Biology subjects

Nguyen, M. D.

Publications and source records attributed to Nguyen, M. D..

6 recordsLinked to original sources

bioRxiv2025

Structure-guided allosteric modulation of the delta opioid receptor

Opioid analgesics remain essential for pain management but are associated with significant adverse effects, including respiratory depression, tolerance, and dependence. The {delta}-opioid receptor ({delta}OR) represents a promising therapeutic target for developing safer opioid analgesics with reduced adverse effects compared to conventional -opioid receptor-targeting drugs. Positive allosteric modulators (PAMs) offer advantages over direct agonists by enhancing endogenous opioid signaling while preserving natural spatiotemporal activation patterns, potentially avoiding tolerance and dependence issues. Here, we present high-resolution cryo-EM structures of {delta}OR complexed with the peptide agonist DADLE and the PAM MIPS3614, revealing a novel lipid-facing allosteric binding site formed by transmembrane helices 2, 3, and 4. MIPS3614 stabilizes the active receptor conformation through a critical hydrogen bond with residue N1313.35 in the conserved sodium binding site, a key regulatory region controlling GPCR activation. Comprehensive mutagenesis, molecular dynamics simulations, and structure-activity relationships validate this proposed mechanism. Structure-guided optimization yielded MIPS3983 with enhanced binding affinity and retained cooperativity. Our findings establish the first molecular framework for {delta}OR allosteric modulation and provide a structural foundation for the rational design of safer opioid therapeutics.

pharmacology and toxicology↗

bioRxiv2025

A sensitive orange fluorescent calcium ion indicator for imaging neural activity

Genetically encoded calcium indicators (GECIs) are vital tools for fluorescence-based visualization of neuronal activity with high spatial and temporal resolution. However, current highest-performance GECIs are predominantly green or red fluorescent, limiting multiplexing options and efficient excitation with fixed-wavelength femtosecond lasers operating at 1030 nm. Here, we introduce OCaMP (also known as O-GECO2), an orange fluorescent GECI engineered from O-GECO1 through targeted substitutions to improve calcium affinity while retaining the favorable photophysical properties of mOrange2. OCaMP exhibits improved two-photon cross-section, responsiveness, photostability, and calcium affinity relative to O-GECO1. In cultured neurons, zebrafish, and mouse cortex, OCaMP outperforms the red GECIs jRCaMP1a and jRGECO1a in sensitivity, kinetics, and signal-to-noise ratio. These properties establish OCaMP as a robust tool for high-fidelity neural imaging optimized for 1030 nm excitation and a compromise-free option within the spectral gap between existing green and red GECIs.

neuroscience↗

bioRxiv2024

The gut microbiota is a determinant of sexual dimorphism in ALS-linked TDP43 mice

The mechanisms underlying the earlier onset and male predominance of amyotrophic lateral sclerosis (ALS), the most common form of human motoneuron disease, are poorly understood. Here we show that the gut microbiota protects against TDP43 toxicity and contributes to the sexual dimorphism in mice expressing a mutant form of TDP43 (A315T) linked to ALS. TDP43 mice raised under germ-free conditions, or treated with antibiotics to deplete the gut microbiota, develop motoneuron disease earlier and show no sex differences in onset and lifespan. Behavioral and histopathological analyses confirm the exacerbation in neurodegeneration caused by the absence of gut microbiota. Castration did not alter disease course of male TDP43 mice, suggesting that male sex hormones do not interact with the gut microbiota to confer disease phenotype. Future identification of gut bacteria species and their mechanisms of action offers a unique opportunity to understand sexual dimorphism in ALS, with the ultimate goal to develop non-invasive and sex-specific treatments for ALS.

physiology↗

bioRxiv2022

A Protocol for Single Nucleus RNAseq from Frozen Skeletal Muscle

Single cell technologies are a method of choice to obtain vast amounts of cell-specific transcriptional information under physiological and diseased states. Myogenic cells are resistant to single nucleus RNA sequencing (snRNAseq) due to their large, multinucleated nature. Here, we report a novel, reliable, and cost-effective method to analyze frozen human skeletal muscle by snRNAseq. This method yields all expected cell types for human skeletal muscle and works on tissue frozen for long periods of time and with significant pathological changes. Our method is ideal for studying banked samples with the intention of studying human muscle disease.

genomics↗

bioRxiv2020

Astrocytes amplify cerebral blood flow elevation to sustained cortical activation in the awake mouse

Brain requires increased local cerebral blood flow (CBF) for as long as necessary during neuronal activation to match O2 and glucose supply with demand - termed functional hyperemia. Ca2+ elevation in astrocytes can drive arteriole dilation to increase CBF, yet affirmative evidence for the necessity of astrocytes in functional hyperemia in vivo is lacking. In awake and active mice, we discovered that functional hyperemia is bimodal with a distinct early and late component whereby arteriole dilation progresses as sensory stimulation is sustained. Clamping astrocyte Ca2+ signaling in vivo by expressing a high-affinity plasma membrane Ca2+ ATPase (CalEx) reduces sustained but not brief sensory-evoked arteriole dilation. Reciprocally, elevating astrocyte free Ca2+ using chemogenetics selectively augments sustained but not brief hyperemia. Neither locomotion, arousal, nor changes in neuronal signaling account for the selective effect of astrocyte Ca2+ on the late phase of the CBF response. Antagonizing NMDA-receptors or epoxyeicosatrienoic acid production reduces only the late component of functional hyperemia, leaving brief increases in CBF to sensory stimulation intact. We propose that a fundamental role of astrocyte Ca2+ is to amplify functional hyperemia when neuronal activation is prolonged.

neuroscience↗

bioRxiv2020

Deregulation of brain endothelial CD2AP in Alzheimer's disease impairs Reelin-mediated neurovascular coupling

Genetic variations in CD2-associated protein (CD2AP) predispose to Alzheimers disease (AD) but the underlying mechanisms remain unknown. Here, we show that a cerebrovascular loss of CD2AP is associated with cognitive decline in AD and that genetic downregulation of CD2AP in brain endothelial cells impairs memory function in two distinct mouse models. Mice with reduced CD2AP in brain microvessels display decreased resting cerebral blood flow, impaired functional hyperemia and vasomotion. In brain endothelial cells, CD2AP regulates the levels and signaling of ApoE receptor 2 elicited by Reelin glycoprotein. Activation of the CD2AP-ApoER2 pathway with Reelin mitigates the toxic effects of A{beta} on resting blood flow and vasomotion of brain vessels depleted of CD2AP. Thus, we demonstrate that deregulation of CD2AP perturbs specific functions and segments of the cerebral microvasculature and propose that targeting CD2AP molecular partners may offer refined therapeutic strategies for the treatment of AD.

neuroscience↗