| 1. |
- Aresh, Bejan, 1984-, et al.
(författare)
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Spinal Cord Interneurons Expressing the Gastrin-Releasing Peptide Receptor Convey Itch Through VGLUT2-Mediated Signaling
- 2017
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Ingår i: Pain. - : Ovid Technologies (Wolters Kluwer Health). - 0304-3959 .- 1872-6623. ; 158:5, s. 945-961
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Tidskriftsartikel (refereegranskat)abstract
- Itch is a sensation that promotes the desire to scratch, which can be evoked by mechanical and chemical stimuli. In the spinal cord, neurons expressing the gastrin-releasing peptide receptor (GRPR) have been identified as specific mediators of itch. However, our understanding of the GRPR population in the spinal cord, and thus how these neurons exercise their functions, is limited. For this purpose, we constructed a Cre line designed to target the GRPR population of neurons (Grpr-Cre). Our analysis revealed that Grpr-Cre cells in the spinal cord are predominantly excitatory interneurons that are found in the dorsal lamina, especially in laminae II-IV. Application of the specific agonist gastrin-releasing peptide induced spike responses in 43.3% of the patched Grpr-Cre neurons, where the majority of the cells displayed a tonic firing property. Additionally, our analysis showed that the Grpr-Cre population expresses Vglut2 mRNA, and mice ablated of Vglut2 in Grpr-Cre cells (Vglut2-lox; Grpr-Cre mice) displayed less spontaneous itch and attenuated responses to both histaminergic and nonhistaminergic agents. We could also show that application of the itch-inducing peptide, natriuretic polypeptide B, induces calcium influx in a subpopulation of Grpr-Cre neurons. To summarize, our data indicate that the Grpr-Cre spinal cord neural population is composed of interneurons that use VGLUT2-mediated signaling for transmitting chemical and spontaneous itch stimuli to the next, currently unknown, neurons in the labeled line of itch.
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| 2. |
- Vieillard, Jennifer, et al.
(författare)
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Adult spinal Dmrt3 neurons receive direct somatosensory inputs from ipsi- and contralateral primary afferents and from brainstem motor nuclei
- 2023
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Ingår i: Journal of Comparative Neurology. - : John Wiley & Sons. - 0021-9967 .- 1096-9861. ; 531:1, s. 5-24
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Tidskriftsartikel (refereegranskat)abstract
- In the spinal cord, sensory-motor circuits controlling motor activity are situated in the dorso-ventral interface. The neurons identified by the expression of the transcription factor Doublesex and mab-3 related transcription factor 3 (Dmrt3) have previously been associated with the coordination of locomotion in horses (Equus caballus, Linnaeus, 1758), mice (Mus musculus, Linnaeus, 1758), and zebrafish (Danio rerio, F. Hamilton, 1822). Based on earlier studies, we hypothesized that, in mice, these neurons may be positioned to receive sensory and central inputs to relay processed commands to motor neurons. Thus, we investigated the presynaptic inputs to spinal Dmrt3 neurons using monosynaptic retrograde replication-deficient rabies tracing. The analysis showed that lumbar Dmrt3 neurons receive inputs from intrasegmental neurons, and intersegmental neurons from the cervical, thoracic, and sacral segments. Some of these neurons belong to the excitatory V2a interneurons and to plausible Renshaw cells, defined by the expression of Chx10 and calbindin, respectively. We also found that proprioceptive primary sensory neurons of type Ia2, Ia3, and Ib, defined by the expression of calbindin, calretinin, and Brn3c, respectively, provide presynaptic inputs to spinal Dmrt3 neurons. In addition, we demonstrated that Dmrt3 neurons receive inputs from brain areas involved in motor regulation, including the red nucleus, primary sensory-motor cortex, and pontine nuclei. In conclusion, adult spinal Dmrt3 neurons receive inputs from motor-related brain areas as well as proprioceptive primary sensory neurons and have been shown to connect directly to motor neurons. Dmrt3 neurons are thus positioned to provide sensory-motor control and their connectivity is suggestive of the classical reflex pathways present in the spinal cord.
