Real-Time Imaging Reveals Augmentation of Glutamate-Induced Ca2+ Transients by the NO-cGMP Pathway in Cerebellar Granule Neurons

Dysfunctions of NO-cGMP signaling have been implicated in various neurological disorders. We have studied the potential crosstalk of cGMP and Ca2+ signaling in cerebellar granule neurons (CGNs) by simultaneous real-time imaging of these second messengers in living cells. The NO donor DEA/NO evoked c...

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Bibliographic Details
Published in:International journal of molecular sciences 2018-07, Vol.19 (8), p.2185
Main Authors: Paolillo, Michael, Peters, Stefanie, Schramm, Andrea, Schlossmann, Jens, Feil, Robert
Format: Article
Language:English
Subjects:
calcium
Calcium ions
Calcium signalling
cerebellar granule cells
Cerebellum
Crosstalk
Cyclic GMP
Data processing
FRET imaging
Granular materials
Granule cells
Guanylate cyclase
guanylyl cyclase
Imaging
Inhibitors
Kinases
Neurological diseases
Neurons
Nitric oxide
Phosphodiesterase
PKG
Protein kinase
Proteins
Real time
Rodents
Second messengers
Transgenic mice
Zaprinast
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Summary:Dysfunctions of NO-cGMP signaling have been implicated in various neurological disorders. We have studied the potential crosstalk of cGMP and Ca2+ signaling in cerebellar granule neurons (CGNs) by simultaneous real-time imaging of these second messengers in living cells. The NO donor DEA/NO evoked cGMP signals in the granule cell layer of acute cerebellar slices from transgenic mice expressing a cGMP sensor protein. cGMP and Ca2+ dynamics were visualized in individual CGNs in primary cultures prepared from 7-day-old cGMP sensor mice. DEA/NO increased the intracellular cGMP concentration and augmented glutamate-induced Ca2+ transients. These effects of DEA/NO were absent in CGNs isolated from knockout mice lacking NO-sensitive guanylyl cyclase. Furthermore, application of the cGMP analogues 8-Br-cGMP and 8-pCPT-cGMP, which activate cGMP effector proteins such as cyclic nucleotide-gated cation channels and cGMP-dependent protein kinases (cGKs), also potentiated glutamate-induced Ca2+ transients. Western blot analysis failed to detect cGK type I or II in our primary CGNs. The addition of phosphodiesterase (PDE) inhibitors during cGMP imaging showed that CGNs degrade cGMP mainly via Zaprinast-sensitive PDEs, most likely PDE5 and/or PDE10, but not via PDE1, 2, or 3. In sum, these data delineate a cGK-independent NO-cGMP signaling cascade that increases glutamate-induced Ca2+ signaling in CGNs. This cGMP–Ca2+ crosstalk likely affects neurotransmitter-stimulated functions of CGNs.
ISSN:1422-0067
1661-6596
1422-0067
DOI:10.3390/ijms19082185