Biology subjects

Gamble, M. C.

Publications and source records attributed to Gamble, M. C..

9 recordsLinked to original sources

bioRxiv2026

Xylazine Reshapes the Sleep and Respiratory Consequences of Chronic Fentanyl Exposure in a Sex-Dependent Manner

Recent shifts in the illicit fentanyl supply have led to widespread exposure to xylazine, yet the biological consequences of chronic fentanyl-xylazine co-exposure remain poorly understood. Here, we compared the behavioral and molecular effects of chronic exposures to fentanyl and xylazine both alone and in combination in male and female mice. Combined fentanyl-xylazine produced distinct, sex-dependent alterations in sleep architecture, including changes in NREMS recovery, REMS dynamics, and reduced sleep bout duration compared with fentanyl alone. The 2-adrenergic receptor antagonist yohimbine partially reversed these sleep alterations in a sex-dependent manner. Fentanyl-xylazine also exacerbated respiratory depression relative to fentanyl alone, particularly in males. In contrast, despite its prevalence in humans no skin lesions were observed. Molecular analyses revealed that fentanyl-xylazine co-administration leads to increased peripheral Fibroblast growth factor 21, a hepatokine linked to reduced opioid preference and alcohol consumption. Together, these findings demonstrate that xylazine substantially alters the physiological and neurobiological consequences of chronic fentanyl exposure in a sex-dependent manner, providing a preclinical framework for understanding the growing public health impact of fentanyl-xylazine co-use.

neuroscience↗

bioRxiv2025

Sex-specific transcriptional signatures of oxycodone persist during withdrawal and abstinence in the suprachiasmatic nucleus of heterogeneous stock rats

Opioid use disorder (OUD) is a major public health issue. Sleep and circadian disruptions are recognized as hallmarks of opioid addiction, often emerging during withdrawal and lasting into abstinence. However, little is known about the impact of opioids on the brains primary circadian pacemaker, the suprachiasmatic nucleus (SCN). We examined SCN transcriptomic changes in genetically diverse heterogeneous stock rats across different opioid physiological and behavioral states (naive, oxycodone intoxication, acute withdrawal, and prolonged abstinence), alongside behavioral assessments. In females, intoxication and withdrawal altered pathways related to neurotransmission, circadian rhythms, and inflammation, while in males, changes involved immune regulation, DNA damage, and metabolism. During abstinence, females showed enrichment in stress-related pathways, particularly those involved in energy metabolism and neurotransmitter function, whereas males exhibited enrichment in pathways related to cellular detoxification and oxidative stress, suggesting lasting, sex-specific effects of oxycodone administration during withdrawal and abstinence. Further, the highest proportion of sex-specific rhythmic differentially expressed genes (DEGs) were identified during abstinence compared to other states, suggesting sex differences in gene expression in the SCN during opioid abstinence. Co-expression network analysis identified a black module linked to synaptic signaling and a red module linked to ciliary function, which were positively and negatively associated with intoxication, respectively. Black module genes were positively correlated with addiction-related behaviors during abstinence, while red module genes inversely correlated with these behaviors during intoxication, linking opioid-induced alterations in the SCN to addiction-like phenotypes. These findings highlight the SCN as a dynamic, sex-specific target of opioid exposure and suggests that SCN alterations may contribute to long-term behavioral and physiological consequences of OUD. HighlightsO_LIDistinct sex specific SCN gene patterns across opioid physiological and behavior C_LIO_LIIntoxication in females increased synaptic, glutamatergic, and addiction pathways C_LIO_LICircadian entrainment pathway enriched in females after intoxication C_LIO_LIRhythmic DE genes may drive sex differences in abstinence C_LIO_LISCN gene expression correlated with addiction-like behaviors C_LI

neuroscience↗

bioRxiv2024

Sleep and circadian rhythm activity alterations during adolescence in a mouse model of neonatal fentanyl withdrawal syndrome

PurposeFentanyl, a highly potent synthetic opioid, is a major contributor to the ongoing opioid epidemic. During adulthood, fentanyl is known to induce pronounced sleep and circadian disturbances during use and withdrawal. Children exposed to opioids in utero are likely to develop neonatal opioid withdrawal syndrome, and display sleep disturbances after birth. However, it is currently unknown how neonatal opioid withdrawal from fentanyl impacts sleep and circadian rhythms in mice later in life. MethodsTo model neonatal opioid withdrawal syndrome, mice were treated with fentanyl from postnatal days 1 through 14, analogous to the third trimester of human gestation. After weaning, fentanyl and saline treated mice underwent non-invasive sleep and circadian rhythm monitoring during adolescence postnatal days 23 through 30. ResultsNeonatal fentanyl exposure led to reduced duration of wake and a decrease in the number of bouts of non-rapid eye movement sleep. Further, neonatally exposed mice displayed an increase in the average duration of rapid eye movement sleep bouts, reflecting an overall increase in the percent time spent in rapid eye movement sleep across days. ConclusionsNeonatal fentanyl exposure leads to altered sleep-wake states during adolescence in mice.

