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J. Biol. Chem., Vol. 278, Issue 45, 44921-44928, November 7, 2003

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Compound Exocytosis and Cumulative Fusion in Eosinophils^*

Ismail Hafez, Andreas Stolpe, and Manfred Lindau

From the School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853

Focal release of cytotoxic proteins by eosinophils onto the target surface plays an important role in parasite killing. Degranulation was stimulated by intracellular application of calcium and guanosine 5'-3-O-(thio)triphosphate via the recording patch pipette or via streptolysin-O permeabilization. Exocytotic fusion was monitored by capacitance measurements, whereas release of fluorescent weak bases, which accumulate selectively within eosinophil granules, was followed by fluorescence imaging. Several distinct types of granule fusion events were directly observed by simultaneous capacitance and fluorescence measurements. These are fusion of a single granule with the plasma membrane, intracellular granule-granule fusion, fusion of large compounds of pre-fused granules with the plasma membrane (compound exocytosis), and sequential fusion of granules to granules previously fused to the plasma membrane. Extensive granule-granule fusion was also observed by electron microscopy of permeabilized cells. All these fusion mechanisms contribute to focal release. The coexistence of distinct modes of exocytosis suggests that their regulation may modulate effector functions of eosinophils during helminth infection and allergic response.

Received for publication, June 9, 2003 , and in revised form, August 14, 2003.

^* This work was supported by National Institutes of Health Grant RO1NS38200 and the Nanobiotechnology Center (National Science Foundation Science and Technology Center supported by Agreement ECS-9876771). 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.

To whom correspondence should be addressed: School of Applied and Engineering Physics, Cornell University, Ithaca, NY 14853. Tel.: 607-255-5264; Fax: 607-2557658; E-mail ml95{at}cornell.edu.

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