The simultaneous measurement of epithelial ion transport and intracellularfree Ca2+ in cultured equine sweat gland secretory epithelium

We explored the relationship between nucleotide-evoked changes in intracell ular free calcium ([Ca2+](i)) and anion secretion by measuring [Ca2+](i) an d I-SC simultaneously in Fura-2-loaded, cultured equine sweat gland epithel ia. Apical ATP, UTP or UDP elicited sustained increases in [Ca2+](i) that w ere initiated by the mobilization of cytoplasmic Ca2+ but maintained by Ca2 + influx. However, although these nucleotides also increased I-SC, this res ponse was transient whereas the [Ca2+](i) signals were sustained. Experimen ts in which external Ca2+ was removed/replaced showed that Ca2+. entering n ucleotide-stimulated cells elicited very little change in I-SC. Cross desen sitization experiments showed that UTP-stimulated epithelia became insensit ive to ATP but that UTP could increase both [Ca2+](i) and I-SC in ATP-stimu lated cells by activating 'pyrimidinoceptors' essentially insensitive to AT P. Thapsigargin evoked a sustained rise in [Ca2+](i) that was accompanied b y a maintained increase in I-SC. However, this increase in I-SC was depende nt upon external Ca2+ and so the responses to nucleotides and thapsigargin have different properties. ATP increased I-SC in thapsigargin-treated cells without causing any rise in [Ca2+](i) while ionomycin increased both param eters. The data therefore show that apical P2Y receptors allow nucleotides to increase I-SC via two mechanisms, one of which appears to be [Ca2+](i)-i ndependent control of anion channels.