neuroscience↗

bioRxiv2024

SleepInvestigatoR: A flexible R function for analyzing scored sleep in rodents

Analyzing scored sleep is a fundamental prerequisite to understanding how sleep changes between health and disease. Classically, this is accomplished by manually calculating various measures (e.g., percent of non-rapid eye movement sleep) from a collection of scored sleep files. This process can be tedious and error prone especially when studies include a large number of animals or involve long recording sessions. To address this issue, we present SleepInvestigatoR, a versatile tool that can quickly organize and analyze multiple scored sleep files into a single output. The function is written in the open-source statistical language R and has a total of 25 parameters that can be set to match a wide variety of experimenter needs. SleepInvestigatoR delivers a total of 22 unique measures of sleep, including all measures commonly reported in the rodent literature. A simple plotting function is also provided to quickly graph and visualize the scored data. All code is designed to be implemented with little formal coding knowledge and step-by-step instructions are provided on the corresponding GitHub page. Overall, SleepInvestigatoR provides the sleep researcher a critical tool to increase efficiency, interpretation, and reproducibility in analyzing scored rodent sleep.

neuroscience↗

bioRxiv2023

Sex-specific Effects of the Endocannabinoid Agonist 2-Arachidonoylglycerol on Sleep and Circadian Disruptions during Fentanyl Withdrawal

Fentanyl has become the leading driver of opioid overdoses. Cessation of opioid use represents a challenge as the experience of withdrawal drives subsequent relapse. One of the most prominent withdrawal symptoms that can contribute to opioid craving and vulnerability to relapse is sleep disruption. The endocannabinoid agonist, 2-Arachidonoylglycerol (2-AG), may promote sleep and reduce withdrawal severity; however, the effects of 2-AG on sleep disruption during opioid withdrawal have yet to be assessed. Here, we investigate the effects of 2-AG administration on sleep-wake behavior and diurnal activity in mice during withdrawal from fentanyl. Sleep-wake activity was continuously recorded before and after chronic fentanyl administration in both male and female C57BL/6J mice. Immediately following cessation of fentanyl administration, 2-AG was administered intraperitoneally to investigate the impact of endocannabinoid agonism on opioid-induced sleep disruption. Female mice maintained higher activity levels in response to chronic fentanyl than male mice. Furthermore, fentanyl increased wake and decreased sleep during the light period and inversely increased sleep and decreased wake in the dark period in both sexes. 2-AG treatment increased arousal and decreased sleep in both sexes during first 24 hrs of withdrawal. On withdrawal day 2, only female showed increased wakefulness with no changes in males, but by withdrawal day 3 male mice displayed decreased rapid-eye movement sleep during the dark period with no changes in female mice. Overall, repeated administration of fentanyl altered sleep and diurnal activity and administration of the endocannabinoid agonist, 2-AG, had sex-specific effects on fentanyl-induced sleep and diurnal changes.

neuroscience↗

bioRxiv2023

Three-dimensional characterization of medium spiny neuron heterogeneity in the adult mouse striatum

Striatal projection neurons (SPNs) are traditionally segregated into two subpopulations expressing dopamine (DA) D1-like or D2-like receptors. However, this dichotomy is challenged by recent evidence. Functional and expression studies raise important questions: do SPNs co-express different DA receptors, and do these differences reflect unique striatal spatial distributions and expression profiles? Using RNAscope in mouse striatum, we report heterogenous SPN subpopulations distributed across dorsal-ventral and rostral-caudal axes. SPN subpopulations co-express multiple DA receptors, including D1 and D2 (D1/2R) and D1 and D3. Our integrative approach using single-nuclei multi-omics analyses provides a simple consensus to describe SPNs across diverse datasets, connecting it to complementary spatial mapping. Combining RNAscope and multi-omics shows D1/2R SPNs further separate into distinct subtypes according to spatial organization and conserved marker genes. Each SPN cell type contributes uniquely to genetic risk for neuropsychiatric diseases. Our results bridge anatomy and transcriptomics to offer new understandings of striatal neuron heterogeneity.

neuroscience↗

bioRxiv2022

A role for the circadian transcription factor NPAS2 in the progressive loss of non-rapid eye movement sleep and increased arousal during fentanyl withdrawal in male mice

