Biology subjects

Ruan, Q. T.

Publications and source records attributed to Ruan, Q. T..

8 recordsLinked to original sources

bioRxiv2021

The methamphetamine-induced RNA targetome of hnRNP H in Hnrnph1 mutants showing reduced dopamine release and behavior

Methamphetamine addiction remains a major public health concern in the United States that has paralleled the opioid epidemic. Psychostimulant use disorders have a heritable genetic component that remains unexplained. Methamphetamine targets membrane and vesicular transporters to increase synaptic dopamine, norepinephrine, and serotonin. We previously identified Hnrnph1 (heterogeneous nuclear ribonucleoprotein H1) as a quantitative trait gene underlying methamphetamine behavioral sensitivity. Hnrnph1 encodes the RNA-binding protein hnRNP H1 that is ubiquitously expressed in neurons throughout the brain. Gene-edited mice with a heterozygous frameshift deletion in Hnrnph1s first coding exon of showed reduced methamphetamine-induced dopamine release and behaviors. To inform the mechanism linking hnRNP H with methamphetamine neurobehavioral effects, we surveyed the mRNA targetome of hnRNP H via cross-linking immunoprecipitation coupled with RNA-sequencing in striatal tissue at baseline and at 30 min post-methamphetamine in wild-type male and female C57BL/6J mice. Methamphetamine induced changes in RNA-binding targets of hnRNP H in mice, including differential binding to 3UTR targets and multiple enriched mRNAs involved in synaptic plasticity. Targetome, transcriptome, and spliceome analyses triangulated on Cacna2d2 as a suggestive target, with differences in hnRNP H binding, gene expression and splicing following methamphetamine treatment (2 mg/kg, i.p.). Furthermore, pre-treatment with pregabalin, an inhibitor of 2{delta}2 and 2{delta}1 voltage-gated calcium channel subunits, attenuated methamphetamine-induced locomotor activity in male and female mice, supporting a role for Cacna2d1/d2 in methamphetamine locomotor stimulant sensitivity. Our study identifies a dynamic hnRNP H RNA targetome that can rapidly and adaptively respond to methamphetamine to regulate gene expression and likely synaptic plasticity and behavior.

neuroscience↗

bioRxiv2021

Sex differences in behavioral and brainstem transcriptomic neuroadaptations following neonatal opioid exposure in outbred mice

The opioid epidemic led to an increase in the number of Neonatal Opioid Withdrawal Syndrome (NOWS) cases in infants born to opioid-dependent mothers. Hallmark features of NOWS include weight loss, severe irritability, respiratory problems, and sleep fragmentation. Mouse models provide an opportunity to identify brain mechanisms that contribute to NOWS. Neonatal outbred Swiss Webster Cartworth Farms White (CFW) mice were administered morphine (15mg/kg, s.c.) twice daily for postnatal days (P) 1-14, an approximate of the third trimester of human gestation. Male and female mice underwent behavioral testing on P7 and P14 to determine the impact of opioid exposure on anxiety and pain sensitivity. Ultrasonic vocalizations (USVs) and daily body weights were also recorded. Brainstems containing pons and medulla were collected during morphine withdrawal on P14 for RNA-sequencing. Morphine induced weight loss from P2-14, which persisted during adolescence (P21) and adulthood (P50). USVs markedly increased at P7 in females, emerging earlier than males. On P7 and P14, both morphine exposed female and male mice displayed hyperalgesia on the hot plate and tail flick assays, with females having greater hyperalgesia than males. Morphine-exposed mice exhibited increased anxiety-like behavior in the open-field arena at P21. Transcriptome analysis of the brainstem (medulla plus pons), an area implicated in opioid withdrawal and NOWS, identified pathways enriched for noradrenergic signaling in females and males. We also found sex-specific pathways related to mitochondrial function and neurodevelopment in females and circadian entrainment in males. Sex-specific transcriptomic neuroadaptations implicate unique neurobiological mechanisms underlying NOWS-like behaviors. SIGNIFICANCE STATEMENTNeonatal opioid withdrawal syndrome (NOWS) is a poorly understood condition that has both a genetic and environmental component and is thought to be mechanistically distinct from opioid withdrawal in adults. The development of murine models for measuring neurobehavioral responses is critical for informing the neurobiological adaptations underlying NOWS. Using outbred mice that more closely model human genetic variation, we discovered a surprising degree of sexual dimorphism in behavioral timing and severity of NOWS-model behaviors as well as transcriptomic adaptations in brain tissue that together suggest distinct mechanisms and sex-specific therapeutics for reversing withdrawal symptoms and restoring brain function.

