Organic electrochemical transistor arrays for real-time mapping of evoked neurotransmitter release in vivo
Abstract
Though neurotransmitters are essential in neural signal transmission, techniques for in vivo analysis are still limited. Here, we describe an organic electrochemical transistor array (OECT-array) technique for monitoring catecholamine neurotransmitters (CA-NTs) in rat brains. The OECT-array is an active sensor with intrinsic amplification capability, allowing real-time and direct readout of transient CA-NT release with a sensitivity of nanomolar range and a temporal resolution of several milliseconds. The device has a working voltage lower than half of that typically used in a prevalent cyclic voltammetry measurement, and operates continuously in vivo for hours without significant signal drift, which is inaccessible for existing methods. With the OECT-array, we demonstrate simultaneous mapping of evoked dopamine release at multiple striatal brain regions in different physiological scenarios, and reveal a complex cross-talk between mesolimbic and nigrostriatal pathways, which is heterogeneously affected by the reciprocal innervation between ventral tegmental area and substantia nigra pars compacta.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided as supplementary files.
Article and author information
Author details
Funding
General Research Funds (11278616)
- Peng Shi
General Research Funds (11218015)
- Peng Shi
General Research Funds (11203017)
- Peng Shi
Health and Medical Research Fund (06172336)
- Peng Shi
Collaborative Research Funds (C5015-15G)
- Feng Yan
- Peng Shi
The funders provided resources for the study design, data collection, and interpretation.
Ethics
Animal experimentation: All experimental procedures involving animals were approved by the university Animal Ethics Committee. Animal licenses, (16-97) in DH/HA&P/8/2/5 Pt.5 and (18-129) in DH/SHS/8/2/5 Pt.4, were approved by Department of Health of the Government of Hong Kong Special Administration Region.
Copyright
© 2020, Xie et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Biochemistry and Chemical Biology
- Computational and Systems Biology
The spike protein is essential to the SARS-CoV-2 virus life cycle, facilitating virus entry and mediating viral-host membrane fusion. The spike contains a fatty acid (FA) binding site between every two neighbouring receptor-binding domains. This site is coupled to key regions in the protein, but the impact of glycans on these allosteric effects has not been investigated. Using dynamical nonequilibrium molecular dynamics (D-NEMD) simulations, we explore the allosteric effects of the FA site in the fully glycosylated spike of the SARS-CoV-2 ancestral variant. Our results identify the allosteric networks connecting the FA site to functionally important regions in the protein, including the receptor-binding motif, an antigenic supersite in the N-terminal domain, the fusion peptide region, and another allosteric site known to bind heme and biliverdin. The networks identified here highlight the complexity of the allosteric modulation in this protein and reveal a striking and unexpected link between different allosteric sites. Comparison of the FA site connections from D-NEMD in the glycosylated and non-glycosylated spike revealed that glycans do not qualitatively change the internal allosteric pathways but can facilitate the transmission of the structural changes within and between subunits.
-
- Biochemistry and Chemical Biology
- Genetics and Genomics
Deep Mutational Scanning (DMS) is an emerging method to systematically test the functional consequences of thousands of sequence changes to a protein target in a single experiment. Because of its utility in interpreting both human variant effects and protein structure-function relationships, it holds substantial promise to improve drug discovery and clinical development. However, applications in this domain require improved experimental and analytical methods. To address this need, we report novel DMS methods to precisely and quantitatively interrogate disease-relevant mechanisms, protein-ligand interactions, and assess predicted response to drug treatment. Using these methods, we performed a DMS of the melanocortin-4 receptor (MC4R), a G-protein-coupled receptor (GPCR) implicated in obesity and an active target of drug development efforts. We assessed the effects of >6600 single amino acid substitutions on MC4R’s function across 18 distinct experimental conditions, resulting in >20 million unique measurements. From this, we identified variants that have unique effects on MC4R-mediated Gαs- and Gαq-signaling pathways, which could be used to design drugs that selectively bias MC4R’s activity. We also identified pathogenic variants that are likely amenable to a corrector therapy. Finally, we functionally characterized structural relationships that distinguish the binding of peptide versus small molecule ligands, which could guide compound optimization. Collectively, these results demonstrate that DMS is a powerful method to empower drug discovery and development.