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Combined Optogenetic Approaches Reveal Quantitative Dynamics of Endogenous Noradrenergic Transmission in the Brain.

ABSTRACT: Little is known about the real-time cellular dynamics triggered by endogenous catecholamine release despite their importance in brain functions. To address this issue, we expressed channelrhodopsin in locus coeruleus neurons and protein kinase-A activity biosensors in cortical pyramidal neurons and combined two-photon imaging of biosensors with photostimulation of locus coeruleus cortical axons, in acute slices and in vivo. Burst photostimulation of axons for 5-10 s elicited robust, minutes-lasting kinase-A activation in individual neurons, indicating that a single burst firing episode of synchronized locus coeruleus neurons has rapid and lasting effects on cortical network. Responses were mediated by ?1 adrenoceptors, dampened by co-activation of ?2 adrenoceptors, and dramatically increased upon inhibition of noradrenaline reuptake transporter. Dopamine receptors were not involved, showing that kinase-A activation was due to noradrenaline release. Our study shows that noradrenergic transmission can be characterized with high spatiotemporal resolution in brain slices and in vivo using optogenetic tools.

SUBMITTER: Nomura S

PROVIDER: S-EPMC7645030 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Combined Optogenetic Approaches Reveal Quantitative Dynamics of Endogenous Noradrenergic Transmission in the Brain.

Nomura Shinobu S Tricoire Ludovic L Cohen Ivan I Kuhn Bernd B Lambolez Bertrand B Hepp Régine R

iScience 20201021 11

Little is known about the real-time cellular dynamics triggered by endogenous catecholamine release despite their importance in brain functions. To address this issue, we expressed channelrhodopsin in locus coeruleus neurons and protein kinase-A activity biosensors in cortical pyramidal neurons and combined two-photon imaging of biosensors with photostimulation of locus coeruleus cortical axons, in acute slices and <i>in vivo</i>. Burst photostimulation of axons for 5-10 s elicited robust, minut ...[more]

PMID: 33196030

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Project description:BackgroundEnvironmental mycobacteria (EM) include species commonly found in various terrestrial and aquatic environments, encompassing animal and human pathogens in addition to saprophytes. Approximately 150 EM species can be separated into fast and slow growers based on sequence and copy number differences of their 16S rRNA genes. Cultivation methods are not appropriate for diversity studies; few studies have investigated EM diversity in soil despite their importance as potential reservoirs of pathogens and their hypothesized role in masking or blocking M. bovis BCG vaccine.MethodsWe report here the development, optimization and validation of molecular assays targeting the 16S rRNA gene to assess diversity and prevalence of fast and slow growing EM in representative soils from semi tropical and temperate areas. New primer sets were designed also to target uniquely slow growing mycobacteria and used with PCR-DGGE, tag-encoded Titanium amplicon pyrosequencing and quantitative PCR.ResultsPCR-DGGE and pyrosequencing provided a consensus of EM diversity; for example, a high abundance of pyrosequencing reads and DGGE bands corresponded to M. moriokaense, M. colombiense and M. riyadhense. As expected pyrosequencing provided more comprehensive information; additional prevalent species included M. chlorophenolicum, M. neglectum, M. gordonae, M. aemonae. Prevalence of the total Mycobacterium genus in the soil samples ranged from 2.3×10(7) to 2.7×10(8) gene targets g(-1); slow growers prevalence from 2.9×10(5) to 1.2×10(7) cells g(-1).ConclusionsThis combined molecular approach enabled an unprecedented qualitative and quantitative assessment of EM across soil samples. Good concordance was found between methods and the bioinformatics analysis was validated by random resampling. Sequences from most pathogenic groups associated with slow growth were identified in extenso in all soils tested with a specific assay, allowing to unmask them from the Mycobacterium whole genus, in which, as minority members, they would have remained undetected.

Dataset Information

Combined Optogenetic Approaches Reveal Quantitative Dynamics of Endogenous Noradrenergic Transmission in the Brain.

Publications

Combined Optogenetic Approaches Reveal Quantitative Dynamics of Endogenous Noradrenergic Transmission in the Brain.

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