neuroscience↗

bioRxiv2020

Fentanyl-induced antinociception, reward, reinforcement, and withdrawal in Hnrnph1 mutant mice

Opioid Use Disorder (OUD) and opioid-related deaths remain a major public health concern in the United States. Both environmental and genetic factors influence risk for OUD. We previously identified Hnrnph1 as a quantitative trait gene underlying the stimulant, rewarding, and reinforcing properties of methamphetamine. Prior work demonstrates that hnRNP H1, the RNA-binding protein encoded by Hnrnph1, post-transcriptionally regulates Oprm1 (mu opioid receptor gene) - the primary molecular target for the therapeutic and addictive properties of opioids. Because genetic variants can exert pleiotropic effects on behaviors induced by multiple drugs of abuse, in the current study, we tested the hypothesis that Hnrnph1 mutants would show reduced behavioral sensitivity to the mu opioid receptor agonist fentanyl. Hnrnph1 mutants showed reduced sensitivity to fentanyl-induced locomotor activity, along with a female-specific reduction in, and a male-specific induction of, locomotor sensitization following three, daily injections (0.2 mg/kg, i.p.). Hnrnph1 mutants also required a higher dose of fentanyl to exhibit opioid reward as measured via conditioned place preference. Male Hnrnph1 mutants showed reduced fentanyl reinforcement. Hnrnph1 mutants also showed reduced sucrose motivation, suggesting a reward deficit. No genotypic differences were observed in baseline thermal nociception, fentanyl-induced antinociception, physical or negative affective signs of opioid dependence, or in sensorimotor gating. In the context of our prior work, these findings suggest that Hnrnph1 dysfunction exerts a selective role in reducing the addiction liability to drugs of abuse (opioids and psychostimulants), which could provide new biological pathways to improve their therapeutic profiles.

neuroscience↗

bioRxiv2020

Functional variants within the 5UTR support reduced exon usage as a molecular mechanism underlying the quantitative trait gene Hnrnph1 for reduced methamphetamine sensitivity

We previously identified a 210 kb region on chromosome 11 (50.37-50.58 Mb, mm10) containing two protein-coding genes (Hnrnph1, Rufy1) that was necessary for reduced methamphetamine-induced locomotor activity in C57BL/6J congenic mice harboring DBA/2J polymorphisms. Gene editing of a small deletion in the first coding exon supported Hnrnph1 as a quantitative trait gene. We have since shown that Hnrnph1 mutants also exhibit reduced methamphetamine-induced reward, reinforcement, and dopamine release. However, the quantitative trait variants (QTVs) that modulate Hnrnph1 function at the molecular level are not known. Nine single nucleotide polymorphisms and seven indels distinguish C57BL/6J from DBA/2J within Hnrnph1, including four variants within the 5 untranslated region (UTR). Here, we show that a 114 kb introgressed region containing Hnrnph1 and Rufy1 was sufficient to cause a decrease in MA-induced locomotor activity. Gene-level transcriptome analysis of striatal tissue from 114 kb congenics versus Hnrnph1 mutants identified a nearly perfect correlation of fold-change in expression for those differentially expressed genes that were common to both mouse lines, indicating functionally similar effects on the transcriptome and behavior. Exon-level analysis (including noncoding exons) revealed decreased 5 UTR usage of Hnrnph1 and immunoblot analysis identified a corresponding decrease in hnRNP H protein in 114 kb congenic mice. Molecular cloning of the Hnrnph1 5 UTR containing all four variants (but none of them individually) upstream of a reporter induced a decrease in reporter signal in both HEK293 and N2a cells, thus identifying a set of QTVs underlying molecular regulation of Hnrnph1.

neuroscience↗

bioRxiv2019

A mutation in Hnrnph1 that decreases methamphetamine-induced reinforcement, reward, and dopamine release and increases synaptosomal hnRNP H and mitochondrial proteins

