Skirzewski M, Karavanova I, Shamir A, Erben L, Garcia-Olivares J, Shin JH, Vullhorst D, Alvarez VA, Amara SG, Buonanno A.
PMID: 28727685 | DOI: 10.1038/mp.2017.132
Genetic variants of Neuregulin 1 (NRG1) and its neuronal tyrosine kinase receptor ErbB4 are associated with risk for schizophrenia, a neurodevelopmental disorder characterized by excitatory/inhibitory imbalance and dopamine (DA) dysfunction. To date, most ErbB4 studies have focused on GABAergic interneurons in the hippocampus and neocortex, particularly fast-spiking parvalbumin-positive (PV+) basket cells. However, NRG has also been shown to modulate DA levels, suggesting a role for ErbB4 signaling in dopaminergic neuron function. Here we report that ErbB4 in midbrain DAergic axonal projections regulates extracellular DA levels and relevant behaviors. Mice lacking ErbB4 in tyrosine hydroxylase-positive (TH+) neurons, but not in PV+ GABAergic interneurons, exhibit different regional imbalances of basal DA levels and fail to increase DA in response to local NRG1 infusion into the dorsal hippocampus, medial prefrontal cortex and dorsal striatum measured by reverse microdialysis. Using Lund Human Mesencephalic (LUHMES) cells, we show that NRG/ErbB signaling increases extracellular DA levels, at least in part, by reducing DA transporter (DAT)-dependent uptake. Interestingly, TH-Cre;ErbB4f/f mice manifest deficits in learning, spatial and working memory-related behaviors, but not in numerous other behaviors altered in PV-Cre;ErbB4f/fmice. Importantly, microinjection of a Cre-inducible ErbB4 virus (AAV-ErbB4.DIO) into the mesencephalon of TH-Cre;ErbB4f/f mice, which selectively restores ErbB4 expression in DAergic neurons, rescues DA dysfunction and ameliorates behavioral deficits. Our results indicate that direct NRG/ErbB4 signaling in DAergic axonal projections modulates DA homeostasis, and that NRG/ErbB4 signaling in both GABAergic interneurons and DA neurons contribute to the modulation of behaviors relevant to psychiatric disorders.
Liu QR, Canseco-Alba A, Zhang HY, Tagliaferro P, Chung M, Dennis E, Sanabria B, Schanz N, Escosteguy-Neto JC, Ishiguro H, Lin Z, Sgro S, Leonard CM, Santos-Junior JG, Gardner EL, Egan JM, Lee JW, Xi ZX, Onaivi ES.
PMID: 29234141 | DOI: 10.1038/s41598-017-17796-y
Cannabinoid CB2 receptors (CB2Rs) are expressed in mouse brain dopamine (DA) neurons and are involved in several DA-related disorders. However, the cell type-specific mechanisms are unclear since the CB2R gene knockout mice are constitutive gene knockout. Therefore, we generated Cnr2-floxed mice that were crossed with DAT-Cre mice, in which Cre- recombinase expression is under dopamine transporter gene (DAT) promoter control to ablate Cnr2 gene in midbrain DA neurons of DAT-Cnr2 conditional knockout (cKO) mice. Using a novel sensitive RNAscope in situ hybridization, we detected CB2R mRNA expression in VTA DA neurons in wildtype and DAT-Cnr2 cKO heterozygous but not in the homozygous DAT-Cnr2 cKO mice. Here we report that the deletion of CB2Rs in dopamine neurons enhances motor activities, modulates anxiety and depression-like behaviors and reduces the rewarding properties of alcohol. Our data reveals that CB2Rs are involved in the tetrad assay induced by cannabinoids which had been associated with CB1R agonism. GWAS studies indicates that the CNR2 gene is associated with Parkinson's disease and substance use disorders. These results suggest that CB2Rs in dopaminergic neurons may play important roles in the modulation of psychomotor behaviors, anxiety, depression, and pain sensation and in the rewarding effects of alcohol and cocaine.
Brindley RL, Bauer MB, Walker LA, Quinlan MA, Carneiro AMD, Sze JY, Blakely RD, Currie KPM.
