Repeated cocaine administration upregulates CB2 receptor expression in striatal medium-spiny neurons that express dopamine D1 receptors in mice
Acta pharmacologica Sinica
Zhang, HY;De Biase, L;Chandra, R;Shen, H;Liu, QR;Gardner, E;Lobo, MK;Xi, ZX;
PMID: 34316031 | DOI: 10.1038/s41401-021-00712-6
Cannabinoid CB2 receptors (CB2R) are importantly involved in drug reward and addiction. However, the cellular mechanisms underlying CB2R action remain unclear. We have previously reported that cocaine self-administration upregulates CB2R expression in midbrain dopamine (DA) neurons. In the present study, we investigated whether cocaine or heroin also alters CB2R expression in striatal medium-spiny neurons that express dopamine D1 or D2 receptors (D1-MSNs, D2-MSNs) and microglia. Due to the concern of CB2R antibody specificity, we developed three mouse CB2-specific probes to detect CB2R mRNA using quantitative RT-PCR and RNAscope in situ hybridization (ISH) assays. We found that a single injection of cocaine failed to alter, while repeated cocaine injections or self-administration dose-dependently upregulated CB2R gene expression in both brain (cortex and striatum) and periphery (spleen). In contrast, repeated administration of heroin produced a dose-dependent reduction in striatal CB2 mRNA expression. RNAscope ISH assays detected CB2R mRNA in striatal D1- and D2-MSNs, not in microglia. We then used transgenic CX3CR1eGFP/+ microglia reporter mice and D1- or D2-Cre-RiboTag mice to purify striatal microglia or ribosome-associated mRNAs from CX3CR1eGFP/+, D1-MSNs, or D2-MSNs, respectively. We found that CB2R upregulation occurred mainly in D1-MSNs, not in D2-MSNs or microglia, in the nucleus accumbens rather than the dorsal striatum. These findings indicate that repeated cocaine exposure may upregulate CB2R expression in both brain and spleen, with regional and cell type-specific profiles. In the striatum, CB2R upregulation occurs mainly in D1-MSNs in the nucleus accumbens. Given the important role of D1-MSNs in brain reward function, the present findings provide new insight into mechanisms by which brain CB2Rs modulate cocaine action.
Plescher M, Seifert G, Hansen JN, Bedner P, Steinhäuser C, Halle A.
PMID: 29493017 | DOI: 10.1002/glia.23318
Microglia, the central nervous system resident innate immune cells, cluster around Aβ plaques in Alzheimer's disease (AD). The activation phenotype of these plaque-associated microglial cells, and their differences to microglia distant to Aβ plaques, are incompletely understood. We used novel three-dimensional cell analysis software to comprehensively analyze the morphological properties of microglia in the TgCRND8 mouse model of AD in spatial relation to Aβ plaques. We found strong morphological changes exclusively in plaque-associated microglia, whereas plaque-distant microglia showed only minor changes. In addition, patch-clamp recordings of microglia in acute cerebral slices of TgCRND8 mice revealed increased K+ currents in plaque-associated but not plaque-distant microglia. Within the subgroup of plaque-associated microglia, two different current profiles were detected. One subset of cells displayed only increased inward currents, while a second subset showed both increased inward and outward currents, implicating that the plaque microenvironment differentially impacts microglial ion channel expression. Using pharmacological channel blockers, multiplex single-cell PCR analysis and RNA fluorescence in situ hybridization, we identified Kir and Kv channel types contributing to the in- and outward K+ conductance in plaque-associated microglia. In summary, we have identified a previously unrecognized level of morphological and electrophysiological heterogeneity of microglia in relation to amyloid plaques, suggesting that microglia may display multiple activation states in AD.
Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology
McNulty, CJ;Fallon, IP;Amat, J;Sanchez, RJ;Leslie, NR;Root, DH;Maier, SF;Baratta, MV;
PMID: 36076018 | DOI: 10.1038/s41386-022-01443-w
Stress-linked disorders are more prevalent in women than in men and differ in their clinical presentation. Thus, investigating sex differences in factors that promote susceptibility or resilience to stress outcomes, and the circuit elements that mediate their effects, is important. In male rats, instrumental control over stressors engages a corticostriatal system involving the prelimbic cortex (PL) and dorsomedial striatum (DMS) that prevent many of the sequelae of stress exposure. Interestingly, control does not buffer against stress outcomes in females, and here, we provide evidence that the instrumental controlling response in females is supported instead by the dorsolateral striatum (DLS). Additionally, we used in vivo microdialysis, fluorescent in situ hybridization, and receptor subtype pharmacology to examine the contribution of prefrontal dopamine (DA) to the differential impact of behavioral control. Although both sexes preferentially expressed D1 receptor mRNA in PL GABAergic neurons, there were robust sex differences in the dynamic properties of prefrontal DA during controllable stress. Behavioral control potently attenuated stress-induced DA efflux in males, but not females, who showed a sustained DA increase throughout the entire stress session. Importantly, PL D1 receptor blockade (SCH 23390) shifted the proportion of striatal activity from the DLS to the DMS in females and produced the protective effects of behavioral control. These findings suggest a sex-selective mechanism in which elevated DA in the PL biases instrumental responding towards prefrontal-independent striatal circuitry, thereby eliminating the protective impact of coping with stress.
Grunddal, KV;Jensen, EP;Ørskov, C;Andersen, DB;Windeløv, JA;Poulsen, SS;Rosenkilde, MM;Knudsen, LB;Pyke, C;Holst, JJ;
PMID: 34662392 | DOI: 10.1210/endocr/bqab216
Therapies based on glucagon-like peptide-1 receptor (GLP-1R) agonism are highly effective in treating type 2 diabetes and obesity, but the localization of GLP-1Rs mediating the antidiabetic and other possible actions of GLP-1 is still debated. The purpose with this study was to identify sites of GLP-1R mRNA and protein expression in the mouse gastrointestinal system by means of GLP-1R antibody immunohistochemistry, Glp1r mRNA fluorescence in situ hybridization, and 125I-exendin (9-39) autoradiography. As expected, GLP-1R staining was observed in almost all β-cells in the pancreatic islets, but more rarely in α- and δ-cells. In the stomach, GLP-1R staining was found exclusively in the gastric corpus mucous neck cells, known to protect the stomach mucosa. The Brunner glands were strongly stained for GLP-1R, and pretreatment with GLP-1 agonist exendin-4 caused internalization of the receptor and mucin secretion, while pretreatment with phosphate-buffered saline or antagonist exendin (9-39) did not. In the intestinal mucosa, GLP-1R staining was observed in intraepithelial lymphocytes, lamina propria lymphocytes, and enteroendocrine cells containing secretin, peptide YY, and somatostatin, but not cholecystokinin. GLP-1R staining was seen in nerve fibers within the choline acetyl transferase- and nitric oxide-positive myenteric plexuses from the gastric corpus to the distal large intestine being strongest in the mid- and hindgut area. Finally, intraperitoneal administration of radiolabeled exendin (9-39) strongly labeled myenteric fibers. In conclusion, this study expands our knowledge of GLP-1R localization and suggests that GLP-1 may serve an important role in modulating gastrointestinal health and mucosal protection.
