2019
DOI: 10.3389/fnsyn.2019.00012
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Imaging and Analysis of Presynaptic Calcium Influx in Cultured Neurons Using synGCaMP6f

Abstract: Presynaptic Ca 2+ influx through voltage-gated calcium channels (VGCCs) is a key step in synaptic transmission that links action potential (AP)-derived depolarization to vesicle release. However, investigation of presynaptic Ca 2+ influx by patch clamp recordings is difficult due to the small size of the majority of synaptic boutons along thin axons that hamper clamp control. Genetically encoded calcium indicators (GECIs) in combination with live cell imaging provi… Show more

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Cited by 26 publications
(26 citation statements)
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“…Therefore, we next tested the consequences of α 2 δ TKO/KD on presynaptic calcium signals. To this end neurons were transfected with the genetically encoded calcium indicator GCaMP6f, coupled to synaptophysin and expressed under the control of a synapsin promoter ( 32 ), together with soluble mCherry to outline neuronal and axonal morphology ( Fig. 2 A ).…”
Section: Resultsmentioning
confidence: 99%
See 1 more Smart Citation
“…Therefore, we next tested the consequences of α 2 δ TKO/KD on presynaptic calcium signals. To this end neurons were transfected with the genetically encoded calcium indicator GCaMP6f, coupled to synaptophysin and expressed under the control of a synapsin promoter ( 32 ), together with soluble mCherry to outline neuronal and axonal morphology ( Fig. 2 A ).…”
Section: Resultsmentioning
confidence: 99%
“…Presynaptic calcium transients were visualized using GCaMP6f coupled to synaptophysin driven by a synapsin promotor as previously described ( 8 , 32 ) ( SI Appendix ).…”
Section: Methodsmentioning
confidence: 99%
“…The reduction in Ca 2+ rise in response to stimulation train is unlikely due to any changes in Ca 2+ influx via voltage gated Ca 2+ channels (VGCCs), since the peak of SyGCaMP6f signals induced by single AP was indistinguishable between WT and KO neurons (Fig. 5E-F) (Brockhaus et al, 2019; Kim and Ryan, 2013).…”
Section: Resultsmentioning
confidence: 99%
“…In addition, expressing GCaMP in specific neurons in vivo revealed molecular mechanisms involved in the regulation of neurotransmitter release (Sgobio et al 2014), and the relationship between SV release and presynaptic calcium changes was clarified by simultaneous imaging of two different fluorescence probes for GCaMP and redshifted reporter of SVs (Jackson and Burrone 2016;Li et al 2011). GCaMP was developed to more accurately measure presynaptic calcium changes in living neurons (Brockhaus et al 2019). However, animals expressing GCaMP displayed aberrant current activity and suppression of SV probability (Singh et al 2018;Steinmetz et al 2017), suggesting that artifacts derived from GCaMP overexpression in vivo should be assessed.…”
Section: Genetically Encoded Calcium Indicator (Geci)mentioning
confidence: 99%