PMID: 29894763 | DOI: 10.1016/j.phrs.2018.06.008
Adrenal chromaffin cells comprise the neuroendocrine arm of the sympathetic nervous system and secrete catecholamines to coordinate the appropriate stress response. Deletion of the serotonin (5-HT) transporter (SERT) gene in mice (SERT-/- mice) or pharmacological block of SERT function in rodents and humans augments this sympathoadrenal stress response (epinephrine secretion). The prevailing assumption is that loss of CNS SERT alters central drive to the peripheral sympathetic nervous system. Adrenal chromaffin cells also prominently express SERT where it might coordinate accumulation of 5-HT for reuse in the autocrine control of stress-evoked catecholamine secretion. To help test this hypothesis, we have generated a novel mouse model with selective excision of SERT in the peripheral sympathetic nervous system (SERTΔTH), generated by crossing floxed SERT mice with tyrosine hydroxylase Cre driver mice. SERT expression, assessed by western blot, was abolished in the adrenal gland but not perturbed in the CNS of SERTΔTH mice. SERT-mediated [3H] 5-HT uptake was unaltered in midbrain, hindbrain, and spinal cord synaptosomes, confirming transporter function was intact in the CNS. Endogenous midbrain and whole blood 5-HT homeostasis was unperturbed in SERTΔTH mice, contrasting with the depleted 5-HT content in SERT-/- mice. Selective SERT excision reduced adrenal gland 5-HT content by ≈ 50% in SERTΔTH mice but had no effect on adrenal catecholamine content. This novel model confirms that SERT expressed in adrenal chromaffin cells is essential for maintaining wild-type levels of 5-HT and provides a powerful tool to help dissect the role of SERT in the sympathetic stress response.
Imaging mass cytometry reveals generalised deficiency in OXPHOS complexes in Parkinson\'s disease
Chen, C;McDonald, D;Blain, A;Sachdeva, A;Bone, L;Smith, ALM;Warren, C;Pickett, SJ;Hudson, G;Filby, A;Vincent, AE;Turnbull, DM;Reeve, AK;
PMID: 33980828 | DOI: 10.1038/s41531-021-00182-x
Here we report the application of a mass spectrometry-based technology, imaging mass cytometry, to perform in-depth proteomic profiling of mitochondrial complexes in single neurons, using metal-conjugated antibodies to label post-mortem human midbrain sections. Mitochondrial dysfunction, particularly deficiency in complex I has previously been associated with the degeneration of dopaminergic neurons in Parkinson's disease. To further our understanding of the nature of this dysfunction, and to identify Parkinson's disease specific changes, we validated a panel of antibodies targeting subunits of all five mitochondrial oxidative phosphorylation complexes in dopaminergic neurons from Parkinson's disease, mitochondrial disease, and control cases. Detailed analysis of the expression profile of these proteins, highlighted heterogeneity between individuals. There is a widespread decrease in expression of all complexes in Parkinson's neurons, although more severe in mitochondrial disease neurons, however, the combination of affected complexes varies between the two groups. We also provide evidence of a potential neuronal response to mitochondrial dysfunction through a compensatory increase in mitochondrial mass. This study highlights the use of imaging mass cytometry in the assessment and analysis of expression of oxidative phosphorylation proteins, revealing the complexity of deficiencies of these proteins within individual neurons which may contribute to and drive neurodegeneration in Parkinson's disease.
International journal of cancer
Mangiaterra, TS;De Dios Soler, M;Oviedo, N;Colli, S;Preciado, MV;Soria, M;Galluzo, L;De Matteo, E;Chabay, P;
PMID: 37318089 | DOI: 10.1002/ijc.34623
In 2017, the World Health Organization (WHO) confirmed a new entity, Epstein Barr virus (EBV) + Diffuse large B cell lymphoma (DLBCL), not otherwise specified (NOS). Traces of EBV transcripts were described in lymphomas, including DLBCL, that were diagnosed as EBV negative by conventional methods. The aim of this study was to detect viral genome by qPCR, as well as LMP1 and EBNA2 transcripts, with a more sensitive method in DLBCL cases from Argentina. Fourteen cases originally considered as EBV negative expressed LMP1 and/or EBNA2 transcripts. In addition, LMP1 and/or EBNA2 transcripts were also observed in bystander cells. However, EBERs+ cells cases by conventional ISH showed higher numbers of cells with LMP1 transcripts and LMP1 protein. In the cases that were EBERS- in tumor cells but with expression of LMP1 and/or EBNA2 transcripts, the viral load was below the limit of detection. This study provides further evidence that EBV could be detected in tumor cells by more sensitive methods. However, higher expression of the most important oncogenic protein, LMP1, as well as increased viral load, are only observed in cases with EBERs+ cells by conventional ISH, suggesting that traces of EBV might not display a key role in DLBCL pathogenesis.
