Inositol 1,4,5-Trisphosphate-dependent Oscillations of Luminal [Ca2+] in Permeabilized HSY Cells

Abstract

In many cell types, stimulation of receptors linked to phosphoinositide signaling pathways results in oscillations of intracellular calcium concentration ([Ca 2ϩ ] i ) due to the periodic release and reuptake of Ca 2ϩ by intracellular stores (1-4). These oscillations often take the form of "base-line spikes" in which periodic spikes of [Ca 2ϩ ] i rise from a base-line [Ca 2ϩ ] i which is close to resting levels. It has been suggested that [Ca 2ϩ ] i oscillations provide a sort of digital encoding of the receptor-mediated signal that may have certain advantages, such as noise reduction and decreased exposure to high intra- ] i such that small increases in [Ca 2ϩ ] i above resting levels would be expected to activate the channel, but large increases would be inhibitory. Several detailed mathematical analyses (9 -11) have demonstrated that a single intracellular Ca 2ϩ pool possessing IP 3 receptors with properties derived from these (and other) empirical measurements can produce [Ca 2ϩ ] i oscillations that closely resemble experimental observations. Thus these calculations substantiate the interesting possibility that [Ca 2ϩ ] i oscillations may occur solely as a result of the intrinsic properties of the IP 3 -sensitive Ca 2ϩ channel. Briefly, the underlying operation of these models is the following. A small initial rise in [Ca 2ϩ ] i due to IP 3 -dependent Ca 2ϩ release results in a rapid Ca 2ϩ -dependent increase in IP 3 -sensitive channel activity. This feed forward effect of Ca 2ϩ on its own release generates the upstroke of a [Ca 2ϩ ] i spike. At this high [Ca 2ϩ ] i a somewhat slower process of Ca 2ϩ feedback inhibition then results in channel closure. Ca 2ϩ reuptake into intracellular stores via the SERCA pump subsequently returns [Ca 2ϩ ] i to near base-line levels where Ca 2ϩ feedback inhibition is slowly relieved, followed by reactivation of IP 3 -induced Ca 2ϩ release and a repetition of the [Ca 2ϩ ] i oscillation cycle. To date experimental studies of [Ca 2ϩ ] i oscillations have utilized intact cells, making it difficult to definitively confirm or exclude the involvement of many intracellular processes in this phenomenon, and consequently complicating tests of proposed mechanisms. In the work presented here we extend these studies to permeabilized cells. For our experiments we have employed the HSY cells, a human salivary epithelial cell line (12) that has been found to be a useful model system for investigations of intracellular Ca 2ϩ signaling events (13-15). We have previously shown (15) that the luminal calcium concentration ([Ca 2ϩ ] L ) within the IP 3 -sensitive intracellular Ca 2ϩ stores of saponin-permeabilized HSY cells can be monitored with the fluorescent Ca 2ϩ indicator Mag-fura-2. Using this permeabi-* The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

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Multiple isoforms of the ryanodine receptor are expressed in rat pancreatic acinar cells

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