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| 3. |
- Bjarnadóttir, Thóra K., et al.
(författare)
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The human and mouse repertoire of the adhesion family of G-protein-coupled receptors
- 2004
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Ingår i: Genomics. - : Elsevier BV. - 0888-7543 .- 1089-8646. ; 84:1, s. 23-33
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Tidskriftsartikel (refereegranskat)abstract
- The adhesion G-protein-coupled receptors (GPCRs) (also termed LN-7TM or EGF-7TM receptors) are membrane-bound proteins with long N-termini containing multiple domains. Here, 2 new human adhesion-GPCRs, termed GPR133 and GPR144, have been found by searches done in the human genome databases. Both GPR133 and GPR144 have a GPS domain in their N-termini, while GPR144 also has a pentraxin domain. The phylogenetic analyses of the 2 new human receptors show that they group together without close relationship to the other adhesion-GPCRs. In addition to the human genes, mouse orthologues to those 2 and 15 other mouse orthologues to human were identified (GPR110, GPR111, GPR112, GPR113, GPR114, GPR115, GPR116, GPR123, GPR124, GPR125, GPR126, GPR128, LEC1, LEC2, and LEC3). Currently the total number of human adhesion-GPCRs is 33. The mouse and human sequences show a clear one-to-one relationship, with the exception of EMR2 and EMR3, which do not seem to have orthologues in mouse. EST expression charts for the entire repertoire of adhesion-GPCRs in human and mouse were established. Over 1600 ESTs were found for these receptors, showing widespread distribution in both central and peripheral tissues. The expression patterns are highly variable between different receptors, indicating that they participate in a number of physiological processes.
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| 4. |
- Caruso, Vanni, et al.
(författare)
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Synaptic changes induced by melanocortin signalling
- 2014
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Ingår i: Nature Reviews Neuroscience. - : Springer Science and Business Media LLC. - 1471-003X .- 1471-0048. ; 15:2, s. 98-110
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Forskningsöversikt (refereegranskat)abstract
- The melanocortin system has a well-established role in the regulation of energy homeostasis, but there is growing evidence of its involvement in memory, nociception, mood disorders and addiction. In this Review, we focus on the role of the melanocortin 4 receptor and provide an integrative view of the molecular mechanisms that lead to melanocortin-induced changes in synaptic plasticity within these diverse physiological systems. We also highlight the importance of melanocortin peptides and receptors in chronic pain syndromes, memory impairments, depression and drug abuse, and the possibility of targeting them for therapeutic purposes.
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| 5. |
- Ceder, Mikaela M., et al.
(författare)
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The glycine receptor alpha 3 subunit mRNA expression shows sex-dependent differences in the adult mouse brain
- 2023
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Ingår i: BMC Neuroscience. - : Springer Nature. - 1471-2202. ; 24:1
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Tidskriftsartikel (refereegranskat)abstract
- Background The glycinergic system plays an important inhibitory role in the mouse central nervous system, where glycine controls the excitability of spinal itch- and pain-mediating neurons. Impairments of the glycine receptors can cause motor and sensory deficits. Glycine exerts inhibition through interaction with ligand-gated ion channels composed of alpha and beta subunits. We have investigated the mRNA expression of the glycine receptor alpha 3 (Glra3) subunit in the nervous system as well as in several peripheral organs of female and male mice.Results Single-cell RNA sequencing (scRNA-seq) data analysis on the Zeisel et al. (2018) dataset indicated widespread but low expression of Glra3 in vesicular glutamate transporter 2 (Vglut2, Slc17a6) positive and vesicular inhibitory amino acid transporter (Viaat, Slc32a1)positive neurons of the mouse central nervous system. Highest occurrence of Glra3 expression was identified in the cortex, amygdala, and striatal regions, as well as in the hypothalamus, brainstem and spinal cord. Bulk quantitative real-time-PCR (qRT-PCR) analysis demonstrated Glra3 expression in cortex, amygdala, striatum, hypothalamus, thalamus, pituitary gland, hippocampus, cerebellum, brainstem, and spinal cord. Additionally, male mice expressed higher levels of Glra3 in all investigated brain areas compared with female mice. Lastly, RNAscope spatially validated Glra3 expression in the areas indicated by the single-cell and bulk analyses. Moreover, RNAscope analysis confirmed co-localization of Glra3 with Slc17a6 or Slc32a1 in the central nervous system areas suggested from the single-cell data.Conclusions Glra3 expression is low but widespread in the mouse central nervous system. Clear sex-dependent differences have been identified, indicating higher levels of Glra3 in several telencephalic and diencephalic areas, as well as in cerebellum and brainstem, in male mice compared with female mice.