Journal of Neuroendocrinology
Watanabe, Y;Fisher, L;Campbell, R;Jasoni, C;
| DOI: 10.1111/jne.13302
Polycystic ovary syndrome (PCOS) is a female endocrine disorder that is associated with prenatal exposure to excess androgens. In prenatally androgenized (PNA) mice that model PCOS, GABAergic neural transmission to and innervation of GnRH neurons is increased. Evidence suggests that elevated GABAergic innervation originates in the arcuate nucleus (ARC). We hypothesised that GABA-GnRH circuit abnormalities are a direct consequence of PNA, resulting from DHT binding to androgen receptor (AR) in the prenatal brain. However, whether prenatal ARC neurons express AR at the time of PNA treatment is presently unknown. We used RNAScope _in situ_ hybridization to localize AR mRNA (_Ar_)-expressing cells in healthy gestational day (GD) 17.5 female mouse brains and to assess co-expression levels in specific neuronal phenotypes. Our study revealed that less than 10% of ARC GABA cells expressed _Ar_. In contrast, we found that ARC kisspeptin neurons, critical regulators of GnRH neurons, were highly co-localised with _Ar_. Approximately 75% of ARC _Kiss1_-expressing cells also expressed _Ar_ at GD17.5, suggesting that ARC kisspeptin neurons are potential targets of PNA. Investigating other neuronal populations in the ARC we found that approximately 50% of pro-opiomelanocortin (_Pomc_) cells, 22% of tyrosine hydroxylase (_Th_) cells, 8% of agouti-related protein (_Agrp_) cells and 8% of somatostatin (_Sst_) cells express _Ar_. Lastly, RNAscope in coronal sections showed _Ar_ expression in the medial preoptic area (mPOA), and the ventral part of the lateral septum (vLS). These _Ar_-expressing regions were highly GABAergic, and 22% of GABA cells in the mPOA and 25% of GABA cells in the vLS also expressed _Ar_. Our findings identify specific neuronal phenotypes in the ARC, mPOA and vLS that are androgen sensitive in late gestation. PNA-induced functional changes in these neurons may be related to the development of impaired central mechanisms associated with PCOS-like features.
Lysko, DE;Talbot, WS;
PMID: 36384112 | DOI: 10.1016/j.celrep.2022.111669
The signaling mechanisms neurons use to modulate myelination of circuits in the central nervous system (CNS) are only partly understood. Through analysis of isoform-specific neuregulin1 (nrg1) mutants in zebrafish, we demonstrate that nrg1 type II is an important regulator of myelination of two classes of spinal cord interneurons. Surprisingly, nrg1 type II expression is prominent in unmyelinated Rohon-Beard sensory neurons, whereas myelination of neighboring interneurons is reduced in nrg1 type II mutants. Cell-type-specific loss-of-function studies indicate that nrg1 type II is required in Rohon-Beard neurons to signal to other neurons, not oligodendrocytes, to modulate spinal cord myelination. Together, our data support a model in which unmyelinated neurons express Nrg1 type II proteins to regulate myelination of neighboring neurons, a mode of action that may coordinate the functions of unmyelinated and myelinated neurons in the CNS.
Acta pharmacologica Sinica
Chen, ZJ;Su, CW;Xiong, S;Li, T;Liang, HY;Lin, YH;Chang, L;Wu, HY;Li, F;Zhu, DY;Luo, CX;
PMID: 36460834 | DOI: 10.1038/s41401-022-01024-z
Chronic pain patients often have anxiety disorders, and some of them suffer from anxiety even after analgesic administration. In this study, we investigated the role of AMPAR-mediated synaptic transmission in the ventromedial prefrontal cortex (vmPFC) in chronic pain-induced persistent anxiety in mice and explored potential drug targets. Chronic inflammatory pain was induced in mice by bilateral injection of complete Freund's adjuvant (CFA) into the planta of the hind paws; anxiety-like behaviours were assessed with behavioural tests; S-nitrosylation and AMPAR-mediated synaptic transmission were examined using biochemical assays and electrophysiological recordings, respectively. We found that CFA induced persistent upregulation of AMPAR membrane expression and function in the vmPFC of anxious mice but not in the vmPFC of non-anxious mice. The anxious mice exhibited higher S-nitrosylation of stargazin (an AMPAR-interacting protein) in the vmPFC. Inhibition of S-nitrosylation by bilaterally infusing an exogenous stargazin (C302S) mutant into the vmPFC rescued the surface expression of GluA1 and AMPAR-mediated synaptic transmission as well as the anxiety-like behaviours in CFA-injected mice, even after ibuprofen treatment. Moreover, administration of ZL006, a small molecular inhibitor disrupting the interaction of nNOS and PSD-95 (20 mg·kg-1·d-1, for 5 days, i.p.), significantly reduced nitric oxide production and S-nitrosylation of AMPAR-interacting proteins in the vmPFC, resulting in anxiolytic-like effects in anxious mice after ibuprofen treatment. We conclude that S-nitrosylation is necessary for AMPAR trafficking and function in the vmPFC under chronic inflammatory pain-induced persistent anxiety conditions, and nNOS-PSD-95 inhibitors could be potential anxiolytics specific for chronic inflammatory pain-induced persistent anxiety after analgesic treatment.
