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The Journal of Neuroscience, March 1, 2003, 23(5):1580

BRIEF COMMUNICATION
Calcium-Dependent Exocytosis of Atrial Natriuretic Peptide from Astrocytes

Mojca Kran^{3, *}, Matja Stenovec^{2, *}, Marko Kreft^{1, 2}, Tina Pangri², Sonja Grilc¹, Philip G. Haydon⁴, and Robert Zorec^{1, 2}

¹ Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Medical Faculty, University of Ljubljana, 1000 Ljubljana, Slovenia, ² Celica Biomedical Sciences Center, 1000 Ljubljana, Slovenia, ³ Institute of Pharmacology and Toxicology, Medical Faculty, University of Ljubljana, Slovenia, and ⁴ Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104

Astrocytes are non-neuronal cells in the CNS, which, like neurons, are capable of releasing neuroactive molecules. However, the mechanism of release is ill defined. In this study, we investigated the mechanism of release of atrial natriuretic peptide (ANP) from cultured cortical astrocytes by confocal microscopy. To study the discharge of this hormone, we transfected astrocytes with a construct to express pro-ANP fused with the emerald green fluorescent protein (ANP.emd). The transfection of cells with ANP.emd resulted in fluorescent puncta in the cytoplasm that represent secretory organelles. If ANP is released by exocytosis, in which the vesicle fuses with the plasma membrane, then the total intensity of the green fluorescing probe should decrease, whereas the vesicle membrane is incorporated into the plasma membrane. To monitor exocytosis, we labeled the membrane with the fluorescent styryldye FM 4-64, a reporter of cumulative exocytosis. The application of ionomycin to elevate cytoplasmic [Ca²⁺] increased the fluorescence intensity of FM 4-64, whereas that of ANP.emd decreased. These effects were not observed in the absence of extracellular Ca²⁺, suggesting that ANP is released by regulated Ca²⁺-dependent exocytosis from astrocytes.

Key words: astrocytes; glia; ANP; exocytosis; FM 4-64; fusion pore

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^* M.K. and M.S. contributed equally to this work.

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Copyright © 2003 Society for Neuroscience  0270-6474/03/2351580-04$05.00/0

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