Nguyen MQ, Wu Y, Bonilla LS, von Buchholtz LJ, Ryba NJP.
PMID: 28957441 | DOI: 10.1371/journal.pone.0185543
The trigeminal ganglion contains somatosensory neurons that detect a range of thermal, mechanical and chemical cues and innervate unique sensory compartments in the head and neck including the eyes, nose, mouth, meninges and vibrissae. We used single-cell sequencing and in situ hybridization to examine the cellular diversity of the trigeminal ganglion in mice, defining thirteen clusters of neurons. We show that clusters are well conserved in dorsal root ganglia suggesting they represent distinct functional classes of somatosensory neurons and not specialization associated with their sensory targets. Notably, functionally important genes (e.g. the mechanosensory channel Piezo2 and the capsaicin gated ion channel Trpv1) segregate into multiple clusters and often are expressed in subsets of cells within a cluster. Therefore, the 13 genetically-defined classes are likely to be physiologically heterogeneous rather than highly parallel (i.e., redundant) lines of sensory input. Our analysis harnesses the power of single-cell sequencing to provide a unique platform for in silico expression profiling that complements other approaches linking gene-expression with function and exposes unexpected diversity in the somatosensory system.
Developmental dynamics : an official publication of the American Association of Anatomists
Kasemeier-Kulesa, JC;Morrison, JA;McKinney, S;Li, H;Gogol, M;Hall, K;Chen, S;Wang, Y;Perera, A;McLennan, R;Kulesa, PM;
PMID: 36840366 | DOI: 10.1002/dvdy.577
The molecular identification of neural progenitor cell populations that connect to establish the sympathetic nervous system (SNS) remains unclear. This is due to technical limitations in the acquisition and spatial mapping of molecular information to tissue architecture.To address this, we applied Slide-seq spatial transcriptomics to intact fresh frozen chick trunk tissue transversely cryo-sectioned at the developmental stage prior to SNS formation. In parallel, we performed age- and location-matched single cell (sc) RNA-seq and 10× Genomics Visium to inform our analysis. Downstream bioinformatic analyses led to the unique molecular identification of neural progenitor cells within the peripheral sympathetic ganglia (SG) and spinal cord preganglionic neurons (PGNs). We then successfully applied the HiPlex RNAscope fluorescence in situ hybridization and multispectral confocal microscopy to visualize 12 gene targets in stage-, age- and location-matched chick trunk tissue sections.Together, these data demonstrate a robust strategy to acquire and integrate single cell and spatial transcriptomic information, resulting in improved resolution of molecular heterogeneities in complex neural tissue architectures. Successful application of this strategy to the developing SNS provides a roadmap for functional studies of neural connectivity and platform to address complex questions in neural development and regeneration.
Ma, S;Skarica, M;Li, Q;Xu, C;Risgaard, RD;Tebbenkamp, ATN;Mato-Blanco, X;Kovner, R;Krsnik, Ž;de Martin, X;Luria, V;Martí-Pérez, X;Liang, D;Karger, A;Schmidt, DK;Gomez-Sanchez, Z;Qi, C;Gobeske, KT;Pochareddy, S;Debnath, A;Hottman, CJ;Spurrier, J;Teo, L;Boghdadi, AG;Homman-Ludiye, J;Ely, JJ;Daadi, EW;Mi, D;Daadi, M;Marín, O;Hof, PR;Rasin, MR;Bourne, J;Sherwood, CC;Santpere, G;Girgenti, MJ;Strittmatter, SM;Sousa, AMM;Sestan, N;
PMID: 36007006 | DOI: 10.1126/science.abo7257
The granular dorsolateral prefrontal cortex (dlPFC) is an evolutionary specialization of primates that is centrally involved in cognition. Here, we assessed over 600,000 single-nucleus transcriptomes from adult human, chimpanzee, macaque, and marmoset dlPFC. While most transcriptomically-defined cell subtypes are conserved, we detected several only in some species and substantial species-specific molecular differences across homologous neuronal, glial and non-neural subtypes. The latter are exemplified by human-specific switching between expression of the neuropeptide somatostatin (SST) and tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine production, in certain interneurons, and also by expression of the neuropsychiatric risk gene FOXP2, which is human-specific in microglia and primate-specific in layer-4 granular neurons. We generated a comprehensive survey of dlPFC cellular repertoire and its shared and divergent features in anthropoid primates.