Molecular Neuropsychiatry
Hu X,. Rocco BR, Fee C, Sibille E.
PMID: - | DOI: 10.1159/000495840
Converging evidence suggests that deficits in somatostatin (SST)-expressing neuron signaling contributes to major depressive disorder. Preclinical studies show that enhancing this signaling, specifically at α5 subunit-containing γ-aminobutyric acid subtype A receptors (α5-GABAARs), provides a potential means to overcome low SST neuron function. The cortical microcircuit comprises multiple subtypes of inhibitory γ-aminobutyric acid (GABA) neurons and excitatory pyramidal cells (PYCs). In this study, multilabel fluorescence in situ hybridization was used to characterize α5-GABAAR gene expression in PYCs and three GABAergic neuron subgroups – vasoactive intestinal peptide (VIP)-, SST-, and parvalbumin (PV)-expressing cells – in the human and mouse frontal cortex. Across species, we found the majority of gene expression in PYCs (human: 39.7%; mouse: 54.14%), less abundant expression in PV neurons (human: 20%; mouse: 16.33%), and no expression in VIP neurons (0%). Only human SST cells expressed GABRA5, albeit at low levels (human: 8.3%; mouse: 0%). Together, this localization suggests potential roles for α5-GABAARs within the cortical microcircuit: (1) regulators of PYCs, (2) regulators of PV cell activity across species, and (3) sparse regulators of SST cell inhibition in humans. These results will advance our ability to predict the effects of pharmacological agents targeting α5-GABAARs, which have shown therapeutic potential in preclinical animal models.
J Ovarian Res. 2015 May 14;8(1):29
Abstract BACKGROUND: Folate receptor alpha (FOLR1/FRA) is expressed in a number of epithelial cancers and in particular epithelial ovarian cancer (EOC), especially of the serous histotype. Recent studies have shown that EOC originates from the fallopian tube fimbriae rather than from epithelial cells lining the ovary. We have previously shown by immunohistochemistry a strong correlation between FRA expression in EOC and normal and fallopian adenocarcinoma. Folate receptor beta (FOLR2/FRB) has been described to be expressed by macrophages both in inflammatory disorders and certain epithelial cancers. Given the high sequence identity of these two folate receptor family members we sought to investigate the architectural and cell-specific expression of these two receptors in gynecologic tissues. METHODS: RNA scope, a novel chromogenic in situ hybridization assay tool, was used to examine expression of the alpha (FOLR1) and beta (FOLR2) isoforms of folate receptor relative to each other as well as to the macrophage markers CD11b and CD68, in samples of normal fallopian tube and fallopian adenocarcinoma as well as normal ovary and EOC. RESULTS: We demonstrated expression of both FOLR1 and FOLR2 in EOC, normal fallopian tube and fallopian adenocarcinoma tissue while very little expression of either marker was observed in normal ovary. Furthermore, FOLR2 was shown to be expressed almost exclusively in macrophages, of both the M1 and M2 lineages, as determined by co-expression of CD11b and/or CD68, with little or no expression in epithelial cells. CONCLUSIONS: These findings further substantiate the hypothesis that the cell of origin of EOC is tubal epithelium and that the beta isoform of folate receptor is primarily restricted to macrophages. Further, macrophages expressing FOLR2 may represent tumor associated or infiltrating macrophages (TAMs) in epithelial cancers.
