Biology subjects

Fox, M. E.

Publications and source records attributed to Fox, M. E..

11 recordsLinked to original sources

bioRxiv2026

Projection-defined ventral tegmental area neurons exhibit distinct molecular adaptations during acute fentanyl withdrawal

Opioid use disorder (OUD) remains a major public health concern, with potent synthetic opioids driving overdose deaths. Understanding how synthetic opioids like fentanyl alter brain circuits to promote persistent use is critical for advancing OUD treatment. The ventral tegmental area (VTA), which projects to several other brain regions including prefrontal cortex (PFC), nucleus accumbens (NAc) and amygdala (Amy), contributes to both the euphoric and dysphoric phases of OUD. Projection target influences the baseline properties and opioid-induced neuroadaptations of VTA neurons, yet less is known about the molecular adaptations that occur across distinct VTA projections during spontaneous opioid withdrawal. Here, we investigated transcriptional responses to acute spontaneous fentanyl withdrawal in projection-defined VTA neurons. Following chronic fentanyl exposure and 24h of spontaneous withdrawal in male and female mice, we sequenced ribosome-associated RNA from VTA neurons projecting to PFC, NAc, or Amy. We found that fentanyl withdrawal produced divergent transcriptional adaptations in VTA neurons projecting to PFC compared with VTA-NAc and VTA-Amy neurons, whereas adaptations in VTA-NAc and VTA-Amy neurons were more concordant. Weighted gene co-expression network analysis (WGCNA) identified fentanyl-responsive modules across VTA projections, with gene ontology analysis linking these modules to distinct biological processes. We further used regulatory enrichment analysis to identify candidate transcriptional regulators associated with fentanyl-responsive co-expression modules. Together, our data demonstrate that VTA neurons undergo distinct molecular adaptations during fentanyl withdrawal based on their projection target, providing new insight into molecular mechanisms relevant to OUD.

neuroscience↗

bioRxiv2025

Projection-defined ventral tegmental area neurons exhibit distinct fentanyl-induced molecular and functional adaptations that differentially support drug-context associations

RationaleSynthetic opioids like fentanyl are contributing to unprecedented overdose rates, yet the neural circuitry underlying fentanyl-associated behaviors remains poorly understood. The ventral tegmental area (VTA) projects to both the nucleus accumbens (NAc) and prefrontal cortex (PFC), forming distinct pathways that are implicated in drug-cue associations, though their specific roles in fentanyl-context encoding are not well defined. ObjectivesThis study aimed to determine how VTA-NAc and VTA-PFC circuits contribute to fentanyl-context associations, and to assess the role of downstream dopamine receptor signaling in fentanyl context-seeking. MethodsMale and female mice underwent fentanyl conditioned place preference (CPP; 0.2 mg/kg). We locally inhibited dopamine D1 or D2 receptors in NAc or PFC during CPP expression. We used fiber photometry calcium imaging to measure activity in VTA-NAc and VTA-PFC projection neurons, and chemogenetic inhibition to suppress activity during CPP expression. ResultsFentanyl CPP expression was attenuated by blocking D1 but not D2 receptors in PFC, and D2 but not D1 receptors in NAc. We found both VTA-NAc and VTA-PFC exhibited increased calcium activity during fentanyl exposure and during entries to the fentanyl-paired context. We further identified a functional role for VTA-NAc, as chemogenetic inhibition of VTA-NAc, but not VTA-PFC, reduced fentanyl context-seeking. ConclusionsWhile both VTA-NAc and VTA-PFC pathways are recruited by fentanyl exposure, fentanyl context-seeking relies on different downstream dopamine receptors in NAc vs PFC. Further, activity in VTA-NAc functionally supports the expression of fentanyl CPP. Together, these findings indicate that VTA circuits differentially contribute to fentanyl context-seeking.

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bioRxiv2025

A Drd1-cre mouse line with nucleus accumbens gene dysregulation exhibits blunted fentanyl seeking

