Biology subjects

Hughes, B. W.

Publications and source records attributed to Hughes, B. W..

7 recordsLinked to original sources

bioRxiv2026

A preclinical platform for focused ultrasound delivery from microplate to awake brain

Background. Focused ultrasound (FUS) has broad therapeutic potential, but its mechanisms of action are poorly understood. Preclinical FUS research enables rigorous interrogation of understudied parameters, but fragmented, single-purpose benchtop devices prevent standardized acoustic exposure across models. Objective. To describe a unified preclinical FUS platform for cell culture, ex vivo tissue, and awake stereotaxic in vivo delivery in one lightweight system, validated using glioblastoma as a disease model. Methods. We performed single-well FUS delivery to 96-well microplates, single-well delivery to freshly resected human glioblastoma tissue, and stereotaxic transcranial delivery to awake, head-fixed mice ambulating freely on a floating cage. Output was independently calibrated in free field, with measured microplate insertion loss. Pulsed low-intensity focused ultrasound (pLIFU) was applied to GL261-bearing mice, resected human glioblastoma, and macrophage cell lines. Results. Awake stereotaxic pLIFU produced acute tumor signal attenuation in GL261-bearing mice (mean log2 fold-change -1.66 versus +5.20 controls, p = 0.016) with bioluminescence evidence consistent with focal targeting. Ex vivo pLIFU produced directionally consistent immunophenotypic shifts in human glioblastoma (increased iNOS+, decreased Arg1+ populations). In vitro, pLIFU reduced CD80 on immunosuppressive macrophages to 73% of baseline in RAW 264.7 cells (p = 0.0008, n = 5) and 64% in THP-1 cells (p = 0.0006, n = 2), with MHC II upregulation in RAW 264.7 (ratio 1.65, p = 0.004) and ARG1 reduction in THP-1 (ratio 0.52, p = 0.0025). Conclusions. A unified preclinical FUS platform validated across in vitro, ex vivo, and awake in vivo applications enables stereotaxically guided transcranial FUS in awake, head-fixed mice ambulating in an exploratory manner during sonication, and single-well microplate treatments.

neuroscience↗

bioRxiv2025

Epigenetic editing of Cartpt promotes acquisition and extinction of cocaine memory

In classic disease models, removing a pathological insult restores homeostasis. Yet, addiction persists far beyond the period of active drug use. Cocaine abstinence induces changes in gene expression and neuronal signaling in reward-related brain regions that limit recovery during abstinence. We found that 2 weeks of abstinence increased Cartpt (cocaine- and amphetamine-regulated transcript) in the mouse nucleus accumbens and decreased repressive H3K27me3 at the Cartpt locus. While endogenous CART peptide is best described for its anorexigenic function, it is also implicated in human addiction and dopamine homeostasis. To test the causal relevance of Cartpt chromatin remodeling, we used CRISPR-based epigenetic editing tools, dCas9-FOG1 and dCas9-JMJC-ZF, to manipulate H3K27me3 at Cartpt in vivo. Enriching H3K27me3 in D1 neurons repressed Cartpt expression and augmented acquisition and extinction of cocaine preference. These results show that CRISPR epigenetic editing can recapitulate endogenous chromatin states to modulate addiction-related behavior, highlighting broad therapeutic potential of both Cartpt and epigenetic editing. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=192 SRC="FIGDIR/small/689329v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1b84229org.highwire.dtl.DTLVardef@1ffc2d7org.highwire.dtl.DTLVardef@50cf00org.highwire.dtl.DTLVardef@1462d10_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Discovery of Small Molecules and a Druggable Groove That Regulate DNA Binding and Release of the AP1 Transcription Factor DeltaFOSB