Li, K;Shi, Y;Gonye, EC;Bayliss, DA;
PMID: 34732535 | DOI: 10.1523/ENEURO.0212-21.2021
Select neuronal populations display steady rhythmic neuronal firing that provides tonic excitation to drive downstream networks and behaviors. In noradrenergic neurons of the locus coeruleus (LC), circadian neurons of the suprachiasmatic nucleus (SCN), and CO2/H+-activated neurons of the brainstem retrotrapezoid nucleus (RTN), large subthreshold membrane potential oscillations contribute to the pacemaker-like action potential discharge. The oscillations and firing in LC and SCN involve contributions from leak sodium (NALCN) and L-type calcium channels while recent work from RTN suggested an additional pivotal role for a secondary calcium-activated and voltage-gated cationic current sensitive to TRPM4 channel blockers. Here, we tested whether TRPM4 contributes to subthreshold oscillations in mouse LC and SCN. By RNAscope in situ hybridization, Trpm4 transcripts were detected in both cell groups. In whole-cell recordings from acute slice preparations, prominent voltage-dependent membrane potential oscillations were revealed in LC and SCN after blocking action potentials. These oscillations were inhibited by two chemically-distinct blockers of TRPM4, 9-phenanthrol (9-pt) and 4-chloro-2-[[2-(2-chlorophenoxy)acetyl]amino]benzoic acid (CBA). Under whole-cell voltage clamp, inward currents evoked by oscillation voltage waveforms were inhibited in LC by blocking L-type calcium channels and TRPM4. These data implicate TRPM4 in the large subthreshold membrane potential oscillations that underlie tonic action potential discharge in LC and SCN, providing a voltage-dependent and calcium-dependent cationic current to augment the depolarizing inward Na+ and Ca2+ currents previously associated with this distinctive electroresponsive property.
Shi, Z;Stornetta, DS;Stornetta, RL;Brooks, VL;
PMID: 34937769 | DOI: 10.1523/ENEURO.0404-21.2021
The arcuate nucleus (ArcN) is an integrative hub for the regulation of energy balance, reproduction, and arterial pressure (AP), all of which are influenced by Angiotensin II (AngII); however, the cellular mechanisms and downstream neurocircuitry are unclear. Here we show that ArcN AngII increases AP in female rats via two phases, both of which are mediated via activation of AngII type 1 receptors (AT1aR): initial vasopressin-induced vasoconstriction, followed by slowly developing increases in sympathetic nerve activity (SNA) and heart rate (HR). In male rats, ArcN AngII evoked a similarly slow increase in SNA, but the initial pressor response was variable. In females, the effects of ArcN AngII varied during the estrus cycle, with significant increases in SNA, HR, and AP occurring during diestrus and estrus, but only increased AP during proestrus. Pregnancy markedly increased the expression of AT1aR in the ArcN with parallel substantial AngII-induced increases in SNA and MAP. In both sexes, the sympathoexcitation relied on suppression of tonic ArcN sympathoinhibitory Neuropeptide Y inputs, and activation of pro-opiomelanocortin (POMC) projections, to the paraventricular nucleus (PVN). Few or no NPY or POMC neurons expressed the AT1aR, suggesting that AngII increases AP and SNA at least in part indirectly via local interneurons, which express tyrosine hydroxylase (TH) and VGat (i.e. GABAergic). ArcN TH neurons release GABA locally, and central AT1aR and TH neurons mediate stress responses; therefore, we propose that TH AT1aR neurons are well situated to locally coordinate the regulation of multiple modalities within the ArcN in response to stress.SIGNIFICANCEThe arcuate nucleus (ArcN) is an integrative hub for the regulation of energy balance, reproduction, and arterial pressure (AP), all of which are influenced by Angiotensin II (AngII). Here we show that ArcN AngII activates AT1aR to increase AP in male and female rats by slowly increasing sympathetic nerve activity. In females, ArcN AngII also evoked an initial pressor response mediated by vasopressin-induced vasoconstriction. Pregnant and estrus females responded more than males, in association with higher ArcN AT1aR expression. AT1aR were identified in ArcN interneurons that express tyrosine hydroxylase (TH) and GABA. Since brain AT1aR and TH mediate stress responses, ArcN AT1aR TH neurons are well situated to locally coordinate autonomic, hormonal, and behavioral responses to stress.
Exp Mol Pathol. 2014 Apr 1;96(3):310-315
Cimino PJ, Zhao G, Wang D, Sehn JK, Lewis JS, Duncavage EJ.
PMID: 24704430 | DOI: 10.1016/j.yexmp.2014.03.010.