The synthetic opioid fentanyl remains abundant in the illicit drug supply, contributing to tens of thousands of overdose deaths every year. Despite this, the neurobiological effects of fentanyl use remain largely understudied. The nucleus accumbens (NAc) is a central locus promoting persistent drug use and relapse, largely dependent on activity of dopamine D1 receptors. NAc D1 receptor-expressing medium spiny neurons (D1-MSNs) undergo molecular and physiological adaptations that contribute to negative affect during fentanyl abstinence, but whether these neuroadaptations also promote fentanyl relapse is unclear. Here, we obtained Drd1-cre120Mxu mice to investigate D1-dependent mechanisms of fentanyl relapse. We serendipitously discovered this mouse line is resistant to fentanyl seeking, despite similar intravenous fentanyl self-administration, and greater fentanyl-induced locomotion, compared to wildtype counterparts. In drug naive mice, we found Drd1-cre120Mxu mice have elevated D1 receptor expression in NAc, alongside increased expression of MSN marker genes Chrm4 and Penk. We show Drd1-cre120Mxu mice have increased sensitivity to the D1 receptor agonist SKF-38393, and exhibit divergent expression of MSN markers, opioid receptors, glutamate receptor subunits, and TrkB after fentanyl self-administration that may underly blunted fentanyl seeking. Finally, we show fentanyl-related behavior is unaltered by chemogenetic manipulation of D1-MSNs in Drd1-cre120Mxu mice. Conversely, chemogenetic stimulation of putative D1-MSNs in wildtype mice recapitulated the blunted fentanyl seeking of Drd1-cre120Mxu mice, supporting a role for aberrant D1-MSN signaling in this behavior. Together, our data uncover alterations in NAc gene expression and function with implications for susceptibility and resistance to developing fentanyl use disorder.

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bioRxiv2023

Transcriptional signatures of fentanyl use in the mouse ventral tegmental area

Synthetic opioids such as fentanyl contribute to the vast majority of opioid-related overdose deaths, but fentanyl use remains broadly understudied. Like other substances with misuse potential, opioids cause lasting molecular adaptations to brain reward circuits, including neurons in the ventral tegmental area (VTA). The VTA contains numerous cell types that play diverse roles in opioid use and relapse, however it is unknown how fentanyl experience alters the transcriptional landscape in specific subtypes. Here, we performed single nuclei RNA sequencing to study transcriptional programs in fentanyl experienced mice. Male and female C57/BL6 mice self-administered intravenous fentanyl (1.5 {micro}g/kg/infusion) or saline for 10 days. After 24 hr abstinence, VTA nuclei were isolated and prepared for sequencing on the 10X platform. We identified different patterns of gene expression across cell types. In dopamine neurons, we found enrichment of genes involved in growth hormone signaling. In dopamine-glutamate-GABA combinatorial neurons, and some GABA neurons, we found enrichment of genes involved in Pi3k-Akt signaling. In glutamate neurons, we found enrichment of genes involved in cholinergic signaling. We identified transcriptional regulators for the differentially expressed genes in each neuron cluster, including downregulation of transcriptional repressor Bcl6, and upregulation of Wnt signaling partner Tcf4. We also compared the fentanyl-induced gene expression changes identified in mouse VTA with a published rat dataset in bulk VTA, and found overlap in genes related to GABAergic signaling and extracellular matrix interaction. Together, we provide a comprehensive picture of how fentanyl self-administration alters the transcriptional landscape of the mouse VTA, that serves for the foundation for future mechanistic studies.

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bioRxiv2022

Sex-specific transcriptomic profiling of reward and sensory brain areas in perinatal fentanyl exposed juvenile mice

Use of the synthetic opioid fentanyl increased [~]300% in the last decade, including among women of reproductive ages. Adverse neonatal outcomes and long-term behavioral disruptions are associated with perinatal opioid exposure. Our previous work demonstrated that perinatal fentanyl exposed mice displayed enhanced negative affect and somatosensory circuit and behavioral disruptions during adolescence. However, little is known about molecular adaptations across brain regions that underlie these outcomes. We performed RNA-sequencing across three reward and two sensory brain areas to study transcriptional programs in perinatal fentanyl exposed juvenile mice. Pregnant dams received 10g/ml fentanyl in the drinking water from embryonic day 0 (E0) through gestational periods until weaning at postnatal day 21 (P21). RNA was extracted from nucleus accumbens (NAc), prelimbic cortex (PrL), ventral tegmental area (VTA), somatosensory cortex (S1) and ventrobasal thalamus (VBT) from perinatal fentanyl exposed mice of both sexes at P35. RNA-sequencing was performed, followed by analysis of differentially expressed genes (DEGs) and gene co-expression networks. Transcriptome analysis revealed DEGs and gene modules significantly associated with exposure to perinatal fentanyl in a sex-wise manner. The VTA had the most DEGs, while robust gene enrichment occurred in NAc. Genes enriched in mitochondrial respiration were pronounced in NAc and VTA of perinatal fentanyl exposed males, extracellular matrix (ECM) and neuronal migration enrichment were pronounced in NAc and VTA of perinatal fentanyl exposed males, while genes associated with vesicular cycling and synaptic signaling were markedly altered in NAc of perinatal fentanyl exposed female mice. In sensory areas from perinatal fentanyl exposed females, we found alterations in mitochondrial respiration, synaptic and ciliary organization processes. Our findings demonstrate distinct transcriptomes across reward and sensory brain regions, with some showing discordance between sexes. These transcriptome adaptations may underlie structural, functional, and behavioral changes observed in perinatal fentanyl exposed mice.

