Behavioral context improves optogenetic stimulation of transplanted dopaminergic cells in unilateral 6-OHDA rats

Anderson, Kevin; Whitehead, Bailey; Petersen, Eric D.; Kemme, Madison R; Wedster, Anna; Hochgeschwender, Ute; Sandstrom, Michael I

doi:10.1016/j.bbr.2022.114279

Cited by 2 publications

(2 citation statements)

References 49 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Additionally, now that we have created BLING variants with diverse properties such as varying levels of background luminescence and responses to their neurotransmitter (Figure ) we can create a variety of neuromodulators based on these sensors tailored to specific applications. These can then be used to improve our prior and ongoing work in neurodegenerative disorders such as spinal cord injury and Parkinson’s disease by allowing the noninvasive current stimulation of neurons to be dependent on endogenous activity. ,,− These new BLINGs also present ample opportunity as multiple starting points for further evolution to create new BLINGs with greater responses to glutamate and lower background. Specifically improving BLINGs to have higher affinity to glutamate, in the 100–500 micromolar range to respond to fluctuations in glutamate on the lower end of the physiological spectrum and by improving membrane trafficking and trafficking the sensor specifically to the synaptic cleft as was recently done with fluorescent glutamate sensors…”

Section: Discussionmentioning

confidence: 99%

Bioluminescent Genetically Encoded Glutamate Indicators for Molecular Imaging of Neuronal Activity

et al. 2023

Self Cite

View full text Add to dashboard Cite

Genetically encoded optical sensors and advancements in microscopy instrumentation and techniques have revolutionized the scientific toolbox available for probing complex biological processes such as release of specific neurotransmitters. Most genetically encoded optical sensors currently used are based on fluorescence and have been highly successful tools for single-cell imaging in superficial brain regions. However, there remains a need to develop new tools for reporting neuronal activity in vivo within deeper structures without the need for hardware such as lenses or fibers to be implanted within the brain. Our approach to this problem is to replace the fluorescent elements of the existing biosensors with bioluminescent elements. This eliminates the need of external light sources to illuminate the sensor, thus allowing deeper brain regions to be imaged noninvasively. Here, we report the development of the first genetically encoded neurotransmitter indicators based on bioluminescent light emission. These probes were optimized by high-throughput screening of linker libraries. The selected probes exhibit robust changes in light output in response to the extracellular presence of the excitatory neurotransmitter glutamate. We expect this new approach to neurotransmitter indicator design to enable the engineering of specific bioluminescent probes for multiple additional neurotransmitters in the future, ultimately allowing neuroscientists to monitor activity associated with a specific neurotransmitter as it relates to behavior in a variety of neuronal and psychiatric disorders, among many other applications.

show abstract

Section: Discussionmentioning

confidence: 99%

Bioluminescent Genetically Encoded Glutamate Indicators for Molecular Imaging of Neuronal Activity

et al. 2023

Self Cite

View full text Add to dashboard Cite

show abstract

“…Efficiently changing membrane potentials in neurons required the use of a brighter light emitter such as the GLuc variant ‘slow burn’ (sbGLuc) 9 in combination with optogenetic channels with higher light sensitivity, such as Volvox channelrhodopsin 1 (VChR1) 10-11 . This combination in LMO3 yielded robust and reliable activation of neurons in numerous in vivo experiments 5-6,12-19 . We and others have since continued to combine more potent light emitters with more light sensitive opsins to generate LMOs with still higher efficacies.…”

Section: Introductionmentioning

confidence: 99%

Engineering luminopsins with improved coupling efficiencies

Slaviero,

Gorantla,

Simkins

et al. 2023

Preprint

Self Cite

View full text Add to dashboard Cite

SignificanceLuminopsins (LMOs) are bioluminescent-optogenetic tools with a luciferase fused to an opsin that allow bimodal control of neurons by providing both optogenetic and chemogenetic access. Determining which design features contribute to the efficacy of LMOs will be beneficial for further improving LMOs for use in research.AimWe investigated the relative impact of luciferase brightness, opsin sensitivity, pairing of emission and absorption wavelength, and arrangement of moieties on the function of LMOs.ApproachWe quantified efficacy of LMOs through whole cell patch clamp recordings in HEK293 cells by determining coupling efficiency, the percentage of maximum LED induced photocurrent achieved with bioluminescent activation of an opsin. We confirmed key results by multielectrode array (MEAs) recordings in primary neurons.ResultsLuciferase brightness and opsin sensitivity had the most impact on the efficacy of LMOs, and N-terminal fusions of luciferases to opsins performed better than C-terminal and multi-terminal fusions. Precise paring of luciferase emission and opsin absorption spectra appeared to be less critical.ConclusionsWhole cell patch clamp recordings allowed us to quantify the impact of different characteristics of LMOs on their function. Our results suggest that coupling brighter bioluminescent sources to more sensitive opsins will improve LMO function. As bioluminescent activation of opsins is most likely based on Förster resonance energy transfer (FRET), the most effective strategy for improving LMOs further will be molecular evolution of luciferase-fluorescent protein-opsin fusions.

show abstract

Behavioral context improves optogenetic stimulation of transplanted dopaminergic cells in unilateral 6-OHDA rats

Cited by 2 publications

References 49 publications

Bioluminescent Genetically Encoded Glutamate Indicators for Molecular Imaging of Neuronal Activity

Bioluminescent Genetically Encoded Glutamate Indicators for Molecular Imaging of Neuronal Activity

Engineering luminopsins with improved coupling efficiencies

Contact Info

Product

Resources

About