be tight-sealed vesicles since they showed saturation in ⁴⁵Ca²⁺ uptake experiments. Inositol 1,4,5-trisphosphate InsP₃, 4 μM, an agonist of InsP_{3}-sensitive Ca²⁺ release channel (InsP₃, receptor), and Ca-ionophore A23187 (10 μM) induced ⁴⁵Ca²⁺ releases of 20% and 50% of stored ⁴⁵Ca²⁺, respectively. The addition of (10 μM InsP₃ also increased the microsomal ATPase activity from 282±8 nmol/min mg protein to 334±21 nmol/min . mg protein in the intact vesicles. Similar increase in the activity was observed by making microsomes leaky (uncoupling) using the Ca-ionophore A23187. ;InsP₃-induced effects were blocked by either thapsigargin or heparin suggesting that: 1) the InsP₃-induced increase in ATPase activity is mediated by microsomal Ca²⁺-ATPase, and 2) dissipation of Ca²⁺ gradient across the microsomal membrane is responsible for the InsP₃-induced effect. In order to test the dependence of the Ca²⁺-ATPase activity on the activity of InsP₃-induced the activity of ATPases was monitored in various concentrations of free Ca²⁺ using EGTA-Ca²⁺ buffers. The Ca²⁺-dependent biphasic change is the well-known character of InsP₃ receptor but not of microsomal Ca²⁺-ATPase in non-excitable cells; however, the activity of microsomal ATPase appeared biphasic and a maxim진 activity of 397±36nmol/min .mg protein was obtained in the solution containing 100 nM free Ca²⁺. Below or above this concentration, the activity of ATPases was lower. These results strongly support a positive correlation of microsomal Ca²⁺-ATPase to the InsP₃ receptors in epithelial microsomes.

키워드

Porcine tracheal epithelium Ca²⁺-ATPase InsP₃ receptor Coupling ⁴⁵Ca²⁺ uptake and release