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bioRxiv2021

Activation of Npas1-neurons in the Ventral Pallidum Mediates Stress Susceptibility

AbstractAltered activity of the ventral pallidum (VP) underlies disrupted motivation in stress and drug exposure. The VP is a very heterogeneous structure comprised of many neuron types with distinct physiological properties and projections. Neuronal PAS 1-positive (Npas1+) VP neurons are thought to send projections to brain regions critical for motivational behavior. While Npas1+ neurons have been characterized in the globus pallidus external, there is limited information on these neurons in the VP. To address this limitation, we evaluated the projection targets of the VP Npas1+ neurons and performed RNA-seq on ribosome-associated mRNA from VP Npas1+ neurons to determine their molecular identity. Finally, we used a chemogenetic approach to manipulate VP Npas1+ neurons during social defeat stress (SDS) and behavioral tasks related to anxiety and motivation in Npas1-Cre mice. We employed a similar approach in females using the chronic witness defeat stress (CWDS). We identified VP Npas1+ projections to the nucleus accumbens, ventral tegmental area, medial and lateral habenula, lateral hypothalamus, thalamus, medial and lateral septum, and periaqueductal gray area. VP Npas1+ neurons displayed distinct transcriptomes representing distinct biological processes. Chemogenetic activation of VP Npas1+ neurons increased susceptibility to a subthreshold (S)SDS and anxiety-like behavior in the elevated plus maze and open field while the inhibition of VP Npas1+ neurons enhanced resilience to chronic (C)SDS and CWDS. Thus, the activity of VP Npas1+ neurons modulates susceptibility to social stressors and anxiety-like behavior. Our studies provide new information into VP Npas1+ neuron circuitry, molecular identity, and their role in stress response.

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bioRxiv2021

Transcriptome profiling of the ventral pallidum reveals a role for pallido-thalamic neurons in cocaine reward

Psychostimulant exposure alters the activity of ventral pallidum (VP) projection-neurons. However, the molecular underpinnings of these circuit dysfunctions are unclear. We used RNA- sequencing to reveal alterations in the transcriptional landscape of the VP that are induced by cocaine self-administration in mice. We then probed gene expression in select VP neuronal subpopulations to isolate a circuit associated with cocaine intake. Finally, we used both overexpression and CRISPR-mediated knockdown to test the role of a gene target on cocaine- mediated behaviors as well as dendritic spine density. Our results showed that a large proportion (55%) of genes associated with structural plasticity were changed 24 hours following cocaine intake. Among them, the transcription factor Nr4a1 (Nuclear receptor subfamily 4, group A, member 1, or Nur77) showed high expression levels. We found that the VP to mediodorsal thalamus (VP[->]MDT) projection neurons specifically were recapitulating this increase in Nr4a1 expression. Overexpressing Nr4a1 in VP[->]MDT neurons enhanced drug-seeking and drug- induced reinstatement, while Nr4a1 knock down prevented self-administration acquisition and subsequent cocaine-mediated behaviors. Moreover, we showed that Nr4a1 negatively regulated spine dynamics in this specific cell subpopulation. Together, our study identifies for the first time the transcriptional mechanisms occurring in VP in drug exposure. Our study provides further understanding on the role of Nr4a1 in cocaine-related behaviors and identifies the crucial role of the VP[->]MDT circuit in drug intake and relapse-like behaviors.

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bioRxiv2020

Divergent profiles of fentanyl withdrawal and associated pain in mice and rats

Opioid abuse has devastating effects on patients, their families, and society. Withdrawal symptoms are severely unpleasant, prolonged, and frequently hinder recovery or lead to relapse. The sharp increase in abuse and overdoses arising from the illicit use of potent and rapidly-acting synthetic opioids, such as fentanyl, highlights the urgency of understanding the withdrawal mechanisms related to these drugs. Progress is impeded by inconsistent reports on opioid withdrawal in different preclinical models. Here, using rats and mice of both sexes, we quantified withdrawal behaviors during spontaneous and naloxone-precipitated withdrawal, following two weeks of intermittent fentanyl exposure. We found that both mice and rats lost weight during exposure and showed increased signs of distress during spontaneous and naloxone precipitated withdrawal. However, these species differed in their expression of withdrawal associated pain, a key contributor to relapse in humans. Spontaneous or ongoing pain was preferentially expressed in rats in both withdrawal conditions, while no change was observed in mice. In contrast, withdrawal associated thermal hyperalgesia was found only in mice. These data suggest that rats and mice diverge in how they experience withdrawal and which aspects of the human condition they most accurately model. These differences highlight each species strengths as model systems and can inform experimental design in studies of opioid withdrawal.