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| 6. |
- Dondalska, Aleksandra, et al.
(författare)
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Amelioration of Compound 48/80-Mediated Itch and LL-37-Induced Inflammation by a Single-Stranded Oligonucleotide
- 2020
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Ingår i: Frontiers in Immunology. - : FRONTIERS MEDIA SA. - 1664-3224. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Numerous inflammatory skin disorders display a high prevalence of itch. The Mas-related G protein coupled receptor X2 (MRGPRX2) has been shown to modulate itch by inducing non-IgE-mediated mast cell degranulation and the release of endogenous inducers of pruritus. Various substances collectively known as basic secretagogues, which include inflammatory peptides and certain drugs, can trigger MRGPRX2 and thereby induce pseudo-allergic reactions characterized by histamine and protease release as well as inflammation. Here, we investigated the capacity of an immunomodulatory single-stranded oligonucleotide (ssON) to modulate IgE-independent mast cell degranulation and, more specifically, its ability to inhibit the basic secretagogues compound 48/80 (C48/80)-and LL-37in vitroandin vivo. We examined the effect of ssON on MRGPRX2 activationin vitroby measuring degranulation in a human mast cell line (LAD2) and calcium influx in MRGPRX2-transfected HEK293 cells. To determine the effect of ssON on itch, we performed behavioral studies in established mouse models and collected skin biopsies for histological analysis. Additionally, with the use of a rosacea mouse model and RT-qPCR, we investigated the effect on ssON on LL-37-induced inflammation. We reveal that both mast cell degranulation and calcium influx in MRGPRX2 transfected HEK293 cells, induced by the antimicrobial peptide LL-37 and the basic secretagogue C48/80, are effectively inhibited by ssON in a dose-dependent manner. Further, ssON demonstrates a capability to inhibit LL-37 and C48/80 activationin vivoin two mouse models. We show that intradermal injection of ssON in mice is able to block itch induced via C48/80 in a dose-dependent manner. Histological staining revealed that ssON inhibits acute mast cell degranulation in murine skin treated with C48/80. Lastly, we show that ssON treatment ameliorates LL-37-induced inflammation in a rosacea mouse model. Since there is a need for new therapeutics targeting non-IgE-mediated activation of mast cells, ssON could be used as a prospective drug candidate to resolve itch and inflammation in certain dermatoses.
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| 7. |
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| 8. |
- Franck, Marina Christina Mikaela, et al.
(författare)
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Urocortin3-expressing neurons in sensory transmission
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Urocortin 3 (UCN3) is a neuropeptide involved in mechanosensation and stress regulation, and Ucn3-Cre neurons have been assigned a role in mechanical itch. Here, we show that Ucn3 marks a population of excitatory neurons in the mouse dorsal horn, divided into two non-overlapping subpopulations expressing protein kinase C g or calretinin/calbindin 2. Chemogenetic activation of spinal Ucn3-Cre neurons evoked a targeted biting/licking behavior towards the corresponding dermatome. Genetic deletion of vesicular glutamate transporter 2 (VGLUT2) in Ucn3-Cre neurons removed the phenotype, showing that the biting/licking behavior is VGLUT2-dependent. Conditional deletion of VGLUT2 did not affect acute thermal or mechanical withdrawal responses, nor thermal withdrawal responses after nerve growth factor-induced hypersensitivity or the prurifensive response to 48/80 or von Frey stimuli applied in nape. Instead, we found that a group of spinal Ucn3 neurons were activated in response to artificial scratching or 48/80-induced itch. Electrophysiological experiments showed that spinal Ucn3 neurons received both glycinergic and GABAergic tonic inhibition, and monosynaptic inputs from both Aβ and C fibers, which could be confirmed by rabies tracing. Spinal Ucn3/Ucn3-Cre neurons thus represent a mechanically sensitive population with several roles in the itch-scratch cycle.