Viral pathogens have been implicated in the development of certain cancers including human papillomavirus (HPV) in squamous cell carcinoma and Epstein-Barr virus (EBV) in Burkitt's lymphoma. The significance of viral pathogens in brain tumors is controversial, and human cytomegalovirus (HCMV) has been associated with glioblastoma (GBM) in some but not all studies, making the role of HCMV unclear. In this study we sought to determine if viral pathogen sequences could be identified in an unbiased manner from previously discarded, unmapped, non-human, next-generation sequencing (NGS) reads obtained from targeted oncology, panel-based sequencing of high grade gliomas (HGGs), including GBMs. Twenty one sequential HGG cases were analyzed by a targeted NGS clinical oncology panel containing 151 genes using DNA obtained from formalin-fixed, paraffin-embedded (FFPE) tissue. Sequencing reads that did not map to the human genome (average of 38,000 non-human reads/case (1.9%)) were filtered and low quality reads removed. Extracted high quality reads were then sequentially aligned to the National Center for Biotechnology Information (NCBI) non-redundant nucleotide (nt and nr) databases. Aligned reads were classified based on NCBI taxonomy database and all eukaryotic viral sequences were further classified into viral families. Two viral sequences (both herpesviruses), EBV and Roseolovirus were detected in 5/21 (24%) cases and in 1/21 (5%) cases, respectively. None of the cases had detectable HCMV. Of the five HGG cases with detectable EBV DNA, four had additional material for EBV in situ hybridization (ISH), all of which were negative for expressed viral sequence. Overall, a similar discovery approach using unmapped non-human NGS reads could be used to discover viral sequences in other cancer types.
Vesicular glutamate transporter modulates sex differences in dopamine neuron vulnerability to age-related neurodegeneration
Buck, SA;Steinkellner, T;Aslanoglou, D;Villeneuve, M;Bhatte, SH;Childers, VC;Rubin, SA;De Miranda, BR;O'Leary, EI;Neureiter, EG;Fogle, KJ;Palladino, MJ;Logan, RW;Glausier, JR;Fish, KN;Lewis, DA;Greenamyre, JT;McCabe, BD;Cheetham, CEJ;Hnasko, TS;Freyberg, Z;
PMID: 33909313 | DOI: 10.1111/acel.13365
Age is the greatest risk factor for Parkinson's disease (PD) which causes progressive loss of dopamine (DA) neurons, with males at greater risk than females. Intriguingly, some DA neurons are more resilient to degeneration than others. Increasing evidence suggests that vesicular glutamate transporter (VGLUT) expression in DA neurons plays a role in this selective vulnerability. We investigated the role of DA neuron VGLUT in sex- and age-related differences in DA neuron vulnerability using the genetically tractable Drosophila model. We found sex differences in age-related DA neurodegeneration and its associated locomotor behavior, where males exhibit significantly greater decreases in both DA neuron number and locomotion during aging compared with females. We discovered that dynamic changes in DA neuron VGLUT expression mediate these age- and sex-related differences, as a potential compensatory mechanism for diminished DA neurotransmission during aging. Importantly, female Drosophila possess higher levels of VGLUT expression in DA neurons compared with males, and this finding is conserved across flies, rodents, and humans. Moreover, we showed that diminishing VGLUT expression in DA neurons eliminates females' greater resilience to DA neuron loss across aging. This offers a new mechanism for sex differences in selective DA neuron vulnerability to age-related DA neurodegeneration. Finally, in mice, we showed that the ability of DA neurons to achieve optimal control over VGLUT expression is essential for DA neuron survival. These findings lay the groundwork for the manipulation of DA neuron VGLUT expression as a novel therapeutic strategy to boost DA neuron resilience to age- and PD-related neurodegeneration.