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bioRxiv2020

Perinatal fentanyl exposure leads to long-lasting impairments in somatosensory circuit function and behavior

One consequence of the opioid epidemic are lasting neurodevelopmental sequelae afflicting adolescents exposed to opioids in the womb. A translationally relevant and developmentally accurate preclinical model is needed to understand the behavioral, circuit, network, and molecular abnormalities resulting from this exposure. By employing a novel preclinical model of perinatal fentanyl exposure, our data reveal that fentanyl has several dose-dependent, developmental consequences to somatosensory function and behavior. Newborn male and female mice exhibit signs of withdrawal and sensory-related deficits that extend at least to adolescence. As fentanyl exposure does not affect dams health or maternal behavior, these effects result from the direct actions of perinatal fentanyl on the pups developing brain. At adolescence, exposed mice exhibit reduced adaptation to sensory stimuli, and a corresponding impairment in primary somatosensory (S1) function. In vitro electrophysiology demonstrates a long-lasting reduction in S1 synaptic excitation, evidenced by decreases in release probability, NMDA receptor-mediated postsynaptic currents, and frequency of miniature excitatory postsynaptic currents, as well as increased frequency of miniature inhibitory postsynaptic currents. In contrast, anterior cingulate cortical neurons exhibit an opposite phenotype, with increased synaptic excitation. Consistent with these canghes, electrocorticograms reveal suppressed ketamine-evoked {gamma} oscillations. Morphological analysis of S1 pyramidal neurons indicate reduced dendritic complexity, dendritic length, and soma size. Further, exposed mice exhibited abnormal cortical mRNA expression of key receptors and neuronal growth and development, changes that were consistent with the electrophysiological and morphological changes. These findings demonstrate the lasting sequelae of perinatal fentanyl exposure on sensory processing and function.

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bioRxiv2019

Enduring consequences of perinatal fentanyl exposure in mice

Opioid use by pregnant women is an understudied consequence associated with the opioid epidemic, resulting in a rise in the incidence of neonatal opioid withdrawal syndrome (NOWS), and lifelong neurobehavioral deficits that result from perinatal opioid exposure. There are few preclinical models that accurately recapitulate human perinatal drug exposure, and few focus on fentanyl, a potent synthetic opioid that is a leading driver of the opioid epidemic. To investigate the consequences of perinatal opioid exposure, we administered fentanyl to mouse dams in their drinking water throughout gestation and until litters were weaned at postnatal day (PD) 21. Fentanyl-exposed dams delivered smaller litters, and had higher litter mortality rates compared to controls. Metrics of maternal care behavior were not affected by the treatment, nor were there differences in dams weight or liquid consumption throughout gestation and 21 days postpartum. Twenty-four hours after weaning and drug cessation, perinatal fentanyl-exposed mice exhibited signs of spontaneous somatic withdrawal behavior, and sex-specific weight fluctuations that normalized in adulthood. At adolescence (PD 35) they displayed elevated anxiety-like behaviors and decreased grooming, assayed in the elevated plus maze and sucrose splash tests. Finally, by adulthood (PD 55) they displayed impaired performance in a two-tone auditory discrimination task. Collectively, our findings suggest that perinatal fentanyl exposed mice exhibit somatic withdrawal behavior and changes across the lifespan reminiscent of humans born with NOWS.

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bioRxiv2019

Individual differences in stereotypy and neuron subtype translatome with TrkB deletion

Motor stereotypies occurring in early-onset neuropsychiatric diseases are associated with dysregulated basal ganglia direct-pathway activity. Disruptions in network connectivity through impaired neuronal structure have been implicated in both rodents and humans. However, the neurobiological mechanisms leading to direct-pathway neuron disconnectivity in stereotypy remain poorly understood. We have a mouse line with Tropomyosin receptor kinase B (TrkB) receptor deletion from D1-expressing cells (D1-Cre-flTrkB) in which a subset of animals shows repetitive rotations and head tics with juvenile onset. Here we demonstrate these behaviors may be associated with abnormal direct-pathway activity by reducing rotations using chemogenetic inhibition of dorsal striatum D1-medium spiny neurons (D1-MSNs) in both juvenile and young adult mice. Taking advantage of phenotypical differences in animals with similar genotype, we then interrogated the D1-MSN specific translatome associated with repetitive behavior by using RNA-sequencing of ribosome-associated mRNA. Detailed translatome analysis followed by multiplexed gene expression assessment revealed profound alterations in neuronal projection and synaptic structure related genes in stereotypy mice. Examination of neuronal morphology demonstrated dendritic atrophy and dendritic spine loss in dorsal striatum D1-MSNs from mice with repetitive behavior. Together, our results uncover phenotype-specific molecular alterations in D1-MSNs that relate to morphological adaptations in mice displaying stereotypy behavior.

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