Nat Neurosci. 2019 Jan;22(1):47-56.
Fu H, Possenti A, Freer R, Nakano Y, Villegas NCH, Tang M, Cauhy PVM, Lassus BA, Chen S, Fowler SL, Figueroa HY, Huey ED, Johnson GVW, Vendruscolo M, Duff KE.
PMID: 30559469 | DOI: 10.1038/s41593-018-0298-7
Excitatory neurons are preferentially impaired in early Alzheimer's disease but the pathways contributing to their relative vulnerability remain largely unknown. Here we report that pathological tau accumulation takes place predominantly in excitatory neurons compared to inhibitory neurons, not only in the entorhinal cortex, a brain region affected in early Alzheimer's disease, but also in areas affected later by the disease. By analyzing RNA transcripts from single-nucleus RNA datasets, we identified a specific tau homeostasis signature of genes differentially expressed in excitatory compared to inhibitory neurons. One of the genes, BCL2-associated athanogene 3 (BAG3), a facilitator of autophagy, was identified as a hub, or master regulator, gene. We verified that reducing BAG3 levels in primary neurons exacerbated pathological tau accumulation, whereas BAG3 overexpression attenuated it. These results define a tau homeostasis signature that underlies the cellular and regional vulnerability of excitatory neurons to tau pathology.
Folorunso, OO;Brown, SE;Baruah, J;Harvey, TL;Jami, SA;Radzishevsky, I;Wolosker, H;McNally, JM;Gray, JA;Vasudevan, A;Balu, DT;
PMID: 37311798 | DOI: 10.1038/s41598-023-35615-5
The proper development and function of telencephalic GABAergic interneurons is critical for maintaining the excitation and inhibition (E/I) balance in cortical circuits. Glutamate contributes to cortical interneuron (CIN) development via N-methyl-D-aspartate receptors (NMDARs). NMDAR activation requires the binding of a co-agonist, either glycine or D-serine. D-serine (co-agonist at many mature forebrain synapses) is racemized by the neuronal enzyme serine racemase (SR) from L-serine. We utilized constitutive SR knockout (SR-/-) mice to investigate the effect of D-serine availability on the development of CINs and inhibitory synapses in the prelimbic cortex (PrL). We found that most immature Lhx6 + CINs expressed SR and the obligatory NMDAR subunit NR1. At embryonic day 15, SR-/- mice had an accumulation of GABA and increased mitotic proliferation in the ganglionic eminence and fewer Gad1 + (glutamic acid decarboxylase 67 kDa; GAD67) cells in the E18 neocortex. Lhx6 + cells develop into parvalbumin (PV+) and somatostatin (Sst+) CINs. In the PrL of postnatal day (PND) 16 SR-/- mice, there was a significant decrease in GAD67+ and PV+, but not SST + CIN density, which was associated with reduced inhibitory postsynaptic potentials in layer 2/3 pyramidal neurons. These results demonstrate that D-serine availability is essential for prenatal CIN development and postnatal cortical circuit maturation.
Godino, A;Salery, M;Durand-de Cuttoli, R;Estill, MS;Holt, LM;Futamura, R;Browne, CJ;Mews, P;Hamilton, PJ;Neve, RL;Shen, L;Russo, SJ;Nestler, EJ;
PMID: 36889314 | DOI: 10.1016/j.neuron.2023.02.013
The complex nature of the transcriptional networks underlying addictive behaviors suggests intricate cooperation between diverse gene regulation mechanisms that go beyond canonical-activity-dependent pathways. Here, we implicate in this process a nuclear receptor transcription factor, retinoid X receptor alpha (RXRα), which we initially identified bioinformatically as associated with addiction-like behaviors. In the nucleus accumbens (NAc) of male and female mice, we show that although its own expression remains unaltered after cocaine exposure, RXRα controls plasticity- and addiction-relevant transcriptional programs in both dopamine receptor D1- and D2-expressing medium spiny neurons, which in turn modulate intrinsic excitability and synaptic activity of these NAc cell types. Behaviorally, bidirectional viral and pharmacological manipulation of RXRα regulates drug reward sensitivity in both non-operant and operant paradigms. Together, this study demonstrates a key role for NAc RXRα in promoting drug addiction and paves the way for future studies of rexinoid signaling in psychiatric disease states.