Khan S, Stott S, Chabrat A, Truckenbrodt AM, Spencer-Dene B, Nave KA, Guillemot F, Levesque M, Ang SL.
PMID: 28130357 | DOI: 10.1523/JNEUROSCI.2414-16.2016
Abstract
Midbrain dopaminergic neurons are highly heterogeneous. They differ in their connectivity and firing patterns, and therefore in their functional properties. The molecular underpinnings of this heterogeneity are largely unknown and there is a paucity of markers that distinguish these functional subsets. In this paper, we report the identification and characterisation of a novel subset of midbrain dopaminergic neurons located in the ventral tegmental area that expresses the basic helix-loop-helix transcription factor, Neurogenic Differentiation Factor-6 (NEUROD6). Retrograde fluorogold tracing experiments demonstrate that Neurod6+ mDA neurons project to two distinct septal regions, the dorsal lateral and intermediate region of the lateral septum. Loss-of-function studies in mice demonstrate that Neurod6 and the closely related family member Neurod1 are both specifically required for the survival of this lateral-septum projecting neuronal subset during development. Our findings underscore the complex organisation of midbrain dopaminergic neurons and provide an entry point for future studies of the functions of the Neurod6+ subset of midbrain dopaminergic neurons.
SIGNIFICANCE STATEMENT:
Midbrain dopaminergic neurons regulate diverse brain functions, including voluntary movement and cognitive and emotive behaviours. These neurons are heterogeneous and distinct subsets are thought to regulate different behaviours. However, we currently lack the means to identify and modify gene function in specific subsets of midbrain dopaminergic neurons. In this study, we identify the transcription factor NEUROD6 as a specific marker for a novel subset of midbrain dopaminergic neurons in the ventral midbrain that project to the lateral septum and we reveal essential roles for Neurod1 and Neurod6 in the survival of these neurons during development. Our findings highlight the molecular and anatomical heterogeneity of midbrain dopaminergic neurons and contribute to a better understanding of this functionally complex group of neurons.
VGLUT2 is a determinant of dopamine neuron resilience in a rotenone model of dopamine neurodegeneration
The Journal of neuroscience : the official journal of the Society for Neuroscience
Buck, SA;Miranda, BR;Logan, RW;Fish, KN;Greenamyre, JT;Freyberg, Z;
PMID: 33893220 | DOI: 10.1523/JNEUROSCI.2770-20.2021
Parkinson's disease (PD) is characterized by progressive dopamine (DA) neuron loss in the substantia nigra pars compacta (SNc). In contrast, DA neurons in the ventral tegmental area (VTA) are relatively protected from neurodegeneration, but the underlying mechanisms for this resilience remain poorly understood. Recent work suggests that expression of the vesicular glutamate transporter 2 (VGLUT2) selectively impacts midbrain DA neuron vulnerability. We investigated whether altered DA neuron VGLUT2 expression determines neuronal resilience in rats exposed to rotenone, a mitochondrial complex I inhibitor and toxicant model of PD. We discovered that VTA/SNc DA neurons that expressed VGLUT2 are more resilient to rotenone-induced DA neurodegeneration. Surprisingly, the density of neurons with detectable VGLUT2 expression in the VTA and SNc increases in response to rotenone. Furthermore, dopaminergic terminals within the nucleus accumbens, where the majority of VGLUT2-expressing DA neurons project, exhibit greater resilience compared to DA terminals in the caudate/putamen. More broadly, VGLUT2-expressing terminals are protected throughout the striatum from rotenone-induced degeneration. Together, our data demonstrate that a distinct subpopulation of VGLUT2-expressing DA neurons are relatively protected from rotenone neurotoxicity. Rotenone-induced upregulation of the glutamatergic machinery in VTA and SNc neurons and their projections may be part of a broader neuroprotective mechanism. These findings offer a putative new target for neuronal resilience that can be manipulated to prevent toxicant-induced DA neurodegeneration in PD.SIGNIFICANCE STATEMENT:Environmental exposures to pesticides contribute significantly to pathological processes that culminate in Parkinson's disease (PD). The pesticide rotenone has been used to generate a PD model that replicates key features of the illness including dopamine neurodegeneration. To date, longstanding questions remain: are there dopamine neuron subpopulations resilient to rotenone, and if so, what are the molecular determinants of this resilience? Here we show that the subpopulation of midbrain dopaminergic neurons that express the vesicular glutamate transporter 2 (VGLUT2) are more resilient to rotenone-induced neurodegeneration. Rotenone also upregulates VGLUT2 more broadly in the midbrain, suggesting VGLUT2 expression generally confers increased resilience to rotenone. VGLUT2 may therefore be a new target for boosting neuronal resilience to prevent toxicant-induced DA neurodegeneration in PD.
