Bright and High-Performance Genetically Encoded Ca²⁺ Indicator Based on mNeonGreen Fluorescent Protein

Abstract

Genetically encodable calcium ion (Ca 2+ ) indicators (GECIs) based on green fluorescent proteins (GFP) are powerful tools for imaging of cell signaling and neural activity in model organisms. Following almost 2 decades of steady improvements in the Aequorea victoria GFP-based GCaMP series of GECIs, the performance of the most recent generation (i.e., jGCaMP7) may have reached its practical limit due to the inherent properties of GFP. In an effort to sustain the steady progression toward ever-improved GECIs, we undertook the development of a new GECI based on the bright monomeric GFP, mNeonGreen (mNG). The resulting indicator, mNG-GECO1, is 60% brighter than GCaMP6s in vitro and provides comparable performance as demonstrated by imaging Ca 2+ dynamics in cultured cells, primary neurons, and in vivo in larval zebrafish. These results suggest that mNG-GECO1 is a promising next-generation GECI that could inherit the mantle of GCaMP and allow the steady improvement of GECIs to continue for generations to come.

Design and Prototyping of Genetically Encoded Arsenic Biosensors Based on Transcriptional Regulator AfArsR

“…The conformational change of AfArsR upon binding to As 3+ was expected to alter the chromophore environment, thus changing the fluorescence emission intensity ( Figure 4 B). This topology has been previously reported for the construction of Ca 2+ , glutamate, K + , and citrate biosensors [ 26 , 29 , 30 , 31 ] among many others. We found that insertion of intact AfArsR into the GFP scaffold resulted in a prototype green fluorescent indicator, designated as GEAR-G1.…”

Medium-throughput zebrafish optogenetic platform identifies deficits in subsequent neural activity following brief early exposure to cannabidiol and Δ9-tetrahydrocannabinol

“…Nevertheless, locomotor activity is not a direct measurement of neural activity. Calcium sensors, such as the GCaMP series 57 , 58 , the more recently introduced GECO 59 , 60 , and CaMPARI, offer opportunity for more direct measures of neural activity. Importantly, CaMPARI provides a temporal-snapshot of neural activity in the freely swimming animal, which can be anesthetized and imaged after the experiment 34 .…”

Medium-throughput zebrafish optogenetic platform identifies deficits in subsequent neural activity following brief early exposure to cannabidiol and Δ-9-tetrahydrocannabinol

“…Nevertheless, locomotor activity is not a direct measurement of neural activity. Calcium sensors, such as the GCaMP series 57,58 , the more recently introduced GECO 59,60 , and CaMPARI, are more accurate. CaMPARI provides a temporal-snapshot of neural activity in the freely swimming animal, which can be anesthetized and imaged after the experiment 34 .…”