Brain Struct Funct. 2014 Nov 27.
de Kloet AD, Wang L, Ludin JA, Smith JA, Pioquinto DJ, Hiller H, Steckelings UM, Scheuer DA, Sumners C, Krause EG.
PMID: 25427952
Angiotensin-II acts at its type-1 receptor (AT1R) in the brain to regulate body fluid homeostasis, sympathetic outflow and blood pressure. However, the role of the angiotensin type-2 receptor (AT2R) in the neural control of these processes has received far less attention, largely because of limited ability to effectively localize these receptors at a cellular level in the brain. The present studies combine the use of a bacterial artificial chromosome transgenic AT2R-enhanced green fluorescent protein (eGFP) reporter mouse with recent advances in in situ hybridization (ISH) to circumvent this obstacle. Dual immunohistochemistry (IHC)/ISH studies conducted in AT2R-eGFP reporter mice found that eGFP and AT2R mRNA were highly co-localized within the brain. Qualitative analysis of eGFP immunoreactivity in the brain then revealed localization to neurons within nuclei that regulate blood pressure, metabolism, and fluid balance (e.g., NTS and median preoptic nucleus [MnPO]), as well as limbic and cortical areas known to impact stress responding and mood. Subsequently, dual IHC/ISH studies uncovered the phenotype of specific populations of AT2R-eGFP cells. For example, within the NTS, AT2R-eGFP neurons primarily express glutamic acid decarboxylase-1 (80.3 ± 2.8 %), while a smaller subset express vesicular glutamate transporter-2 (18.2 ± 2.9 %) or AT1R (8.7 ± 1.0 %). No co-localization was observed with tyrosine hydroxylase in the NTS. Although AT2R-eGFP neurons were not observed within the paraventricular nucleus (PVN) of the hypothalamus, eGFP immunoreactivity is localized to efferents terminating in the PVN and within GABAergic neurons surrounding this nucleus. These studies demonstrate that central AT2R are positioned to regulate blood pressure, metabolism, and stress responses.
Kim, JS;Williams, KC;Kirkland, RA;Schade, R;Freeman, KG;Cawthon, CR;Rautmann, AW;Smith, JM;Edwards, GL;Glenn, TC;Holmes, PV;de Lartigue, G;de La Serre, CB;
PMID: 37380023 | DOI: 10.1016/j.molmet.2023.101764
Obesity is associated with deficits in reward which have been linked to compensatory overeating. The vagus nerve is a direct neural pathway that conveys post-ingestive feedback from the gut to the brain, including the reward regions, and vagal activation causes stereotypical reward behaviors. Chronic high fat (HF) feeding alters vagal signaling potentially dampening food-associated reward. Microbiota composition changes rapidly with HF feeding, and a HF-type microbiota is sufficient to alter vagal structure and function. However, whether microbiota-driven alterations in vagal signaling affect host appetitive feeding behavior is unknown. Here, we investigate if microbiota composition modulates reward signaling and assess the role of the vagus in mediating microbiota to brain communication. Male germ-free Fisher rats were colonized with gastrointestinal contents from chow (low fat (LF) ConvLF) or HF (ConvHF) fed rats. Following colonization, ConvHF rats consumed significantly more food than ConvLF animals. ConvHF rats displayed lower feeding-induced extracellular DOPAC levels (a metabolite of dopamine) in the Nucleus Accumbens (NAc) as well as reduced motivation for HF foods compared to ConvLF rats. Dopamine receptor 2 (DDR2) expression levels in the NAc were also significantly lower in ConvHF animals. Similar deficits were observed in conventionally raised HF fed rats, showing that diet-driven alteration in reward can be initiated via microbiota. Selective gut to brain deafferentation restored DOPAC levels, DRD2 expression, and motivational drive in ConvHF rats. We concluded from these data that a HF-type microbiota is sufficient to alter appetitive feeding behavior and that bacteria to reward communication is mediated by the vagus nerve.