Choi E, Lantz TL, Vlacich G, Keeley TM, Samuelson LC, Coffey RJ, Goldenring JR, Powell AE.
PMID: 28814482 | DOI: 10.1136/gutjnl-2017-313874
Abstract
OBJECTIVE:
Lrig1 is a marker of proliferative and quiescent stem cells in the skin and intestine. We examined whether Lrig1-expressing cells are long-lived gastric progenitors in gastric glands in the mouse stomach. We also investigated how the Lrig1-expressing progenitor cells contribute to the regeneration of normal gastric mucosa by lineage commitment to parietal cells after acute gastric injury in mice.
DESIGN:
We performed lineage labelling using Lrig1-CreERT2/+;R26R-YFP/+ (Lrig1/YFP) or R26R-LacZ/+ (Lrig1/LacZ) mice to examine whether the Lrig1-YFP-marked cells are gastric progenitor cells. We studied whether Lrig1-YFP-marked cells give rise to normal gastric lineage cells in damaged mucosa using Lrig1/YFP mice after treatment with DMP-777 to induce acute injury. We also studied Lrig1-CreERT2/CreERT2 (Lrig1 knockout) mice to examine whether the Lrig1 protein is required for regeneration of gastric corpus mucosa after acute injury.
RESULTS:
Lrig1-YFP-marked cells give rise to gastric lineage epithelial cells both in the gastric corpus and antrum, in contrast to published results that Lgr5 only marks progenitor cells within the gastric antrum. Lrig1-YFP-marked cells contribute to replacement of damaged gastric oxyntic glands during the recovery phase after acute oxyntic atrophy in the gastric corpus. Lrig1 null mice recovered normally from acute gastric mucosal injury indicating that Lrig1 protein is not required for lineage differentiation. Lrig1+ isthmal progenitor cells did not contribute to transdifferentiating chief cell lineages after acute oxyntic atrophy.
CONCLUSIONS:
Lrig1 marks gastric corpus epithelial progenitor cells capable of repopulating the damaged oxyntic mucosa by differentiating into normal gastric lineage cells in mouse stomach.
Neurogastroenterol Motil. 2018 Oct 12:e13479.
Russell JP, Mohammadi E, Ligon C, Latorre R, Johnson AC, Hoang B, Krull D, Ho MW, Eidam HS, DeMartino MP, Cheung M, Oliff AI, Kumar S, Greenwood-Van Meerveld B.