Individual variation in the addiction liability of amphetamines has a heritable genetic component. We previously identified Hnrnph1 (heterogeneous nuclear ribonucleoprotein H1) as a quantitative trait gene underlying decreased methamphetamine-induced locomotor activity in mice. Here, mice (both male and female) with a heterozygous mutation in the first coding exon of Hnrnph1 (H1+/-) showed reduced methamphetamine reinforcement and intake and dose-dependent changes in methamphetamine reward as measured via conditioned place preference. Furthermore, H1+/- mice showed a robust decrease in methamphetamine-induced dopamine release in the nucleus accumbens with no change in baseline extracellular dopamine, striatal whole tissue dopamine, dopamine transporter protein, or dopamine uptake. Immunohistochemical and immunoblot staining of midbrain dopaminergic neurons and their forebrain projections for tyrosine hydroxylase did not reveal any major changes in staining intensity, cell number, or in the number of forebrain puncta. Surprisingly, there was a two-fold increase in hnRNP H protein in the striatal synaptosome of H1+/- mice with no change in whole tissue levels. To gain insight into the molecular mechanisms linking increased synaptic hnRNP H with decreased methamphetamine-induced dopamine release and behaviors, synaptosomal proteomic analysis identified an increased baseline abundance of several mitochondrial complex I and V proteins that rapidly decreased at 30 min post-methamphetamine administration in H1+/- mice. In contrast, the much lower level of basal synaptosomal mitochondrial proteins in wild-type mice showed a rapid increase in response to methamphetamine. We conclude that H1+/- decreases methamphetamine-induced dopamine release, reward, and reinforcement and induces dynamic changes in basal and methamphetamine-induced synaptic mitochondrial function.\n\nSIGNIFICANCE STATEMENTMethamphetamine dependence is a significant public health concern with no FDA-approved treatment. We discovered a role for the RNA binding protein hnRNP H in methamphetamine reward and reinforcement. Hnrnph1 mutation also blunted methamphetamine-induced dopamine release in the nucleus accumbens - a key neurochemical event contributing to methamphetamine addiction liability. Finally, Hnrnph1 mutants showed a marked increase in basal level of synaptosomal hnRNP H and mitochondrial proteins that decreased in response to methamphetamine whereas wild-type mice showed a methamphetamine-induced increase in synaptosomal mitochondrial proteins. Thus, we identified a potential role for hnRNP H in basal and dynamic mitochondrial function that informs methamphetamine-induced cellular adaptations associated with reduced addiction liability.

neuroscience↗

bioRxiv2018

Changes in neuronal immunofluorescence in the C- versus N-terminal domains of hnRNP H following D1 dopamine receptor activation

RNA binding proteins are a diverse class of proteins that regulate all aspects of RNA metabolism. Accumulating studies indicate that heterogeneous nuclear ribonucleoproteins are associated with cellular adaptations in response to drugs of abuse. We recently mapped and validated heterogeneous nuclear ribonucleoprotein H1 (Hnrnph1) as a quantitative trait gene underlying differential behavioral sensitivity to methamphetamine. The molecular mechanisms by which hnRNP H1 alters methamphetamine behaviors are unknown but could involve preand/or post-synaptic changes in protein localization and function. Methamphetamine initiates post-synaptic D1 dopamine receptor signaling indirectly by binding to pre-synaptic dopamine transporters and vesicular monoamine transporters of midbrain dopaminergic neurons which triggers revers e transport and accumulation of dopamine at the synapse. Here, we examined changes in neuronal localization of hnRNP H in primary rat cortical neurons that express dopamine receptors that can be modulated by the D1 or D2 dopamine receptor agonists SKF38393 and (-)-Quinpirole HCl, respectively. Basal immunostaining of hnRNP H was localized primarily to the nucleus. D1 dopamine receptor activation induced an increase in hnRNP H nuclear immunostaining as detected by immunocytochemistry with a C-domain directed antibody containing epitope near the glycine-rich domain but not with an N-domain specific antibody. Although there was no change in hnRNP H protein in the nucleus or cytoplasm, there was a decrease in Hnrnph1 transcript following D1 receptor stimulation. Taken together, these results suggest that D1 receptor activation increases availability of the hnRNP H C-terminal epitope, which could potentially reflect changes in protein-protein interactions. Thus, D1 receptor signaling could represent a key molecular post-synaptic event linking Hnrnph1 polymorphisms to drug-induced behavior.\n\nHighlightsO_LIWe investigated immunofluorescence and localization of hnRNP H following dopamine receptor activation in primary rat cortical neurons.\nC_LIO_LIActivation of D1 dopamine receptor increased nuclear immunofluorescence of the C-terminal domain but not the N-terminal domain of hnRNP H with no change in either nuclear or cytoplasmic levels of hnRNP H protein.\nC_LIO_LID1 dopamine signaling could alter interaction of hnRNP H with other proteins.\nC_LI