Reiner, BC;Zhang, Y;Stein, LM;Perea, ED;Arauco-Shapiro, G;Ben Nathan, J;Ragnini, K;Hayes, MR;Ferraro, TN;Berrettini, WH;Schmidt, HD;Crist, RC;
PMID: 36075888 | DOI: 10.1038/s41398-022-02135-1
Opioid exposure is known to cause transcriptomic changes in the nucleus accumbens (NAc). However, no studies to date have investigated cell type-specific transcriptomic changes associated with volitional opioid taking. Here, we use single nucleus RNA sequencing (snRNAseq) to comprehensively characterize cell type-specific alterations of the NAc transcriptome in rats self-administering morphine. One cohort of male Brown Norway rats was injected with acute morphine (10 mg/kg, i.p.) or saline. A second cohort of rats was allowed to self-administer intravenous morphine (1.0 mg/kg/infusion) for 10 consecutive days. Each morphine-experienced rat was paired with a yoked saline control rat. snRNAseq libraries were generated from NAc punches and used to identify cell type-specific gene expression changes associated with volitional morphine taking. We identified 1106 differentially expressed genes (DEGs) in the acute morphine group, compared to 2453 DEGs in the morphine self-administration group, across 27 distinct cell clusters. Importantly, we identified 1329 DEGs that were specific to morphine self-administration. DEGs were identified in novel clusters of astrocytes, oligodendrocytes, and D1R- and D2R-expressing medium spiny neurons in the NAc. Cell type-specific DEGs included Rgs9, Celf5, Oprm1, and Pde10a. Upregulation of Rgs9 and Celf5 in D2R-expressing neurons was validated by RNAscope. Approximately 85% of all oligodendrocyte DEGs, nearly all of which were associated with morphine taking, were identified in two subtypes. Bioinformatic analyses identified cell type-specific upstream regulatory mechanisms of the observed transcriptome alterations and downstream signaling pathways, including both novel and previously identified molecular pathways. These findings show that volitional morphine taking is associated with distinct cell type-specific transcriptomic changes in the rat NAc and highlight specific striatal cell populations and novel molecular substrates that could be targeted to reduce compulsive opioid taking.
Caprioli D, Venniro M, Zhang M, Bossert JM, Warren BL, Hope BT, Shaham Y.
PMID: 27980115 | DOI: 10.1523/JNEUROSCI.3091-16.2016
We recently developed a rat model of incubation of methamphetamine craving after choice-based voluntary abstinence. Here, we studied the role of dorsolateral and dorsomedial striatum (DLS, DMS) in this incubation.We trained rats to self-administer palatable food pellets (6 days, 6-h/d) and methamphetamine (12 days, 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/day) for 19 days. We used in situ hybridization to measure co-labeling of the activity marker Fos with Drd1 and Drd2 in DMS and DLS after the tests. Based on the in situ hybridization co-labeling results, we tested the causal role of DMS D1- and D2-family 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 days of voluntary abstinence than after 1 day (incubation of methamphetamine craving). The 'incubated' response was associated with increased Fos expression in DMS but not DLS; Fos was co-labeled 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 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.
