Blank PS et al. (1998), The calcium sensitivity of individual secretory...

ECB-ART-37033

J Gen Physiol 1998 Nov 01;1125:569-76. doi: 10.1085/jgp.112.5.569.

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The calcium sensitivity of individual secretory vesicles is invariant with the rate of calcium delivery.

Blank PS , Vogel SS , Cho MS , Kaplan D , Bhuva D , Malley J , Zimmerberg J .

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Differences in the calcium sensitivity of individual secretory vesicles can explain a defining feature of calcium-regulated exocytosis, a graded response to calcium. The role of the time dependence of calcium delivery in defining the observed differences in the calcium sensitivity of sea urchin egg secretory vesicles in vitro was examined. The calcium sensitivity of individual secretory vesicles (i.e., the distribution of calcium thresholds) is invariant over a range of calcium delivery rates from faster than micromolar per millisecond to slower than micromolar per second. Any specific calcium concentration above threshold triggers subpopulations of vesicles to fuse, and the size of these subpopulations is independent of the time course required to reach that calcium concentration. All evidence supports the hypothesis that the magnitude of the free calcium is the single controlling variable that determines the fraction of vesicles that fuse, and that this fraction is established before the application of calcium. Submaximal responses to calcium cannot be attributed to alterations in the calcium sensitivity of individual secretory vesicles arising from the temporal properties of the calcium delivery. Models that attempt to explain the cessation of fusion using changes in the distribution of calcium thresholds arising from the rate of calcium delivery and/or adaptation are not applicable to this system, and thus cannot be general.

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Genes referenced: LOC100887844 LOC100893907

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	Figure 2. Interrupted ramp. Interrupting a calcium ramp (arrow) and maintaining this calcium concentration (solid line) results in submaximal extents of fusion that are indistinguishable from those produced by step changes to the same calcium concentration. See Fig. 8 A.
	Figure 8. (A) Overlap between the extent of fusion after an interrupted ramp (â–ª) and the underlying calcium activity curve derived from step changes in the calcium concentration (solid line). Overlap between the extent of fusion after short pulses of UV (â€¢) and the underlying calcium activity curve derived from step changes in the calcium concentration shifted by 0.2 pCa units (dotted line). This shift in the activity curve is consistent with the known errors in determining the free calcium concentration using Rhod-2 fluorescence and is not significant. (B) The change in calcium (Î”pCa) required to detect a 10% change in fusion beginning at any initial value of fusion is shown (solid line). The curve is derived from the log-normal cumulative distribution function that describes the calcium activation curve (d% fusion/d pCa) and the points represent the data from individual photolysis experiments.
	Figure 7. Correlation between calcium challenges. The correlation between the calcium in the first challenge and the calcium at the onset of fusion after the second challenge is shown. All points, except one, lie to the left of the identity line, indicating that a higher calcium concentration is required for subsequent fusion.
	Figure 3. Continuation of an interrupted ramp. (A) The dynamic response of the remaining unfused vesicles is not altered when the ramp is continued. (B) The slope of the cross-correlation between the last 30% fusion response for interrupted (solid line) and continuous (dotted line) ramps using normalized time as the parametric variable is not significantly different from one. The identity line is included for comparison.
	Figure 4. Response to a calcium ramp generated by perfusion. (A) The dynamic response to calcium ramps of identical magnitude but different rates of delivery. (B) The difference in pCa at 50% fusion in the dynamic response to the two ramps is not significant. The slope of the cross-correlation between the fusion response at the two different calcium-delivery rates using normalized time as the parametric variable is consistent with no significant differences between the dynamic response for 11- and 23-min ramps.

	Figure 6. Double challenge with photolysis of DM-nitrophen. (A) The release of calcium after photolysis of DM-nitrophen leads to submaximal extents of fusion. The time-dependent changes in the calcium concentration during the two challenges are indicated. The calcium concentration during the second challenge increases to concentrations >60 Î¼M (data not shown). Calcium concentration is monitored using Rhod-2 fluorescence. (B) The calcium at the onset of fusion after the second challenge is indicated by point 3. The horizontal lines define the peak-to-peak noise envelope and the first detectable fusion event is indicated. Point 1 marks the beginning of UV irradiation, point 2 marks the time at which the calcium concentration matches the peak transient (first challenge), and point 3 marks the onset of fusion. The data were collected using a sample time of 0.02 s with an offset of 0.01 s between light scattering and fluorescence data.

References [+] :

Blank, Submaximal responses in calcium-triggered exocytosis are explained by differences in the calcium sensitivity of individual secretory vesicles. 1998, Pubmed, Echinobase