PMID: 30311722 | DOI: 10.1111/nmo.13479
Abstract BACKGROUND: The expression of RET in the developing enteric nervous system (ENS) suggests that RET may contribute to adult intestinal function. ENS cholinergic nerves play a critical role in the control of colonic function through the release of acetylcholine (ACh). In the current study, we hypothesized that a RET-mediated mechanism may regulate colonic ion transport and motility through modulation of cholinergic nerves. METHODS: The effect of RET inhibition on active ion transport was assessed electrophysiologically in rat colonic tissue mounted in Ussing chambers via measurements of short circuit current (Isc) upon electrical field stimulation (EFS) or pharmacologically with cholinergic agonists utilizing a gastrointestinal (GI)-restricted RET inhibitor. We assessed the effect of the RET inhibitor on propulsive motility via quantification of fecal pellet output (FPO) induced by the acetylcholinesterase inhibitor neostigmine. KEY RESULTS: We found that enteric ganglia co-expressed RET and choline acetyltransferase (ChAT) transcripts. In vitro, the RET kinase inhibitor GSK3179106 attenuated the mean increase in Isc induced by either EFS or carbachol but not bethanechol. In vivo, GSK3179106 significantly reduced the prokinetic effect of neostigmine. CONCLUSION AND INFERENCES: Our findings provide evidence that RET-mediated mechanisms regulate colonic function by maintaining cholinergic neuronal function and enabling ACh-evoked chloride secretion and motility. We suggest that modulating the cholinergic control of the colon via a RET inhibitor may represent a novel target for the treatment of intestinal disorders associated with increased secretion and accelerated GI transit such as irritable bowel syndrome with diarrhea (IBS-D).
Proc Natl Acad Sci U S A. 2014 Nov 3.
Zhang HY, Gao M, Liu QR, Bi GH, Li X, Yang HJ, Gardner EL, Wu J, Xi ZX.
PMID: 25368177 | DOI: 201413210
Cannabinoid CB2 receptors (CB2Rs) have been recently reported to modulate brain dopamine (DA)-related behaviors; however, the cellular mechanisms underlying these actions are unclear. Here we report that CB2Rs are expressed in ventral tegmental area (VTA) DA neurons and functionally modulate DA neuronal excitability and DA-related behavior. In situ hybridization and immunohistochemical assays detected CB2 mRNA and CB2R immunostaining in VTA DA neurons. Electrophysiological studies demonstrated that activation of CB2Rs by JWH133 or other CB2R agonists inhibited VTA DA neuronal firing in vivo and ex vivo, whereas microinjections of JWH133 into the VTA inhibited cocaine self-administration. Importantly, all of the above findings observed in WT or CB1 -/- mice are blocked by CB2R antagonist and absent in CB2 -/- mice. These data suggest that CB2R-mediated reduction of VTA DA neuronal activity may underlie JWH133's modulation of DA-regulated behaviors.
Ortega-de San Luis C, Sanchez-Garcia MA, Nieto-Gonzalez JL, García-Junco-Clemente P, Montero-Sanchez A, Fernandez-Chacon R, Pascual A.
PMID: 30058223 | DOI: 10.1111/acel.12821
The striatum integrates motor behavior using a well-defined microcircuit whose individual components are independently affected in several neurological diseases. The glial cell line-derived neurotrophic factor (GDNF), synthesized by striatal interneurons, and Sonic hedgehog (Shh), produced by the dopaminergic neurons of the substantia nigra (DA SNpc), are both involved in the nigrostriatal maintenance but the reciprocal neurotrophic relationships among these neurons are only partially understood. To define the postnatal neurotrophic connections among fast-spiking GABAergic interneurons (FS), cholinergic interneurons (ACh), and DA SNpc, we used a genetically induced mouse model of postnatal DA SNpc neurodegeneration and separately eliminated Smoothened (Smo), the obligatory transducer of Shh signaling, in striatal interneurons. We show that FS postnatal survival relies on DA SNpc and is independent of Shh signaling. On the contrary, Shh signaling but not dopaminergic striatal innervation is required to maintain ACh in the postnatal striatum. ACh are required for DA SNpc survival in a GDNF-independent manner. These data demonstrate the existence of three parallel but interdependent neurotrophic relationships between SN and striatal interneurons, partially defined by Shh and GDNF. The definition of these new neurotrophic interactions opens the search for new molecules involved in the striatal modulatory circuit maintenance with potential therapeutic value.
