Investigating the developmental, structural and functional complexity of mammalian tissues and organs depends on identifying and gaining experimental access to diverse cell populations. Here, we ...describe a set of recombinase-responsive fluorescent indicator alleles in mice that significantly extends our ability to uncover cellular diversity by exploiting the intrinsic genetic signatures that uniquely define cell types. Using a recombinase-based intersectional strategy, these new alleles uniquely permit non-invasive labeling of cells defined by the overlap of up to three distinct gene expression domains. In response to different combinations of Cre, Flp and Dre recombinases, they express eGFP and/or tdTomato to allow the visualization of full cellular morphology. Here, we demonstrate the value of these features through a proof-of-principle analysis of the central noradrenergic system. We label previously inaccessible subpopulations of noradrenergic neurons to reveal details of their three-dimensional architecture and axon projection profiles. These new indicator alleles will provide experimental access to cell populations at unprecedented resolution, facilitating analysis of their developmental origin and anatomical, molecular and physiological properties.
Contextual fear learning is heavily dependent on the hippocampus. Despite evidence that catecholamines contribute to contextual encoding and memory retrieval, the precise temporal dynamics of their ...release in the hippocampus during behavior is unknown. In addition, new animal models are required to probe the effects of altered catecholamine synthesis on release dynamics and contextual learning.
We generated two new mouse models of altered locus coeruleus norepinephrine (LC-NE) synthesis and utilized them together with GRABNE and GRABDA sensors and in vivo fiber photometry to investigate norepinephrine (NE) and dopamine (DA) release dynamics in dorsal hippocampal CA1 during contextual fear conditioning.
We report that aversive foot-shock increases both NE and DA release in dorsal CA1, while freezing behavior associated with recall of fear memory is accompanied by decreased release. Moreover, we find that freezing at the recent timepoint is sensitive to both partial and complete loss of LC-NE synthesis throughout prenatal and postnatal development, similar to prior observations of mice with global loss of NE synthesis beginning postnatally. In contrast, freezing at the remote timepoint is compromised only by complete loss of LC-NE synthesis beginning prenatally.
Overall, these findings provide novel insights into the role of NE in contextual fear and the precise temporal dynamics of both NE and DA during freezing behavior, and highlight a complex relationship between genotype, sex, and NE signaling.
The default mode network (DMN) of the brain is functionally associated with a wide range of behaviors. In this study, we used functional magnetic resonance imaging (fMRI), positron emission ...tomography (PET), and spectral fiber photometry to investigate the selective neuromodulatory effect of norepinephrine (NE)-releasing noradrenergic neurons in the locus coeruleus (LC) on the mouse DMN. Chemogenetic-induced tonic LC activity decreased cerebral blood volume (CBV) and glucose uptake and increased synchronous low-frequency fMRI activity within the frontal cortices of the DMN. Fiber photometry results corroborated these findings, showing that LC-NE activation induced NE release, enhanced calcium-weighted neuronal spiking, and reduced CBV in the anterior cingulate cortex. These data suggest that LC-NE alters conventional coupling between neuronal activity and CBV in the frontal DMN. We also demonstrated that chemogenetic activation of LC-NE neurons strengthened functional connectivity within the frontal DMN, and this effect was causally mediated by reduced modulatory inputs from retrosplenial and hippocampal regions to the association cortices of the DMN.
Objective. To estimate performance characteristics and diagnostic value of immunofluorescent islet cell antibody (ICA) assay, immunoradiometric glutamic acid decarboxylase antibody (GADA) assay, and ...ELISA tyrosine phosphatase IA-2 antibody (IA-2A) and insulin antibody (IA) assays.Research Design and Methods. Antibodies were tested in 438 children and adolescents with newly diagnosed diabetes mellitus (DM) type 1, and in 891 subjects without DM type 1. ICA were determined by the classic indirect immunofluorescent method recommended by the Juvenile Diabetes Foundation International, GADA were determined with the Immunotech IRMA Anti-GAD kit, and IA-2A and IA were determined with Medizym Anti-IA2 and Orgentec Anti-Insulin ELISA kits, respectively. Sensitivity (Se), specificity (Sp), positive predictive value (PPV) and negative predictive value (NPV) of the tests were estimated with contingency tables. Diagnostic accuracy was estimated from areas under receiver operating curves (AUC).Results. ICA test was of the greatest diagnostic value (Se=88%, Sp=96%, PPV=96%, NPV=94%, AUC=0,94), followed by IA-2A (Se=66%, Sp=98%, PPV=98%, NPV=59%, AUC=0,82) and GADA (Se=73%, Sp=84%, PPV=75%, NPV=83%, AUC=0,79). IA test exhibited a very low Se (4,3%) and lacked diagnostic accuracy (AUC=0,5).Conclusions. We recommend to use ICA, IA-2A and GADA tests surveyed in our study for diagnosis of DM type 1 and differential diagnosis of DM. We don’t recommend IA testing with an Orgentec Anti-Insulin ELISA kit for usage in clinical practice.
Objective. To estimate performance characteristics and diagnostic value of immunofluorescent islet cell antibody (ICA) assay, immunoradiometric glutamic acid decarboxylase antibody (GADA) assay, and ...ELISA tyrosine phosphatase IA-2 antibody (IA-2A) and insulin antibody (IA) assays.Research Design and Methods. Antibodies were tested in 438 children and adolescents with newly diagnosed diabetes mellitus (DM) type 1, and in 891 subjects without DM type 1. ICA were determined by the classic indirect immunofluorescent method recommended by the Juvenile Diabetes Foundation International, GADA were determined with the Immunotech IRMA Anti-GAD kit, and IA-2A and IA were determined with Medizym Anti-IA2 and Orgentec Anti-Insulin ELISA kits, respectively. Sensitivity (Se), specificity (Sp), positive predictive value (PPV) and negative predictive value (NPV) of the tests were estimated with contingency tables. Diagnostic accuracy was estimated from areas under receiver operating curves (AUC).Results. ICA test was of the greatest diagnostic value (Se=88%, Sp=96%, PPV=96%, NPV=94%, AUC=0,94), followed by IA-2A (Se=66%, Sp=98%, PPV=98%, NPV=59%, AUC=0,82) and GADA (Se=73%, Sp=84%, PPV=75%, NPV=83%, AUC=0,79). IA test exhibited a very low Se (4,3%) and lacked diagnostic accuracy (AUC=0,5).Conclusions. We recommend to use ICA, IA-2A and GADA tests surveyed in our study for diagnosis of DM type 1 and differential diagnosis of DM. We don’t recommend IA testing with an Orgentec Anti-Insulin ELISA kit for usage in clinical practice.
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