AbstractO_ST_ABSRationaleC_ST_ABSSynthetic opioids like fentanyl are contributing to the rise in rates of opioid use disorder and drug overdose deaths. Sleep dysfunction and circadian rhythm disruption may worsen during opioid withdrawal and persist during abstinence. Severe and persistent sleep and circadian alterations are putative factors in opioid craving and relapse. However, very little is known about the impact of fentanyl on sleep architecture and sleep-wake cycles, particularly opioid withdrawal. Further, circadian rhythms regulate sleep-wake cycles, and the circadian transcription factor, neuronal PAS domain 2 (NPAS2) is involved in the modulation of sleep architecture and drug reward. Here, we investigate the role of NPAS2 in fentanyl-induced sleep alterations. ObjectivesTo determine the effect of fentanyl administration and withdrawal on sleep architecture, and the role of NPAS2 as a factor in fentanyl-induced sleep changes. MethodsElectroencephalography (EEG) and electromyography (EMG) was used to measure non-rapid eye movement sleep (NREMS) and rapid eye movement sleep (REMS) at baseline and following acute and chronic fentanyl administration in wild-type and NPAS2-deficient male mice. ResultsAcute and chronic administration of fentanyl led to increased wake and arousal in both wild-type and NPAS2-deficient mice, an effect that was more pronounced in NPAS2-deficient mice. Chronic fentanyl administration led to decreased NREMS, which persisted during withdrawal, progressively decreasing from day 1 to 4 of withdrawal. The impact of fentanyl on NREMS and arousal was more pronounced in NPAS2-deficient mice. ConclusionsChronic fentanyl disrupts NREMS, leading to a progressive loss of NREMS during subsequent days of withdrawal. Loss of NPAS2 exacerbates the impact of fentanyl on sleep and wake, revealing a potential role for the circadian transcription factor in opioid-induced sleep changes.

neuroscience↗

bioRxiv2020

Characterization of basal forebrain glutamate neurons suggests a role in control of arousal and avoidance behavior

The basal forebrain (BF) is involved in arousal, attention, and reward processing but the role of individual BF subtypes is still being uncovered. Glutamatergic neurons are the least well-understood of the three major BF neurotransmitter phenotypes. Here we analyzed the distribution, size, calcium-binding protein content and projections of the major group of BF glutamate neurons expressing the vesicular glutamate transporter subtype 2 (vGluT2) and tested the functional effect of activating them. Mice expressing Cre recombinase under control of the vGluT2 promoter were crossed with a reporter strain expressing the red fluorescent protein, tdTomato, to generate vGluT2-cre-tdTomato mice. Immunohistochemical staining for choline acetyltransferase and a cross with mice expressing green fluorescent protein selectively in GABAergic neurons confirmed cholinergic, GABAergic and vGluT2+ neurons represent separate BF subpopulations. Subsets of BF vGluT2+ neurons expressed the calcium binding proteins calbindin or calretinin, suggesting that multiple subtypes of BF vGluT2+ neurons exist. Anterograde tracing using adeno-associated viral vectors expressing channelrhodopsin2- enhanced yellow fluorescent fusion proteins revealed major projections of BF vGluT2+ neurons to neighboring BF cholinergic and parvalbumin neurons, as well as to extra-BF areas involved in the control of arousal and regions responding to aversive or rewarding stimuli such as the lateral habenula and ventral tegmental area. Optogenetic activation of BF vGluT2 neurons in a place preference paradigm elicited a striking avoidance of the area where stimulation was given. Together with previous optogenetic findings suggesting an arousal-promoting role, our findings suggest BF vGluT2 neurons play a dual role in promoting wakefulness and avoidance behavior.

neuroscience↗

bioRxiv2019

Basal forebrain parvalbumin neurons mediate arousals from sleep induced by hypercarbia or auditory stimuli

Brief arousals from sleep in patients with sleep apnea and other disorders prevent restful sleep, and contribute to cognitive, metabolic and physiologic dysfunction. Little is currently known about which neural systems mediate these brief arousals, hindering the development of treatments. The basal forebrain (BF) receives inputs from many nuclei of the ascending arousal system. These inputs include the brainstem parabrachial neurons which promote arousal in response to elevated blood carbon dioxide levels, as seen in sleep apnea. Optical inhibition of the terminals of parabrachial neurons in the BF impairs cortical arousals to hypercarbia, but which cell types within the BF mediate cortical arousals in response to hypercarbia or other sensory stimuli is unknown. Here using optogenetic techniques in mice, we show that BF parvalbumin (PV) neurons fulfill several criteria for a system mediating brief arousals from sleep. Optical stimulation of BF PV neurons during the light period, when mice normally sleep, caused rapid transitions to wakefulness and increased wake bout durations. Unlike many other ascending arousal systems, arousals induced by stimulation of BF PV neurons were brief, resulting in only a small (13.6%) increase in the total amount of wakefulness. Bilateral optical inhibition of BF PV neurons increased the latency to arousal produced by hypercarbia or auditory stimuli. Thus, BF PV neurons are an important component of the brain circuitry which generates brief arousals from sleep in response to internal and external sensory stimuli.

neuroscience↗