Pancani, T;Day, M;Tkatch, T;Wokosin, DL;González-Rodríguez, P;Kondapalli, J;Xie, Z;Chen, Y;Beaumont, V;Surmeier, DJ;
PMID: 36914640 | DOI: 10.1038/s41467-023-36556-3
Huntington's disease (HD) is a progressive, neurodegenerative disease caused by a CAG triplet expansion in huntingtin. Although corticostriatal dysfunction has long been implicated in HD, the determinants and pathway specificity of this pathophysiology are not fully understood. Here, using a male zQ175+/- knock-in mouse model of HD we carry out optogenetic interrogation of intratelencephalic and pyramidal tract synapses with principal striatal spiny projection neurons (SPNs). These studies reveal that the connectivity of intratelencephalic, but not pyramidal tract, neurons with direct and indirect pathway SPNs increased in early symptomatic zQ175+/- HD mice. This enhancement was attributable to reduced pre-synaptic inhibitory control of intratelencephalic terminals by striatal cholinergic interneurons. Lowering mutant huntingtin selectively in striatal cholinergic interneurons with a virally-delivered zinc finger repressor protein normalized striatal acetylcholine release and intratelencephalic functional connectivity, revealing a node in the network underlying corticostriatal pathophysiology in a HD mouse model.
Brain : a journal of neurology
Wlaschin, JJ;Donahue, C;Gluski, J;Osborne, JF;Ramos, LM;Silberberg, H;Le Pichon, CE;
PMID: 36342754 | DOI: 10.1093/brain/awac415
Amyotrophic lateral sclerosis or ALS is a devastating and fatal neurodegenerative disease of motor neurons with very few treatment options. We had previously found that motor neuron degeneration in a mouse model of ALS can be delayed by deleting the axon damage sensor MAP3K12 or Dual Leucine Zipper Kinase (DLK)1. However, DLK is also involved in axon regeneration2-5, prompting us to ask whether combining DLK deletion with a way to promote axon regeneration would result in greater motor neuron protection. To achieve this, we used a mouse line that constitutively expresses ATF3, a master regulator of regeneration in neurons6,7. Although there is precedence for each individual strategy in the SOD1G93A mouse model of ALS1,8, these have not previously been combined. By several lines of evidence including motor neuron electrophysiology, histology and behavior, we observed a powerful synergy when combining DLK deletion with ATF3 expression. The combinatorial strategy resulted in significant protection of motor neurons with fewer undergoing cell death, reduced axon degeneration, and preservation of motor function and connectivity to muscle. This study provides a demonstration of the power of combinatorial therapy to treat neurodegenerative disease.
Pandey, M;Zhang, JH;Adikaram, PR;Kittock, CM;Lue, N;Awe, AM;Degner, KN;Jacob, N;Staples, JN;Thomas, R;Kohnen, AB;Ganesan, S;Kabat, J;Chen, CK;Simonds, WF;
PMID: 37219953 | DOI: 10.1172/jci.insight.134685
Mechanical, thermal, and chemical pain sensation is conveyed by primary nociceptors, a subset of sensory afferent neurons. The intracellular regulation of the primary nociceptive signal is an area of active study. We report here the discovery of a Gβ5-dependent regulatory pathway within mechanical nociceptors that restrains anti-nociceptive input from metabotropic GABA-B receptors. In mice with conditional knockout (cKO) of Gnb5 targeted to peripheral sensory neurons, we demonstrate the impairment of mechanical, thermal, and chemical nociception. We further report the specific loss of mechanical nociception in Rgs7-Cre+/-; Gnb5fl/fl mice but not in Rgs9-Cre+/-; Gnb5fl/fl mice, suggesting that Gβ5 might specifically regulate mechanical pain in Rgs7+ cells. Additionally, Gβ5-dependent and Rgs7-associated mechanical nociception is dependent upon GABA-B receptor signaling since both were abolished by treatment with a GABA-B receptor antagonist and since cKO of Gβ5 from sensory cells or from Rgs7+ cells potentiated the analgesic effects of GABA-B agonists. Following activation by the Mrgprd agonist β-alanine, enhanced sensitivity to inhibition by baclofen was observed in primary cultures of Rgs7+ sensory neurons harvested from Rgs7-Cre+/-; Gnb5fl/fl mice. Taken together, these results suggest that the targeted inhibition of Gβ5 function in Rgs7+ sensory neurons might provide specific relief for mechanical allodynia, including that contributing to chronic neuropathic pain, without reliance on exogenous opioids.
