J Neurosci. 2015 May 27;35(21):8232-44.
Li X, Rubio FJ, Zeric T, Bossert JM, Kambhampati S, Cates HM, Kennedy PJ, Liu QR, Cimbro R, Hope BT, Nestler EJ, Shaham Y.
PMID: 26016895 | DOI: 10.1038/jid.2015.200.
Cue-induced methamphetamine seeking progressively increases after withdrawal (incubation of methamphetamine craving), but the underlying mechanisms are largely unknown. We determined whether this incubation is associated with alterations in candidate genes in dorsal striatum (DS), a brain area implicated in cue- and context-induced drug relapse. We first measured mRNA expression of 24 candidate genes in whole DS extracts after short (2 d) or prolonged (1 month) withdrawal in rats following extended-access methamphetamine or saline (control condition) self-administration (9 h/d, 10 d). We found minimal changes. Next, using fluorescence-activated cell sorting, we compared gene expression in Fos-positive dorsal striatal neurons, which were activated during "incubated" cue-induced drug-seeking tests after prolonged withdrawal, with nonactivated Fos-negative neurons. We found significant increases in mRNA expression of immediate early genes (Arc, Egr1), Bdnf and its receptor (Trkb), glutamate receptor subunits (Gria1, Gria3, Grm1), and epigenetic enzymes (Hdac3, Hdac4, Hdac5, GLP, Dnmt3a, Kdm1a) in the Fos-positive neurons only. Using RNAscope to determine striatal subregion and cell-type specificity of the activated neurons, we measured colabeling of Fos with Drd1 and Drd2 in three DS subregions. Fos expression was neither subregion nor cell-type specific (52.5 and 39.2% of Fos expression colabeled with Drd1 and Drd2, respectively). Finally, we found that DS injections of SCH23390 (C17H18ClNO), a D1-family receptor antagonist known to block cue-induced Fos induction, decreased incubated cue-induced methamphetamine seeking after prolonged withdrawal. Results demonstrate a critical role of DS in incubation of methamphetamine craving and that this incubation is associated with selective gene-expression alterations in cue-activated D1- and D2-expressing DS neurons.
Biological Psychiatry Global Open Science
Guerri, L;Dobbs, L;da Silva e Silva, D;Meyers, A;Ge, A;Lecaj, L;Djakuduel, C;Islek, D;Hipolito, D;Martinez, A;Shen, P;Marietta, C;Garamszegi, S;Capobianco, E;Jiang, Z;Schwandt, M;Mash, D;Alvarez, V;Goldman, D;
| DOI: 10.1016/j.bpsgos.2022.08.010
Background A salient effect of addictive drugs is to hijack the dopamine reward system, an evolutionarily conserved driver of goal-directed behavior and learning. Reduced dopamine type-II receptor (D2R) availability in the striatum is an important pathophysiological mechanism for addiction that is both consequential and causal for other molecular, cellular, and neuronal network differences etiologic for this disorder. Here, we sought to identify gene expression changes attributable to innate low expression of the Drd2 gene in the striatum and specific to striatal indirect medium spiny neurons (iMSNs). Methods Cre-conditional, Translating Ribosome Affinity Purification (TRAP) was used to purify and analyze the translatome (ribosome-bound mRNA) of iMSNs from mice with low/heterozygous or wild-type Drd2 expression in iMSNs. Complementary electrophysiological recordings and gene expression analysis of postmortem brain tissue from human cocaine users were performed. Results Innate low expression of Drd2 in iMSNs led to differential expression of genes involved in GABA and cAMP signaling, neural growth, lipid metabolism, neural excitability, and inflammation. Creb1 was identified as a likely upstream regulator, among others. In human brain, expression of FXYD2, a modulatory subunit of the Na/K pump, was negatively correlated with DRD2 mRNA expression. In iMSN-TRAP-Drd2HET mice, increased Cartpt and reduced S100a10 (p11) expression recapitulated previous observations in cocaine paradigms. Electrophysiology experiments supported a higher GABA tone in iMSN-Drd2HET mice. Conclusion This study provides strong molecular evidence that in addiction inhibition by the indirect pathway is constitutively enhanced through neural growth and increased GABA signaling.
Caprioli D, Venniro M, Zhang M, Bossert JM, Warren BL, Hope BT, Shaham Y.
