Biology subjects

Restrepo, D.

Publications and source records attributed to Restrepo, D..

27 records · Page 2Linked to original sources

bioRxiv2020

Transcriptional profiling reveals TRPM5-expressing cells involved in viral infection in the olfactory epithelium

BackgroundUnderstanding viral infection of the olfactory epithelium is essential because the olfactory nerve is an important route of entry for viruses to the central nervous system. Specialized chemosensory epithelial cells that express the transient receptor potential cation channel subfamily M member 5 (TRPM5) are found throughout the airways and intestinal epithelium and are involved in responses to viral infection. ResultsHerein we performed deep transcriptional profiling of olfactory epithelial cells sorted by flow cytometry based on the expression of mCherry as a marker for olfactory sensory neurons and for eGFP in OMP-H2B::mCherry/TRPM5-eGFP transgenic mice (Mus musculus). We find profuse expression of transcripts involved in inflammation, immunity and viral infection in TRPM5-expressing microvillous cells. ConclusionOur study provides new insights into a potential role for TRPM5-expressing microvillous cells in viral infection of the olfactory epithelium. We find that, as found for solitary chemosensory cells (SCCs) and brush cells in the airway epithelium, and for tuft cells in the intestine, the transcriptome of TRPM5-expressing microvillous cells indicates that they are likely involved in the inflammatory response elicited by viral infection of the olfactory epithelium.

neuroscience↗

bioRxiv2020

Hyperexcitability and loss of feedforward inhibition in the Fmr1KO lateral amygdala.

Fragile X Syndrome (FXS) is a neurodevelopmental disorder characterized by intellectual disability, autism spectrum disorders (ASDs), and anxiety disorders. The disruption in the function of the FMR1 gene results in a range of alterations in cellular and synaptic function. Previous studies have identified dynamic alterations in inhibitory neurotransmission in early postnatal development in the amygdala of the mouse model of FXS. Yet little is known how these changes alter microcircuit development and plasticity in the lateral amygdala (LA). Using whole-cell patch clamp electrophysiology, we demonstrate that principal neurons (PNs) in the LA exhibit hyperexcitability with a concomitant increase in the synaptic strength of excitatory synapses in the BLA. Further, reduced feed-forward inhibition appears to enhance synaptic plasticity in the FXS amygdala. These results demonstrate that plasticity is enhanced in the amygdala of the juvenile Fmr1 KO mouse and that E/I imbalance may underpin anxiety disorders commonly seen in FXS and ASDs.

neuroscience↗

bioRxiv2020

An optogenetic switch for the Set2 methyltransferase provides evidence for rapid transcription-dependent and independent dynamics of H3K36 methylation

Histone H3 lysine 36 methylation (H3K36me) is a conserved histone modification associated with transcription and DNA repair. Although the effects of H3K36 methylation have been studied, the short-term genome-wide dynamics of H3K36me deposition and removal are not known. We established rapid and reversible optogenetic control for Set2, the sole H3K36 methyltransferase in yeast, by fusing the enzyme with the light activated nuclear shuttle (LANS) domain. Early H3K36me3 dynamics identified rapid methylation in vivo, with total H3K36me3 levels correlating with RNA abundance. Although genes exhibited disparate levels of H3K36 methylation, relative rates of H3K36me3 accumulation were largely linear and consistent across genes, suggesting a rate-limiting mechanism for H3K36me3 deposition. Removal H3K36me2/3 was also rapid and highly dependent on the demethylase Rph1. However, the per-gene rate of H3K36me3 loss weakly correlated with RNA abundance and followed exponential decay, suggesting H3K36 demethylases act in a global, stochastic manner.

molecular biology↗

bioRxiv2019

Molecular layer interneurons in the cerebellum encode for valence in associative learning

We used two-photon microscopy to study the role of ensembles of cerebellar molecular layer interneurons (MLIs) in a go-no go task where mice obtain a sugar water reward if they lick a spout in the presence of the rewarded odorant and avoid a time out when they refrain from licking for the unrewarded odorant. In naive animals the MLI responses did not differ between the odorants. With learning, the rewarded odorant elicited a large increase in MLI calcium responses, and the identity of the odorant could be decoded from the differential response. Importantly, MLIs switched odorant responses when the valence of the stimuli was reversed. Finally, mice took a longer time to refrain from licking in the presence of the unrewarded odorant and had difficulty becoming proficient when MLIs were inhibited by chemogenetic intervention. Our findings support a role for MLIs in learning valence in the cerebellum.