Xue, T;Wang, X;Hu, Y;Cheng, Y;Li, H;Shi, Y;Wang, L;Yin, D;Cui, D;
PMID: 36291328 | DOI: 10.3390/brainsci12101395
The brain is susceptible to perturbations of redox balance, affecting neurogenesis and increasing the risks of psychiatric disorders. Thioredoxin-interacting protein (TXNIP) is an endogenous inhibitor of the thioredoxin antioxidant system. Its deletion or inhibition suggests protection for a brain with ischemic stroke or Alzheimer's disease. Combined with conditional knockout mice and schizophrenia samples, we aimed to investigate the function of TXNIP in healthy brain and psychiatric disorders, which are under-studied. We found TXNIP was remarkedly expressed in the prefrontal cortex (PFC) during healthy mice's prenatal and early postnatal periods, whereas it rapidly decreased throughout adulthood. During early life, TXNIP was primarily distributed in inhibitory and excitatory neurons. Contrary to the protective effect, the embryonic deletion of TXNIP in GABAergic (gamma-aminobutyric acid-ergic) neurons enhanced oxidative stress in PV+ interneurons of aging mice. The deleterious impact was brain region-specific. We also investigated the relationship between TXNIP and schizophrenia. TXNIP was significantly increased in the PFC of schizophrenia-like mice after MK801 administration, followed by oxidative stress. First episode and drug naïve schizophrenia patients with a higher level of plasma TXNIP displayed severer psychiatric symptoms than patients with a low level. We indicated a bidirectional function of TXNIP in the brain, whose high expression in the early stage is protective for development but might be harmful in a later period, associated with mental disorders.
Yu, B;Zhang, Q;Lin, L;Zhou, X;Ma, W;Wen, S;Li, C;Wang, W;Wu, Q;Wang, X;Li, XM;
PMID: 36788214 | DOI: 10.1038/s41421-022-00506-y
The amygdala, or an amygdala-like structure, is found in the brains of all vertebrates and plays a critical role in survival and reproduction. However, the cellular architecture of the amygdala and how it has evolved remain elusive. Here, we generated single-nucleus RNA-sequencing data for more than 200,000 cells in the amygdala of humans, macaques, mice, and chickens. Abundant neuronal cell types from different amygdala subnuclei were identified in all datasets. Cross-species analysis revealed that inhibitory neurons and inhibitory neuron-enriched subnuclei of the amygdala were well-conserved in cellular composition and marker gene expression, whereas excitatory neuron-enriched subnuclei were relatively divergent. Furthermore, LAMP5+ interneurons were much more abundant in primates, while DRD2+ inhibitory neurons and LAMP5+SATB2+ excitatory neurons were dominant in the human central amygdalar nucleus (CEA) and basolateral amygdalar complex (BLA), respectively. We also identified CEA-like neurons and their species-specific distribution patterns in chickens. This study highlights the extreme cell-type diversity in the amygdala and reveals the conservation and divergence of cell types and gene expression patterns across species that may contribute to species-specific adaptations.
Frontiers in synaptic neuroscience
Garcia DuBar, S;Cosio, D;Korthas, H;Van Batavia, JP;Zderic, SA;Sahibzada, N;Valentino, RJ;Vicini, S;
PMID: 34675794 | DOI: 10.3389/fnsyn.2021.754786
The pontine nuclei comprising the locus coeruleus (LC) and Barrington's nucleus (BRN) amongst others form the neural circuitry(s) that coordinates arousal and voiding behaviors. However, little is known about the synaptic connectivity of neurons within or across these nuclei. These include corticotropin-releasing factor (CRF+) expressing neurons in the BRN that control bladder contraction and somatostatin expressing (SST+) neurons whose role in this region has not been discerned. To determine the synaptic connectivity of these neurons, we employed optogenetic stimulation with recordings from BRN and LC neurons in brain stem slices of channelrhodopsin-2 expressing SST or CRF neurons. Optogenetic stimulation of CRF+ BRN neurons of Crf Cre ;chr2-yfp mice had little effect on either CRF+ BRN neurons, CRF- BRN neurons, or LC neurons. In contrast, in Sst Cre ;chr2-yfp mice light-activated inhibitory postsynaptic currents (IPSCs) were reliably observed in a majority of LC but not BRN neurons. The GABAA receptor antagonist, bicuculline, completely abolished the light-induced IPSCs. To ascertain if these neurons were part of the neural circuitry that controls the bladder, the trans-synaptic tracer, pseudorabies virus (PRV) was injected into the bladder wall of Crf Cre ;tdTomato or Sst Cre ;tdTomato mice. At 68-72 h post-viral infection, PRV labeled neurons were present only in the BRN, being preponderant in CRF+ neurons with few SST+ BRN neurons labeled from the bladder. At 76 and 96 h post-virus injection, increased labeling was observed in both BRN and LC neurons. Our results suggest SST+ neurons rather than CRF+ neurons in BRN can regulate the activity of LC neurons.