neuroscience↗

bioRxiv2018

Cyfip1 haploinsufficiency increases compulsive-like behavior and paternally inherited palatable food intake: Implications for Prader-Willi Syndrome

Binge eating (BE) is a heritable trait associated with eating disorders and involves rapid consumption of large quantities of food. We identified cytoplasmic FMRP-interacting protein 2 (Cyfip2) as a major genetic factor underlying BE and concomitant compulsive-like behaviors in mice. CYFIP2 is a gene homolog of CYFIP1 - one of four paternally-deleted genes in patients with the more severe Type I Prader-Willi Syndrome (PWS). PWS is a neurodevelopmental disorder where 70% of cases involve paternal deletion of 15q11-q13. PWS symptoms include hyperphagia, obesity (if untreated), cognitive deficits, and obsessive-compulsive behaviors. We tested whether Cyfip1 haploinsufficiency (+/-) would enhance premorbid compulsive-like behavior and palatable food (PF) intake in a parent-of-origin-selective manner. We tested Cyfip1+/- mice on a C57BL/6N (N) background that were homozygous for the BE-associated missense mutation in Cyfip2 (S968F) as well as mice that we backcrossed to homozygosity for the C57BL/6J (J) allele at Cyfip2 (Cyfip2J/J). Cyfip1+/- mice showed increased compulsive-like behavior on both backgrounds, increased PF consumption on the Cyfip2N/N background in a paternally-enhanced manner, and decreased PF consumption in male Cyfip1+/- mice on the Cyfip2J/J background in a maternally selective manner. In the hypothalamus, there was a maternally-enhanced reduction of Cyfip1 transcription, but a paternally-enhanced reduction in CYFIP1 protein. In the nucleus accumbens, there was a maternally-enhanced reduction in CYFIP1 protein. Together, increased compulsive-like behavior, parent-of-origin-, and genetic background-dependent effects of Cyfip1 haploinsufficiency on PF consumption implicate CYFIP1 in behaviors in neurodevelopmental disorders involving reduced expression of CYFIP1, including PWS, Fragile X Syndrome, and 15q11.2 Microdeletion Syndrome.

genetics↗

bioRxiv2017

Genetic differences in the behavioral organization of binge eating, conditioned food reward, and compulsive-like eating in C57BL/6J and DBA/2J strains

Binge eating (BE) is a heritable symptom of eating disorders associated with anxiety, depression, malnutrition, and obesity. Genetic analysis of BE could facilitate therapeutic discovery. We used an intermittent, limited access BE paradigm involving sweetened palatable food (PF) to examine genetic differences in BE, conditioned food reward, and compulsive-like eating between C57BL/6J (B6J) and DBA/2J (D2J) inbred mouse strains. D2J mice showed a robust escalation in intake and conditioned place preference for the PF-paired side. D2J mice also showed a unique style of compulsive-like eating in the light/dark conflict test where they rapidly hoarded and consumed PF in the preferred unlit environment. BE and compulsive-like eating exhibited narrow-sense heritability estimates between 56 and 73 percent. To gain insight into the genetic basis, we phenotyped and genotyped a small cohort of 133 B6J x D2J-F2 mice at the peak location of three quantitative trait loci (QTL) previously identified in F2 mice for sweet taste (chromosome 4: 156 Mb), bitter taste (chromosome 6: 133 Mb) and behavioral sensitivity to drugs of abuse (chromosome 11: 50 Mb). The D2J allele on chromosome 6 was associated with greater PF intake on training days and greater compulsive-like PF intake, but only in males, suggesting that decreased bitter taste may increase BE in males. The D2J allele on chromosome 11 was associated with an increase in final PF intake and slope of escalation across days. Future studies employing larger crosses and genetic reference panels comprising B6J and D2J alleles will identify causal genes and neurobiological mechanisms.

animal behavior and cognition↗