neuroscience↗

bioRxiv2019

Learning improves decoding of odor identity with phase-referenced oscillations in the olfactory bulb

Local field potential oscillations reflect temporally coordinated neuronal ensembles-- coupling distant brain regions, gating processing windows, and providing a reference for spike timing-based codes. In phase amplitude coupling (PAC), the amplitude of the envelope of a faster oscillation is larger within a phase window of a slower carrier wave. Here, we characterized PAC, and the related theta phase-referenced high gamma and beta power (PRP), in the olfactory bulb of mice learning to discriminate odorants. PAC changes throughout learning, and odorant-elicited changes in PRP increase for rewarded and decrease for unrewarded odorants. Contextual odorant identity (is the odorant rewarded?) can be decoded from peak PRP in animals proficient in odorant discrimination, but not in naive mice. As the animal learns to discriminate the odorants the dimensionality of PRP decreases. Therefore, modulation of phase-referenced chunking of information in the course of learning plays a role in early sensory processing in olfaction. SignificanceEarly processing of olfactory information takes place in circuits undergoing slow frequency theta oscillations generated by the interplay of olfactory input modulated by sniffing and centrifugal feedback from downstream brain areas. Studies in the hippocampus and cortex suggest that different information "chunks" are conveyed at different phases of the theta oscillation. Here we show that in the olfactory bulb, the first processing station in the olfactory system, the amplitude of high frequency gamma oscillations encodes for information on whether an odorant is rewarded when it is observed at the peak phase of the theta oscillation. Furthermore, encoding of information by the theta phase-referenced gamma oscillations becomes more accurate as the animal learns to differentiate two odorants.

neuroscience↗

bioRxiv2019

Differential Expression of Mucins in Murine Olfactory Versus Respiratory Epithelium

Mucins are a key component of the airway surface liquid and serve many functions. Given the numerous differences in olfactory versus respiratory nasal epithelia, we hypothesized that mucins would be differentially expressed between these two areas. Secondarily, we evaluated for changes in mucin expression with radiation exposure, given the clinical observations of nasal dryness, altered mucus rheology, and smell loss in radiated patients. Immunofluorescence staining was performed in a mouse model to determine the expression of mucins 1, 2, 5AC and 5B in nasal respiratory and olfactory epithelia of control mice and one week after exposure to 8 gy of radiation. Mucins 1, 5AC and 5B exhibited differential expression between olfactory and respiratory epithelium while mucin 2 showed no difference. Within the olfactory epithelium, mucin 1 was located in a lattice-like pattern around gaps corresponding to dendritic knobs of olfactory sensory neurons, whereas in respiratory epithelium it was only intermittently expressed. Mucin 5AC was expressed by subepithelial glands in both epithelial types but to a higher degree in the olfactory epithelium. Mucin 5B was expressed by submucosal glands in the olfactory epithelium but by surface epithelial cells in respiratory epithelium. At one-week after exposure to single-dose 8 gy of radiation, no qualitative effects were seen on mucin expression. Our findings demonstrate that murine olfactory and respiratory epithelia express mucins differently, and characteristic patterns of mucins 1, 5AC, and 5B can be used to define the underlying epithelium. Radiation (8 gy) does not appear to affect mucin expression at one week.\n\nAuthor RolesChristopher Kennel conceived, organized and executed the study, performed the analysis, and contributed to the manuscript.\n\nElizabeth Gould conceived and executed the study, and contributed to the manuscript.\n\nDiego Restrepo conceived and executed the study, supervised the experiments, reviewed the analysis, and contributed to the manuscript.\n\nErnesto Salcedo performed experiments and reviewed the manuscript.\n\nThad Vickery performed experiments and reviewed the manuscript.\n\nEric Larson performed experiments and reviewed the manuscript.\n\nVijay Ramakrishnan conceived and executed the study, reviewed the analysis, and contributed to the manuscript.\n\nAll authors discussed the results and implications and contributed to the final manuscript.

