'Trigger' Events Precede Calcium Puffs in Xenopus Oocytes

¹ University of California, Irvine
² Bristol University

^* To whom correspondence should be addressed. E-mail: heather{at}uci.edu.

Submitted on May 11, 2006
Revised on June 26, 2006
Accepted on 30 August 2006

	Abstract

The liberation of calcium ions sequestered in the endoplasmic reticulum through inositol 1,4,5-trisphosphate receptors/channels (IP₃Rs) results in a spatio-temporal hierarchy of calcium signaling events that range from single channel openings, through local Ca²⁺ puffs believed to arise from several to tens of clustered IP₃Rs, to global calcium waves. Using high-resolution confocal linescan imaging and a sensitive Ca²⁺ indicator dye (Fluo-4-dextran) we show that puffs are often preceded by small, transient Ca²⁺ elevations that we christen trigger events. The magnitude of triggers is consistent with their arising from the opening of a single IP₃ receptor/channel, and we propose that they initiate puffs by recruiting neighboring IP₃Rs within the cluster by a regenerative process of Ca²⁺-induced Ca²⁺ release. Puff amplitudes (fluorescence ratio change) are on average about 6-times greater than that of the triggers, suggesting that at least 6 IP₃Rs may simultaneously be open during a puff. Trigger events have average durations of about 12 ms, as compared to 19 ms for the mean rise-time of puffs, and their spatial extent is about 3-times smaller than puffs (respective widths at half peak amplitude 0.6 and 1.6 µm). All these parameters were relatively independent of IP₃ concentration, although the proportion of puffs showing resolved triggers was greatest (~ 80%) at low [IP₃]. Because Ca²⁺ puffs constitute the building blocks from which cellular IP₃-mediated Ca²⁺ signals are constructed, the events that initiate them are likely to be of fundamental importance for cell signaling. Moreover, the trigger events provide a useful yardstick by which to derive information regarding the number and spatial arrangement of IP₃Rs within clusters.

Key Words: IP3, blips, calcium channels, confocal

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