{Delta}FOSB, a member of the AP1 family of transcription factors, mediates long-term neuroadaptations underlying drug addiction, seizure-related cognitive decline, dyskinesias, and several other chronic conditions. AP1 transcription factors are notoriously difficult to modulate pharmacologically due to the absence of well-defined binding pockets. Here, we identify a novel site on {Delta}FOSB, located outside the DNA-binding cleft, that accommodates small molecules. We show that sulfonic acid-containing compounds bind to this site via an induced-fit mechanism, reorienting side chains critical for DNA binding, and that they may hinder the {Delta}FOSB bZIP -helix from binding to the major groove of DNA. In vivo, direct administration of one such compound, JPC0661, into the brain reduces {Delta}FOSB occupancy at genomic AP1 consensus sites by approximately 60% as determined by CUT&RUN-sequencing. These findings suggest that DNA binding and release by AP1 transcription factors can be controlled via small molecules that dock into a novel site that falls outside of the DNA-binding cleft. Minimal sequence conservation across 29 bZIP domain-containing transcription factors in this druggable groove suggests that it can be exploited to develop AP1-subunit-selective compounds. Our studies thus reveal a novel strategy to design small-molecule inhibitors of {Delta}FOSB and other members of the bZIP transcription factor family.

neuroscience↗

bioRxiv2025

Efficient in vivo pharmacological inhibition of deltaFOSB, an AP1 transcription factor, in brain

{Delta}FOSB, an unusually stable member of the AP1 family of transcription factors, mediates long-term maladaptations that play a key role in the pathogenesis of drug addiction, cognitive decline, dyskinesias, and several other chronic neurological and psychiatric conditions. We have recently identified that 2-phenoxybenzenesulfonic acid-containing compounds disrupt the binding of {Delta}FOSB to DNA in vitro in cell-based assays, and one such compound, JPC0661, disrupts {Delta}FOSB binding to genomic DNA in vivo in mouse brain with partial efficiency. JPC0661 binds to a groove outside of the DNA-binding cleft of the {Delta}FOSB/JUND bZIP heterodimer in the co-crystal structure. Here, we generated a panel of analogs of JPC0661 with the goal of establishing structure-activity relationships and improving its in vivo efficacy by replacing the amino-pyrazolone cap moiety with various substituents. We show that one such analog, YL0441, disrupts the binding of {Delta}FOSB to DNA in vitro and in vivo, and suppresses {Delta}FOSB-function in cell-based assays. Importantly, infusion of YL0441 into the hippocampus of APP mice (a mouse model for Alzheimers disease) leads to virtually complete loss of {Delta}FOSB bound to genomic DNA by CUT&RUN sequencing. Our findings corroborate that DNA binding/release of AP1 transcription factors can be controlled via small molecules, even by analogs of a compound that binds to a groove outside of the DNA-binding cleft, and that our lead can be optimized via medicinal chemistry to yield a highly efficacious inhibitor of {Delta}FOSB function in vivo. These findings define a strategy to design small-molecule inhibitors for other AP1- and AP1-related transcription factors. IN BRIEFWe demonstrate the creation of a highly effective inhibitor, YL0441, of {Delta}FOSB, an AP1 transcription factor, which decreases the number of {Delta}FOSB-bound sites to genomic DNA by [~]94% upon in vivo infusion to the hippocampus of APP mice, a mouse model for Alzheimers disease. This work generates a highly novel probe compound to assess the therapeutic value of {Delta}FOSB in vivo, a transcription factor with a critical role in mediating long-term changes in gene expression in several neuropsychiatric disorders in addition to Alzheimers disease, including drug addiction, seizure-related cognitive decline, and dyskinesias.