Incerto-thalamic modulation of fear via GABA and dopamine
Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology
Venkataraman, A;Hunter, SC;Dhinojwala, M;Ghebrezadik, D;Guo, J;Inoue, K;Young, LJ;Dias, BG;
PMID: 33864008 | DOI: 10.1038/s41386-021-01006-5
Fear generalization and deficits in extinction learning are debilitating dimensions of Post-Traumatic Stress Disorder (PTSD). Most understanding of the neurobiology underlying these dimensions comes from studies of cortical and limbic brain regions. While thalamic and subthalamic regions have been implicated in modulating fear, the potential for incerto-thalamic pathways to suppress fear generalization and rescue deficits in extinction recall remains unexplored. We first used patch-clamp electrophysiology to examine functional connections between the subthalamic zona incerta and thalamic reuniens (RE). Optogenetic stimulation of GABAergic ZI → RE cell terminals in vitro induced inhibitory post-synaptic currents (IPSCs) in the RE. We then combined high-intensity discriminative auditory fear conditioning with cell-type-specific and projection-specific optogenetics in mice to assess functional roles of GABAergic ZI → RE cell projections in modulating fear generalization and extinction recall. In addition, we used a similar approach to test the possibility of fear generalization and extinction recall being modulated by a smaller subset of GABAergic ZI → RE cells, the A13 dopaminergic cell population. Optogenetic stimulation of GABAergic ZI → RE cell terminals attenuated fear generalization and enhanced extinction recall. In contrast, optogenetic stimulation of dopaminergic ZI → RE cell terminals had no effect on fear generalization but enhanced extinction recall in a dopamine receptor D1-dependent manner. Our findings shed new light on the neuroanatomy and neurochemistry of ZI-located cells that contribute to adaptive fear by increasing the precision and extinction of learned associations. In so doing, these data reveal novel neuroanatomical substrates that could be therapeutically targeted for treatment of PTSD.
McKinnon C, De Snoo ML, Gondard E, Neudorfer C, Chau H, Ngana SG, O'Hara DM, Brotchie JM, Koprich JB, Lozano AM, Kalia LV, Kalia SK
PMID: 32059750 | DOI: 10.1186/s40478-020-0894-0
Parkinson's disease is a progressive neurodegenerative disorder characterised by the accumulation of misfolded ?-synuclein in selected brain regions, including the substantia nigra pars compacta (SNpc), where marked loss of dopaminergic neurons is also observed. Yet, the relationship between misfolded ?-synuclein and neurotoxicity currently remains unclear. As the principal route for degradation of misfolded proteins in mammalian cells, the ubiquitin-proteasome system (UPS) is critical for maintenance of cellular proteostasis. Misfolded ?-synuclein impairs UPS function and contributes to neuronal death in vitro. Here, we examine its effects in vivo using adeno-associated viruses to co-express A53T ?-synuclein and the ubiquitinated reporter protein UbG76V-GFP in rat SNpc. We found that ?-synuclein over-expression leads to early-onset catalytic impairment of the 26S proteasome with associated UPS dysfunction, preceding the onset of behavioural deficits and dopaminergic neurodegeneration. UPS failure in dopaminergic neurons was also associated with selective accumulation of ?-synuclein phosphorylated at the serine 129 residue, which has previously been linked to increased neurotoxicity. Our study highlights a role for ?-synuclein in disturbing proteostasis which may contribute to neurodegeneration in vivo
Liu, Z;Yang, N;Dong, J;Tian, W;Chang, L;Ma, J;Guo, J;Tan, J;Dong, A;He, K;Zhou, J;Cinar, R;Wu, J;Salinas, AG;Sun, L;Kumar, M;Sullivan, BT;Oldham, BB;Pitz, V;Makarious, MB;Ding, J;Kung, J;Xie, C;Hawes, SL;Wang, L;Wang, T;Chan, P;Zhang, Z;Le, W;Chen, S;Lovinger, DM;Blauwendraat, C;Singleton, AB;Cui, G;Li, Y;Cai, H;Tang, B;
PMID: 35715418 | DOI: 10.1038/s41467-022-31168-9
Endocannabinoid (eCB), 2-arachidonoyl-glycerol (2-AG), the most abundant eCB in the brain, regulates diverse neural functions. Here we linked multiple homozygous loss-of-function mutations in 2-AG synthase diacylglycerol lipase β (DAGLB) to an early onset autosomal recessive Parkinsonism. DAGLB is the main 2-AG synthase in human and mouse substantia nigra (SN) dopaminergic neurons (DANs). In mice, the SN 2-AG levels were markedly correlated with motor performance during locomotor skill acquisition. Genetic knockdown of Daglb in nigral DANs substantially reduced SN 2-AG levels and impaired locomotor skill learning, particularly the across-session learning. Conversely, pharmacological inhibition of 2-AG degradation increased nigral 2-AG levels, DAN activity and dopamine release and rescued the locomotor skill learning deficits. Together, we demonstrate that DAGLB-deficiency contributes to the pathogenesis of Parkinsonism, reveal the importance of DAGLB-mediated 2-AG biosynthesis in nigral DANs in regulating neuronal activity and dopamine release, and suggest potential benefits of 2-AG augmentation in alleviating Parkinsonism.
