Abstract
Extracellular nucleotides such as ATP have been shown to regulate ion transport processes in a variety of epithelia. This effect is mediated by the activation of plasma membrane P2Y receptors, which leads to Ca(2+) signaling cascade. Ion transport processes (e.g. activation of apical calcium-dependent Cl(-) channels) are then stimulated via an increase in [Ca(2+)](i). Many polarized epithelia express apical and/or basolateral P2Y receptors. To test whether apical and basolateral stimulation of P2Y receptors elicit polarized Ca(2+) signaling and anion secretion, we simultaneously measured the two parameters in polarized epithelia. Although activation of P2Y receptors located at both apical and basolateral membranes evoked an increase in [Ca(2+)](i), only apical P2Y receptors-coupled Ca(2+) release stimulated an increase in anion secretion. Moreover, the calcium influx evoked by apical and basolateral P2Y receptor stimulation is predominately via the basolateral membrane domain. It appears that the apical P2Y receptor-regulated Ca(2+) release and activation of apical Cl(-) channels is compartmentalized in polarized epithelia with basolateral P2Y-stimulated Ca(2+) release failing to activate anion secretion. These data suggest that there may be two distinct ATP-releasable Ca(2+) pools, each coupled to apical and basolateral membrane receptor but linked to the same calcium influx pathway located at the basolateral membrane.
Highlights
There is, a paucity of information on the mechanisms of stimulus-secretion coupling in polarized epithelia that expressed both apical and basolateral P2Y receptors
The calcium influx evoked by apical and basolateral P2Y receptor stimulation is predominately via the basolateral membrane domain. It appears that the apical P2Y receptor-regulated Ca2؉ release and activation of apical Cl؊ channels is compartmentalized in polarized epithelia with basolateral P2Y-stimulated Ca2؉ release failing to activate anion secretion
Reversing the sequence of Ca2ϩ, the readdition showed similar results (n ϭ 8, data not shown). These results indicate that in polarized epithelia, the stimulation with apical ATP leads to the activation of transmembrane Ca2ϩ influx pathway, which is located mainly in the basolateral membrane
Summary
There is, a paucity of information on the mechanisms of stimulus-secretion coupling in polarized epithelia that expressed both apical and basolateral P2Y receptors. The Ca2ϩmediated ClϪ secretion activated by various nucleotides was explored in an equine sweat gland epithelial cell line with emphasis on the polarized Ca2ϩ mobilization and influx in response to apical versus basolateral P2Y receptor stimulation. A change in [Ca2ϩ]i (Fig. 3E) and ISC (Fig. 3F) was quantified in control monolayers and in monolayers pretreated with apical or basolateral ATP before the activation of the P2Y receptors localized in the contralateral membrane.
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