neuroscience↗

bioRxiv2023

The Role of Ryanodine Receptor 2 in Drug-Associated Learning

Type-2 ryanodine receptor (RyR2) ion channels facilitate the release of Ca2+ from stores and serve an important function in neuroplasticity. The role for RyR2 in hippocampal-dependent learning and memory is well established and chronic hyperphosphorylation of RyR2 (RyR2P) is associated with pathological calcium leakage and cognitive disorders, including Alzheimers disease. By comparison, little is known about the role of RyR2 in the ventral medial prefrontal cortex (vmPFC) circuitry important for working memory, decision making, and reward seeking. Here, we evaluated the basal expression and localization of RyR2 and RyR2P in the vmPFC. Next, we employed an operant model of sucrose, cocaine, or morphine self-administration (SA) followed by a (reward-free) recall test, to reengage vmPFC neurons and reactivate reward-seeking and re-evaluated the expression and localization of RyR2 and RyR2P in vmPFC. Under basal conditions, RyR2 was expressed in pyramidal cells but not regularly detected in PV/SST interneurons. On the contrary, RyR2P was rarely observed in PFC somata and was restricted to a different subcompartment of the same neuron - the apical dendrites of layer-5 pyramidal cells. Chronic SA of drug (cocaine or morphine) and nondrug (sucrose) rewards produced comparable increases in RyR2 protein expression. However, recalling either drug reward impaired the usual localization of RyR2P in dendrites and markedly increased its expression in somata immunoreactive for Fos, a marker of highly activated neurons. These effects could not be explained by chronic stress or drug withdrawal and instead appeared to require a recall experience associated with prior drug SA. In addition to showing the differential distribution of RyR2/RyR2P and affirming the general role of vmPFC in reward learning, this study provides information on the propensity of addictive drugs to redistribute RyR2P ion channels in a neuronal population engaged in drug-seeking. Hence, focusing on the early impact of addictive drugs on RyR2 function may serve as a promising approach to finding a treatment for substance use disorders.

neuroscience↗

bioRxiv2022

NPAS4 supports drug-cue associations and relapse-like behavior through regulation of the cell type-specific activation balance in the nucleus accumbens

Use of addictive substances creates powerful drug-cue associations that often trigger relapse. Drug seeking is gated in the nucleus accumbens (NAc) by competing activation of D1 dopamine receptor-expressing medium spiny neurons (D1-MSNs) that promote, and D2 dopamine receptor-expressing neurons (D2-MSNs) that oppose, drug seeking. We show here that the ensemble of neurons in the NAc that induce the neuronal activity-regulated transcription factor, Neuronal PAS Domain Protein 4 (NPAS4), is required for cocaine-context associations. In addition, NPAS4 functions within NAc D2-MSNs to govern the activation balance of NAc D1-MSNs and D2-MSNs necessary for drug-context memories and cue-induced cocaine, but not sucrose, seeking. NPAS4 regulates drug-cue associations and preponderant D1-MSN activation by influencing a program of gene expression that blocks cocaine-induced potentiation of prefrontal cortical excitatory drive onto D2-MSNs. Together our findings reveal that NPAS4 is a key player governing NAc MSN cell-type activation balance and promoting drug-cue associations and relapse vulnerability.

neuroscience↗

bioRxiv2021

Non-coding enhancer RNA regulates NPAS4 in the mPFC to control chronic stress-induced anhedonia-like behavior

Chronic stress can produce reward system deficits (i.e. anhedonia) and other common symptoms associated with depressive disorders, as well as neural circuit hypofunction in the medial prefrontal cortex (mPFC). However, the molecular mechanisms by which chronic stress promotes depressive-like behavior and hypofrontality remain unclear. We show here that the neuronal activity-regulated transcription factor, NPAS4, in the mPFC is regulated by chronic social defeat stress (CSDS), and its required in this brain region for CSDS-induced changes in sucrose preference and natural reward motivation. Interestingly, NPAS4 is not required for CSDS-induced social avoidance or anxiety-like behavior. We also find that mPFC NPAS4 is required for CSDS-induced reduction of pyramidal cell dendritic spine density, revealing a relationship between mPFC dendritic spine changes and anhedonia-like behavior, but not social avoidance behavior. Finally, transcriptomic analysis from the mPFC revealed that NPAS4 influences expression of numerous genes linked to glutamatergic synapses and ribosomal function, as well as many dysregulated genes observed in common neuropsychiatric disorders, including depression. Together our findings reveal an essential role for the activity-regulated transcription factor, NPAS4, in chronic stress-induced mPFC hypofunction and anhedonia.

neuroscience↗