neuroscience↗

bioRxiv2018

Homeostatic Inhibitory Control of Cortical Hyperexcitability in Fragile X Syndrome

In mouse models of Fragile X Syndrome (FXS), cellular and circuit hyperexcitability are a consequence of altered brain development [reviewed in (Contractor et al., 2015)]. Mechanisms that favor or hinder plasticity of synapses could affect neuronal excitability. This includes inhibitory long term depression (I-LTD) - a heterosynaptic form of plasticity that requires the activation of metabotropic glutamate receptors (mGluRs). Differential circuit maturation leads to shifted time points for critical periods of synaptic plasticity across multiple brain regions (Harlow et al., 2010; He et al., 2014), and disruptions of the development of excitatory and inhibitory synaptic function are also observed both during development and into adulthood (Vislay et al., 2013). However, little is known about how this hyperexcitable environment affects inhibitory synaptic plasticity. Our results demonstrate that the somatosensory cortex of the Fmr1 KO mouse model of FXS exhibits increased GABAergic spontaneous activity, a faulty mGluR-mediated inhibitory input and impaired plasticity processes. We find the overall diminished mGluR activation in the Fmr1 KO mice leads to both a decreased spontaneous inhibitory postsynaptic input to principal cells and also to a disrupted form of inhibitory long term depression (I-LTD). In cortical synapses, this I-LTD is dependent on mGluR activation and the mobilization endocannabinoids (eCBs). Notably, these data suggest enhanced hyperexcitable phenotypes in FXS may be homeostatically counterbalanced by the inhibitory drive of the network and its altered response to mGluR modulation.\n\nSignificance StatementFragile X Syndrome is a pervasive neurodevelopmental disorder characterized by intellectual disability, autism, epilepsy, anxiety and altered sensory sensitivity. In both in vitro and in vivo recordings in the somatosensory cortex of the Fmr1 knockout mouse model of Fragile X Syndrome we show that hyperexcitable network activity contributes to ineffective synaptic plasticity at inhibitory synapses. This increased excitability prevents cortical circuits from adapting to sensory information via ineffective plasticity mechanisms.

neuroscience↗

bioRxiv2018

Cell type specific control of basolateral amygdala plasticity via feedforward inhibition

The basolateral amygdala (BLA) plays a vital role in associating specific sensory stimuli with salient valence information. Excitatory principal neurons (PNs) undergo plastic changes to encode this integrated sensory-valence information; however, local BLA inhibitory interneurons (INs) gate the plasticity of the PNs via feed forward inhibition (FFI). Despite extensive literature implicating parvalbumin expressing (PV+) INs in FFI in cortex and hippocampus, prior anatomical experiments in BLA implicate somatostatin expressing (Sst+) INs in BLA. In the present study, we combined patch clamp electrophysiology with chemogenetics, unsupervised cluster analysis, and predictive modeling and found that a previously unreported subpopulation of fast-spiking Sst+ INs mediate BLA FFI and gate plasticity.

neuroscience↗

bioRxiv2018

Three dimensional two-photon imaging of neuronal activity in freely moving mice using a miniature fiber coupled microscope with active axial-scanning

We present a miniature head mounted two-photon fiber-coupled microscope (2P-FCM) for neuronal imaging with active axial focusing enabled using a miniature electrowetting lens. Full three-dimensional two-photon imaging of GCaMP6s showing individual neuron activity in multiple focal planes was achieved in a freely-moving mouse. Two-color simultaneous imaging of GFP and tdTomato fluorescence is also demonstrated. Additionally, dynamic control of the axial scanning of the electrowetting lens allows tilting of the focal plane enabling cells in multiple focal planes to be imaged simultaneously. Two-photon imaging allows increased penetration depth in tissue yielding a working distance of 450 m with an additional 180 m of active axial focusing. The objective NA is 0.45 with a lateral resolution of 1.8 m, an axial resolution of 10 m, and a field-of-view of 240 m diameter. The 2P-FCM has a weight of only [~]2.5 g and is capable of repeatable and stable head-attachment. The 2P-FCM with dynamic axial scanning provides a new capability to record from functionally distinct neuronal layers, opening new opportunities in neuroscience research.

neuroscience↗