Reversible desensitization of inositol trisphosphate-induced calcium release provides a mechanism for repetitive calcium spikes

Abstract

Repetitive transient increases in cytosolic calcium concentration (calcium spikes or calcium oscillations) are a common mode of signal transduction in receptor-mediated cell activation. Repetitive calcium spikes are initiated by phospholipase C-mediated production of inositol 1,4,5- trisphosphate (InsP3) and are thought to be generated by a positive feedback mechanism in which calcium potentiates its own release, a negative feedback mechanism by which calcium release is terminated, and a slow recovery process that defines the time interval between calcium spikes. The molecular mechanisms that terminate each calcium spike and define the spike frequency are not yet known. Here we show, in intact rat basophilic leukemia cells, that calcium responses induced by InsP3 are diminished for a period of 30- 60 s following an InsP3-induced calcium spike. The sensitivity of calcium release for InsP3 was probed by UV laser-mediated photorelease of InsP3, and calcium responses were monitored by fluorescence calcium imaging. A maximal loss in sensitivity (desensitization) was observed for InsP3 increases that resulted in a near maximal calcium spike and was expressed as an 80100% reduction in the calcium response to an equal amount of InsP3, released 10 s after the first UV pulse. When the amount of released InsP3 in the second pulse was increased 2-3-fold, desensitization was overcome and a second calcium response of equal amplitude to the first was produced. A power dependence of 3.2 was measured between the amount of released InsP3 and the amplitude of the triggered calcium response, explaining how a small decrease in InsP3 sensitivity can lead to a nearly complete reduction in the calcium response. Desensitization was abolished by the addition of the calcium buffers BAPTA and EGTA and could be induced by microinjection of calcium, suggesting that it is a calcium-dependent process. Half-maximal desensitization was observed at a free calcium concentration of 290 nM and increased with a power of 3.7 with peak calcium concentration. These studies suggest that reversible desensitization of InsP3-induced calcium release serves as a 'saw-tooth' parameter that controls the termination of each spike and the frequency of calcium spikes.