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| 9. |
- Fredriksson, Robert, et al.
(författare)
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Novel human G protein-coupled receptors with long N-terminals containing GPS domains and Ser/Thr-rich regions
- 2002
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Ingår i: FEBS Letters. - 0014-5793 .- 1873-3468. ; 531:3, s. 407-414
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Tidskriftsartikel (refereegranskat)abstract
- We report eight novel members of the superfamily of human G protein-coupled receptors (GPCRs) found by searches in the human genome databases, termed GPR97, GPR110, GPR111, GPR112, GPR113, GPR114, GPR115 and GPR116. Phylogenetic analysis shows that these are additional members of a family of GPCRs with long N-termini, previously termed EGF-7TM, LNB-7TM, B2 or LN-7TM. Five of the receptors form their own phylogenetic cluster, while three others form a cluster with the previously reported HE6 and GPR56 (TM7XN1). All the receptors have a GPS domain in their N-terminus and long Ser/Thr-rich regions forming mucin-like stalks. GPR113 has a hormone binding domain and one EGF domain. GPR112 has over 20 Ser/Thr repeats and a pentraxin domain. GPR116 has two immunoglobulin-like repeats and a SEA box. We found several human EST sequences for most of the receptors showing differential expression patterns, which may indicate that some of these receptors participate in reproductive functions while others are more likely to have a role in the immune system.
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| 10. |
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| 11. |
- Fredriksson, Robert, et al.
(författare)
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The G-Protein-Coupled Receptors in the Human Genome Form Five Main Families : Phylogenetic Analysis, Paralogon Groups, and Fingerprints
- 2003
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Ingår i: Molecular Pharmacology. - : American Society for Pharmacology & Experimental Therapeutics (ASPET). - 0026-895X .- 1521-0111. ; 63:6, s. 1256-1272
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Tidskriftsartikel (refereegranskat)abstract
- The superfamily of G-protein-coupled receptors (GPCRs) is very diverse in structure and function and its members are among the most pursued targets for drug development. We identified more than 800 human GPCR sequences and simultaneously analyzed 342 unique functional nonolfactory human GPCR sequences with phylogenetic analyses. Our results show, with high bootstrap support, five main families, named glutamate, rhodopsin, adhesion, frizzled/taste2, and secretin, forming the GRAFS classification system. The rhodopsin family is the largest and forms four main groups with 13 sub-branches. Positions of the GPCRs in chromosomal paralogons regions indicate the importance of tetraploidizations or local gene duplication events for their creation. We also searched for "fingerprint" motifs using Hidden Markov Models delineating the putative inter-relationship of the GRAFS families. We show several common structural features indicating that the human GPCRs in the GRAFS families share a common ancestor. This study represents the first overall map of the GPCRs in a single mammalian genome. Our novel approach of analyzing such large and diverse sequence sets may be useful for studies on GPCRs in other genomes and divergent protein families.
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| 12. |
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| 13. |
- Freitag, Fabio Batista, et al.
(författare)
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Spinal gastrin releasing peptide receptor expressing interneurons are controlled by local phasic and tonic inhibition
- 2019
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Dorsal horn gastrin-releasing peptide receptor (GRPR) neurons have a central role in itch transmission. Itch signaling has been suggested to be controlled by an inhibitory network in the spinal dorsal horn, as increased scratching behavior can be induced by pharmacological disinhibition or ablation of inhibitory interneurons, but the direct influence of the inhibitory tone on the GRPR neurons in the itch pathway have not been explored. Here we have investigated spinal GRPR neurons through in vitro and bioinformatical analysis. Electrophysiological recordings revealed that GRPR neurons receive local spontaneous excitatory inputs transmitted by glutamate and inhibitory inputs by glycine and GABA, which were transmitted either by separate glycinergic and GABAergic synapses or by glycine and GABA co-releasing synapses. Additionally, all GRPR neurons received both glycine- and GABA-induced tonic currents. The findings show a complex inhibitory network, composed of synaptic and tonic currents that gates the excitability of GRPR neurons, which provides direct evidence for the existence of an inhibitory tone controlling spontaneous discharge in an itch-related neuronal network in the spinal cord. Finally, calcium imaging revealed increased levels of neuronal activity in Grpr-Cre neurons upon application of somatostatin, which provides direct in vitro evidence for disinhibition of these dorsal horn interneurons.