Quina LA1, Walker A1, Morton G1, Han V1, Turner EE2,3
PMID: 32332079 | DOI: 10.1523/ENEURO.0527-19.2020
The lateral habenula (LHb) sends complex projections to several areas of the mesopontine tegmentum, the raphe, and the hypothalamus. However, few markers have been available to distinguish subsets of LHb neurons that may serve these pathways. In order to address this complexity, we examined the mouse and rat LHb for neurons that express the GABA biosynthesis enzymes glutamate decarboxylase 1 and 2 (GAD1, GAD2), and the vesicular GABA transporter (VGAT). The mouse LHb contains a population of neurons that express GAD2, while the rat LHb contains discrete populations of neurons that express GAD1 and VGAT. However, we could not detect single neurons in either species that co-express a GABA synthetic enzyme and VGAT, suggesting that these LHb neurons do not use GABA for conventional synaptic transmission. Instead, all of the neuronal types expressing a GABAergic marker in both species showed co-expression of the glutamate transporter VGluT2. Anterograde tract-tracing of the projections of GAD2-expressing LHb neurons in Gad2Cre mice, combined with retrograde tracing from selected downstream nuclei, show that LHb-GAD2 neurons project selectively to the midline structures in the mesopontine tegmentum, including the median raphe and nucleus incertus, and only sparsely innervate the hypothalamus, rostromedial tegmental nucleus, and ventral tegmental area. Postsynaptic recording of LHb-GAD2 neuronal input to tegmental neurons confirms that glutamate, not GABA, is the fast neurotransmitter in this circuit. Thus GAD2 expression can serve as a marker for functional studies of excitatory neurons serving specific LHb output pathways in mice.SIGNFICANCE STATEMENT The lateral habenula provides a major link between subcortical forebrain areas and the dopamine (DA) and serotonin (5HT) systems of the midbrain and pons, and it has been implicated in reward mechanisms and the regulation of mood states. Few markers have been available for the specific cell types and complex output pathways of the lateral habenula. Here we examined the expression of genes mediating GABAergic and glutamatergic transmission in the mouse and rat LHb, where no neurons in either species expressed a full complement of GABAergic markers, and all expressed the glutamatergic marker VGluT2. Consistent with this, in mice the LHb GAD2 neurons are excitatory and appear to use only glutamate for fast synaptic transmission.
Charbogne P, Gardon O, Martín-García E, Keyworth HL, Matsui A, Mechling AE, Bienert T, Nasseef T, Robé A, Moquin L, Darcq E, Hamida SB, Robledo P, Matifas A, Befort K, Gavériaux-Ruff , Harsan LA, Von Everfeldt D, Hennig J, Gratton A, Kitchen I, Bailey A,
PMID: - | DOI: 10.1016/j.biopsych.2016.12.022
Background
Mu opioid receptors (MORs) are central to pain control, drug reward and addictive behaviors, but underlying circuit mechanisms have been poorly explored by genetic approaches. Here we investigate the contribution of MORs expressed in GABAergic forebrain neurons to major biological effects of opiates, and also challenge the canonical disinhibition model of opiate reward.
Methods
We used Dlx5/6-mediated recombination to create conditional Oprm1 mice in GABAergic forebrain neurons. We characterized the genetic deletion by histology, electrophysiology and microdialysis, probed neuronal activation by c-Fos immunohistochemistry and resting state-functional magnetic resonance imaging, and investigated main behavioral responses to opiates, including motivation to obtain heroin and palatable food.
Results
Mutant mice showed MOR transcript deletion mainly in the striatum. In the ventral tegmental area (VTA), local MOR activity was intact, and reduced activity was only observed at the level of striatonigral afferents. Heroin-induced neuronal activation was modified at both sites, and whole-brain functional networks were altered in live animals. Morphine analgesia was not altered, neither was physical dependence to chronic morphine. In contrast, locomotor effects of heroin were abolished, and heroin-induced catalepsy was increased. Place preference to heroin was not modified, but remarkably, motivation to obtain heroin and palatable food was enhanced in operant self-administration procedures.
Conclusions
Our study reveals dissociable MOR functions across mesocorticolimbic networks. Thus beyond a well-established role in reward processing, operating at the level of local VTA neurons, MORs also moderate motivation for appetitive stimuli within forebrain circuits that drive motivated behaviors.
Patrick Card J, Johnson AL, Llewellyn-Smith IJ, Zheng H, Anand R, Brierley DI, Trapp S, Rinaman L.
