The effect of L-type Ca2+ channel blockers on anoxia-induced increases in intracellular Ca2+ concentration in rabbit proximal tubule cells in primary culture.
Ca2+ channel blockers (CCB) have been shown to be protective against ischaemic damage of the kidney, suggesting an important role for intracellular Ca2+ ([Ca2+]i) in generating cell damage. To delineate the mechanism behind this protective effect, we studied [Ca2+]i in cultured proximal tubule (PT) cells during anoxia in the absence of glycolysis and the effect of methoxyverapamil (D 600) and felodipine on [Ca2+]i during anoxia. A method was developed whereby [Ca2+]i in cultured PT cells could be measured continuously with a fura-2 imaging technique during anoxic periods up to 60 min. Complete absence of O2 was realised by inclusion of a mixture of oxygenases in an anoxic chamber. [Ca2+]i in PT cells started to rise after 10 min of anoxia and reached maximal levels at 30 min, which remained stable up to 60 min. The onset of this increase and the maximal levels reached varied markedly among individual cells. The mean values for normoxic and anoxic [Ca2+]i were 118 +/- 2 (n = 98) and 662 +/- 22 (n = 160) nM, respectively. D 600 (1 microM), but not felodipine (10 microM), significantly reduced basal [Ca2+]i in normoxic incubations. During anoxia 1 microM and 100 microM D 600 significantly decreased anoxic [Ca2+]i levels by 22 and 63% respectively. Felodipine at 10 microM was as effective as 1 microM D 600. Removal of extracellular Ca2+ and addition of 0.1 mM La3+ completely abolished anoxia-induced increases in [Ca2+]i. We conclude that anoxia induces increases in [Ca2+]i in rabbit PT cells in primary culture, which results from Ca2+ influx. Since this Ca2+ influx is partially inhibited by low doses of CCBs, L-type Ca2+ channels may be involved.
Department of Physiology, University of Nijmegen, The Netherlands.
U M
UM
Rose
R J
RJ
Bindels
A
A
Vis
J W
JW
Jansen
C H
CH
Van Os
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D000242:Cyclic AMP / Q000096:biosynthesis; D005609:Free Radicals; D016257:Fura-2; D006728:Hormones / Q000494:pharmacology; D000860:Hypoxia / Q000378:metabolism*; D007124:Immunoenzyme Techniques; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008856:Microscopy, Fluorescence; D011817:Rabbits
D002121:Calcium Channel Blockers; D005609:Free Radicals; D006728:Hormones; D000804:Angiotensin II; D000242:Cyclic AMP; D007770:L-Lactate Dehydrogenase; D001920:Bradykinin; D002118:Calcium; D016257:Fura-2