PMID: 28123032 | DOI: 10.1523/JNEUROSCI.3091-16.2017
Abstract
We recently developed a rat model of incubation of methamphetamine craving after choice-based voluntary abstinence. Here, we studied the role of dorsolateral striatum (DLS) and dorsomedial striatum (DMS) in this incubation. We trained rats to self-administer palatable food pellets (6 d, 6 h/d) and methamphetamine (12 d, 6 h/d). We then assessed relapse to methamphetamine seeking under extinction conditions after 1 and 21 abstinence days. Between tests, the rats underwent voluntary abstinence (using a discrete choice procedure between methamphetamine and food; 20 trials/d) for 19 d. We used in situ hybridization to measure the colabeling of the activity marker Fos with Drd1 and Drd2 in DMS and DLS after the tests. Based on the in situ hybridization colabeling results, we tested the causal role of DMS D1 and D2family receptors, and DMS neuronal ensembles in "incubated" methamphetamine seeking, using selective dopamine receptor antagonists (SCH39166 or raclopride) and the Daun02 chemogenetic inactivation procedure, respectively. Methamphetamine seeking was higher after 21 d of voluntary abstinence than after 1 d (incubation of methamphetamine craving). The incubated response was associated with increased Fos expression in DMS but not in DLS; Fos was colabeled with both Drd1 and Drd2 DMS injections of SCH39166 or raclopride selectively decreased methamphetamine seeking after 21 abstinence days. In Fos-lacZ transgenic rats, selective inactivation of relapse test-activated Fos neurons in DMS on abstinence day 18 decreased incubated methamphetamine seeking on day 21. Results demonstrate a role of DMS dopamine D1 and D2 receptors in the incubation of methamphetamine craving after voluntary abstinence and that DMS neuronal ensembles mediate this incubation.
SIGNIFICANCE STATEMENT:
In human addicts, abstinence is often self-imposed and relapse can be triggered by exposure to drug-associated cues that induce drug craving. We recently developed a rat model of incubation of methamphetamine craving after choice-based voluntary abstinence. Here, we used classical pharmacology, in situ hybridization, immunohistochemistry, and the Daun02 inactivation procedure to demonstrate a critical role of dorsomedial striatum neuronal ensembles in this new form of incubation of drug craving.
Bernanke, A;Burnette, E;Murphy, J;Hernandez, N;Zimmerman, S;Walker, QD;Wander, R;Sette, S;Reavis, Z;Francis, R;Armstrong, C;Risher, ML;Kuhn, C;
PMID: 34898621 | DOI: 10.1371/journal.pone.0260577
Females are more affected by psychiatric illnesses including eating disorders, depression, and post-traumatic stress disorder than males. However, the neural mechanisms mediating these sex differences are poorly understood. Animal models can be useful in exploring such neural mechanisms. Conditioned taste aversion (CTA) is a behavioral task that assesses how animals process the competition between associated reinforcing and aversive stimuli in subsequent task performance, a process critical to healthy behavior in many domains. The purpose of the present study was to identify sex differences in this behavior and associated neural responses. We hypothesized that females would value the rewarding stimulus (Boost ) relative to the aversive stimulus (LiCl) more than males in performing CTA. We evaluated behavior (Boost intake, LiCl-induced behaviors, ultrasonic vocalizations (USVs), CTA performance) and Fos activation in relevant brain regions after the acute stimuli [acute Boost (AB), acute LiCl (AL)] and the context-only task control (COT), Boost only task (BOT) and Boost -LiCl task (BLT). Acutely, females drank more Boost than males but showed similar aversive behaviors after LiCl. Females and males performed CTA similarly. Both sexes produced 55 kHz USVs anticipating BOT and inhibited these calls in the BLT. However, more females emitted both 22 kHz and 55 kHz USVs in the BLT than males: the latter correlated with less CTA. Estrous cycle stage also influenced 55 kHz USVs. Fos responses were similar in males and females after AB or AL. Females engaged the gustatory cortex and ventral tegmental area (VTA) more than males during the BOT and males engaged the amygdala more than females in both the BOT and BLT. Network analysis of correlated Fos responses across brain regions identified two unique networks characterizing the BOT and BLT, in both of which the VTA played a central role. In situ hybridization with RNAscope identified a population of D1-receptor expressing cells in the CeA that responded to Boost and D2 receptor-expressing cells that responded to LiCl. The present study suggests that males and females differentially process the affective valence of a stimulus to produce the same goal-directed behavior.
Matsushima, A;Pineda, SS;Crittenden, JR;Lee, H;Galani, K;Mantero, J;Tombaugh, G;Kellis, M;Heiman, M;Graybiel, AM;
PMID: 36650127 | DOI: 10.1038/s41467-022-35752-x
Striatal projection neurons (SPNs), which progressively degenerate in human patients with Huntington's disease (HD), are classified along two axes: the canonical direct-indirect pathway division and the striosome-matrix compartmentation. It is well established that the indirect-pathway SPNs are susceptible to neurodegeneration and transcriptomic disturbances, but less is known about how the striosome-matrix axis is compromised in HD in relation to the canonical axis. Here we show, using single-nucleus RNA-sequencing data from male Grade 1 HD patient post-mortem brain samples and male zQ175 and R6/2 mouse models, that the two axes are multiplexed and differentially compromised in HD. In human HD, striosomal indirect-pathway SPNs are the most depleted SPN population. In mouse HD models, the transcriptomic distinctiveness of striosome-matrix SPNs is diminished more than that of direct-indirect pathway SPNs. Furthermore, the loss of striosome-matrix distinction is more prominent within indirect-pathway SPNs. These results open the possibility that the canonical direct-indirect pathway and striosome-matrix compartments are differentially compromised in late and early stages of disease progression, respectively, differentially contributing to the symptoms, thus calling for distinct therapeutic strategies.