PMID: 30019398 | DOI: 10.1002/cne.24482
Glutamatergic neurons that express pre-proglucagon (PPG) and are immunopositive (+) for glucagon-like peptide-1 (i.e., GLP-1+ neurons) are located within the caudal nucleus of the solitary tract (cNTS) and medullary reticular formation in rats and mice. GLP-1 neurons give rise to an extensive central network in which GLP-1 receptor (R) signaling suppresses food intake, attenuates rewarding, increases avoidance, and stimulates stress responses, partly via . GLP-1R signaling within the cNTS. In mice, noradrenergic (A2) cNTS neurons express GLP-1R, whereas PPG neurons do not. In the present study, confocal microscopy in rats confirmed that prolactin-releasing peptide (PrRP)+ A2 neurons are closely apposed by GLP-1+ axonal varicosities. Surprisingly, GLP-1+ appositions were also observed on dendrites of PPG/GLP-1+ neurons in both species, and electron microscopy in rats revealed that GLP-1+ boutons form asymmetric synaptic contacts with GLP-1+ dendrites. However, RNAscope confirmed that rat GLP-1 neurons do not express GLP-1R mRNA. Similarly, Ca2+ imaging of somatic and dendritic responses in mouse ex vivo slices confirmed that PPG neurons do not respond directly to GLP-1, and a mouse cross-breeding strategy revealed that fewer than 1% of PPG neurons co-express GLP-1R. Collectively, these data suggest that GLP-1R signaling pathways modulate the activity of PrRP+ A2 neurons, and also reveal a local "feed-forward" synaptic network among GLP-1 neurons that apparently does not utilize GLP-1R signaling. This local GLP-1 network may instead use glutamatergic signaling to facilitate dynamic and potentially selective recruitment of GLP-1 neural populations that shape behavioral and physiological responses to internal and external challenges.
Teague, C;Picone, J;Wright, W;Browne, C;Silva, G;Futamura, R;Minier-Toribio, A;Estill, M;Ramakrishnan, A;Martinez-Rivera, F;Godino, A;Parise, E;Schmidt, K;Pulido, N;Lorsch, Z;Kim, J;Shen, L;Neve, R;Dong, Y;Nestler, E;Hamilton, P;
| DOI: 10.1016/j.biopsych.2022.07.022
Background Over the course of chronic drug use, brain transcriptional neuroadaptation are thought to contribute to a change in drug use behavior over time. The function of the transcription factor CREB within the nucleus accumbens (NAc) has been well documented in opposing the rewarding properties of many classes of drugs, yet the gene targets through which CREB causally manifests these lasting neuroadaptations remain unknown. Here, we identify zinc finger protein 189 (Zfp189) as a CREB target gene that is transcriptionally responsive to acute and chronic cocaine use within mouse NAc. Methods To query the role of the CREB-Zfp189 interaction in cocaine use, we virally delivered modified CRISPR/dCas9 constructs, capable of selectively localizing CREB to the Zfp189 gene promoter in the NAc of mice. Results We observe that CREB binding to the Zfp189 promoter increases Zfp189 expression and diminishes the reinforcing responses to cocaine. We show further that NAc Zfp189 expression is increased within D1 medium spiny neurons (MSNs) in response to acute cocaine, but increased in both D1 and D2 MSNs in response to chronic cocaine. CREB-mediated induction of Zfp189 potentiates electrophysiological activity of D1 and D2 MSNs - recapitulating the known effect of CREB on these neurons. Lastly, targeting CREB to the Zfp189 promoter within NAc Drd2-expressing neurons, but not Drd1-expressing neurons, was sufficient to diminish cocaine-conditioned behaviors. Conclusions Together, these findings point to the CREB-Zfp189 interaction within NAc Drd2+ neurons as a molecular signature of chronic cocaine use that is causal in counteracting the reinforcing effects of cocaine.
Vectorology for Optogenetics and Chemogenetics
Lin, J;Dimidschstein, J;
| DOI: 10.1007/978-1-0716-2918-5_9
Recombinant adeno-associated viruses can be coupled with short regulatory elements to restrict viral expression to specific cellular populations. These viral vectors can be used as tools for basic research to dissect many aspects of the biology of specific cellular subtypes in health and disease, and across species. A handful of enhancers have already been described in the nervous system, and recent studies suggest that transcriptomic and epigenetic data can be leveraged to systematize the discovery of novel elements to restrict viral expression to any cell type. However, a thorough characterization of the expression profile conferred by these short sequences is required to demonstrate their utility in the experimental context in which they will be ultimately used. Here we describe a complete guide to select, screen, and validate the expression profile of enhancers to target specific subtypes of neurons.
