Autoregulation in PC12 cells via P2Y receptors: Evidence for non-exocytotic nucleotide release from neuroendocrine cells |
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Authors: | Simon Hussl Helmut Kubista Stefan Boehm |
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Institution: | (1) Institute of Pharmacology, Medical University of Vienna, Waehringerstrasse 13a, 1090 Vienna, Austria;(2) Institute of Experimental and Clinical Pharmacology, Medical University of Graz, Universitaetsplatz 4, 8010 Graz, Austria |
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Abstract: | Nucleotides are released not only from neurons, but also from various other types of cells including fibroblasts, epithelial,
endothelial and glial cells. While ATP release from non-neural cells is frequently Ca2+ independent and mostly non-vesicular, neuronal ATP release is generally believed to occur via exocytosis. To evaluate whether
nucleotide release from neuroendocrine cells might involve a non-vesicular component, the autocrine/paracrine activation of
P2Y12 receptors was used as a biosensor for nucleotide release from PC12 cells. Expression of a plasmid coding for the botulinum
toxin C1 light chain led to a decrease in syntaxin 1 detected in immunoblots of PC12 membranes. In parallel, spontaneous as
well as depolarization-evoked release of previously incorporated 3H]noradrenaline from transfected cells was significantly reduced in comparison with the release from untransfected cells,
thus indicating that exocytosis was impaired. In PC12 cells expressing the botulinum toxin C1 light chain, ADP reduced cyclic
AMP synthesis to the same extent as in non-transfected cells. Likewise, the enhancement of cyclic AMP synthesis either due
to the blockade of P2Y12 receptors or due to the degradation of extracellular neucleotides by apyrase was not different between non-transfected and
botulinum toxin C1 light chain expressing cells. However, the inhibition of cyclic AMP synthesis caused by depolarization-evoked
release of endogenous nucleotides was either abolished or greatly reduced in cells expressing the botulinum toxin C1 light
chain. Together, these results show that spontaneous nucleotide release from neuroendocrine cells may occur independently
of vesicle exocytosis, whereas depolarization-evoked nucleotide release relies predominantly on exocytotic mechanisms. |
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Keywords: | Cyclic AMP Nucleotide release PC12 cells P2Y receptor Vesicle exocytosis |
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