Downs, AM;Donsante, Y;Jinnah, HA;Hess, EJ;
PMID: 35314320 | DOI: 10.1016/j.nbd.2022.105699
Trihexyphenidyl (THP), a non-selective muscarinic receptor (mAChR) antagonist, is commonly used for the treatment of dystonia associated with TOR1A, otherwise known as DYT1 dystonia. A better understanding of the mechanism of action of THP is a critical step in the development of better therapeutics with fewer side effects. We previously found that THP normalizes the deficit in striatal dopamine (DA) release in a mouse model of TOR1A dystonia (Tor1a+/ΔE knockin (KI) mice), revealing a plausible mechanism of action for this compound, considering that abnormal DA neurotransmission is consistently associated with many forms of dystonia. However, the mAChR subtype(s) that mediate the rescue of striatal dopamine release remain unclear. In this study we used a combination of pharmacological challenges and cell-type specific mAChR conditional knockout mice of either sex to determine which mAChR subtypes mediate the DA release-enhancing effects of THP. We determined that THP acts in part at M4 mAChR on striatal cholinergic interneurons to enhance DA release in both Tor1a+/+ and Tor1a+/ΔE KI mice. Further, we found that the subtype selective M4 antagonist VU6021625 recapitulates the effects of THP. These data implicate a principal role for M4 mAChR located on striatal cholinergic interneurons in the mechanism of action of THP and suggest that subtype selective M4 mAChR antagonists may be effective therapeutics with fewer side effects than THP for the treatment of TOR1A dystonia.
A cyclic AMP related gene network in microglia is inversely regulated by morphine tolerance and withdrawal
Biological Psychiatry Global Open Science
Coffey, K;Lesiak, A;Marx, R;Vo, E;Garden, G;Neumaier, J;
| DOI: 10.1016/j.bpsgos.2021.07.011
Background Microglia have recently been implicated in opioid dependence and withdrawal. Mu Opioid (MOR) receptors are expressed in microglia, and microglia form intimate connections with nearby neurons. Accordingly, opioids have both direct (MOR mediated) and indirect (neuron-interaction mediated) effects on microglia function. Methods To investigate this directly, we used RNA sequencing of ribosome-associated RNAs from striatal microglia (RiboTag-Seq) after the induction of morphine tolerance and followed by naloxone precipitated withdrawal (n=16). We validated the RNA-Seq data by combining fluorescent in-situ hybridization with immunohistochemistry for microglia (n=18). Finally, we expressed and activated the Gi/o-coupled hM4Di DREADD receptor in CX3CR1-expressing cells during morphine withdrawal (n=18). Results We detected large, inverse changes in RNA translation following opioid tolerance and withdrawal. WGCNA analysis revealed an intriguing network of cAMP-associated genes that are known to be involved in microglial motility, morphology, and interactions with neurons that were downregulated with morphine tolerance and upregulated rapidly by withdrawal. Three-dimensional histological reconstruction of microglia allowed for volumetric, visual colocalization of mRNA within individual microglia that validated our bioinformatics results. Direct activation of Gi/o-coupled DREADD receptors in CX3CR1-expressing cells exacerbated signs of opioid withdrawal rather than mimicking the effects of morphine. Conclusions These results indicate that Gi-signaling and cAMP-associated gene networks are inversely engaged during opioid tolerance and early withdrawal, perhaps revealing a role of microglia in mitigating the consequences of opioids.
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RNAscope™ LS 2.5 Probe- Mm-Hesx1
?Cat No. 451878
Automated Assay for Leica Systems - RNAscope
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RNAscope™ 2.5 VS Probe- Mm-Hesx1
?Cat No. 451879
Automated Assay for Ventana Systems - RNAscope
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RNAscope™ HiPlex CS Probe- Mm-Hesx1-T2
?Cat No. 451877-T2
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RNAscope™ HiPlex CS Probe- Mm-Hesx1-T3
?Cat No. 451877-T3
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RNAscope™ HiPlex CS Probe- Mm-Hesx1-T4
?Cat No. 451877-T4
