The number and spatial distribution of IP₃ Receptors underlying calcium puffs in Xenopus Oocytes

¹ University of California, Irvine

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

Submitted on May 12, 2006
Revised on June 26, 2006
Accepted on 29 August 2006

	Abstract

Calcium puffs are local Ca²⁺ release events that arise from a cluster of inositol 1,4,5-trisphosphate receptor/channels (IP₃Rs) and serve as a basic 'building block' from which global Ca²⁺ waves are generated. Important questions remain as to the number of IP₃Rs that open during a puff, their spatial distribution within a cluster, and how much Ca²⁺ current flows through each channel. The recent discovery of 'trigger' events - small Ca²⁺ signals that immediately precede puffs and are interpreted to arise through opening of single IP₃R channels - now provides a useful yardstick by which to calibrate the Ca²⁺ flux underlying puffs. Here, we describe a deterministic numerical model to simulate puffs and trigger events. Based on confocal linescan imaging in Xenopus oocytes, we simulated Ca²⁺ release in two sequential stages; representing the trigger by the opening of a single IP₃R in the center of a cluster for 12 ms, followed by the concerted opening of some number of IP₃Rs for 19 ms representing the rising phase of the puff. The diffusion of Ca²⁺ and Ca²⁺-bound indicator dye were modeled in a 3-dimensional cytosolic volume in the presence of immobile and mobile Ca²⁺ buffers, and were used to predict the observed fluorescence signal after blurring by the microscope point-spread function. An optimal correspondence with experimental measurements of puff spatial width and ratio of puff/trigger amplitudes was obtained assuming that puffs arise from the synchronous opening of 25-35 IP₃Rs, each carrying a Ca²⁺ current about 0.4 pA, with the channels distributed through a 300-800 nm diameter cluster.

Key Words: Calcium, Channel, IP3, Model, Puffs, Xenopus oocyte

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