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| 14. |
- Freitag, Fabio B., et al.
(författare)
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Targeting barrel field spiny stellate cells using a vesicular monoaminergic transporter 2-Cre mouse line
- 2021
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Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- Rodent primary somatosensory cortex (S1) is organized in defined layers, where layer IV serves as the main target for thalamocortical projections. Serotoninergic signaling is important for the organization of thalamocortical projections and consequently proper barrel field development in rodents, and the vesicular monoamine transporter 2 (VMAT2) can be detected locally in layer IV S1 cortical neurons in mice as old as P10, but the identity of the Vmat2-expressing neurons is unknown. We here show that Vmat2 mRNA and also Vmat2-Cre recombinase are still expressed in adult mice in a sub-population of the S1 cortical neurons in the barrel field. The Vmat2-Cre cells showed a homogenous intrinsically bursting firing pattern determined by whole-cell patch-clamp, localized radial densely spinous basal dendritic trees and almost exclusively lack of apical dendrite, indicative of layer IV spiny stellate cells. Single cell mRNA sequencing analysis showed that S1 cortical Vmat2-Cre;tdTomato cells express the layer IV marker Rorb and mainly cluster with layer IV neurons, and RNAscope analysis revealed that adult Vmat2-Cre neurons express Vmat2 and vesicular glutamate transporter 1 (Vglut1) and Vglut2 mRNA to a high extent. In conclusion, our analysis shows that cortical Vmat2 expression is mainly confined to layer IV neurons with morphological, electrophysiological and transcriptional characteristics indicative of spiny stellate cells.
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| 15. |
- Freitag, Fabio
(författare)
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Targeting and driving somatosensory neurons
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Pain and itch are two distinct sensations, but the fundamental question of how our nervous system distinguishes the processing and encoding of their related information is still far to be clearly delineated. At the spinal cord level, evidences have pointed out specific groups of neurons expressing the gastrin releasing peptide (Grp) and its receptor (Grpr) as responsible for carrying specifically itch-related information. Such important findings suggest a labeled line for itch and hypothesize the existence of separate pathways transmitting different sensory modalities already at this stage. Aiming at digging further on the pain/itch dualism, the present thesis focused first in addressing the GRPR-expressing dorsal horn interneurons and its roles in itch transmission. In the paper I, we observed that this population is composed mainly by excitatory interneurons, transmits itch through glutamate, and is at least partly downstream to the natriuretic peptide b (NPPB) signaling. Interestingly, increasing amount of behavior evidences have suggested that itch-related information is under local inhibition in the dorsal horn, since decrease of the local inhibitory tone by the peptide somatostatin is able to potentiate itch sensation in mice. In the paper II we complement these findings by showing in vitro that the itch-related GRPR-expressing dorsal horn neurons are under local tonic and phasic inhibition, besides being partly activated by somatostatin, corroborating that this population is indeed part of the disinhibition-induced itch circuitry. In order to confirm the itch-specific phenotype related to GRPR-expressing neurons and extend this theory to the rodent orofacial area, in the paper III we showed a new method developed to target and manipulate the orofacial-related trigeminal neurons. By using this method, we unexpectedly observed a functional switch in the GRPR population, from itch-related in the spinal cord to pain-related in the trigeminal nucleus caudalis, suggesting a labeled line of orofacial pain in this brainstem nucleus. As in the trigeminal nuclei, neuronal circuitry formed by defined cell types transmitting pain- and itch-related information has not been addressed yet in the somatosensory cortex. In the paper IV, we offer a mouse genetic tool that enables the target of barrel field spiny stellate cells, opening for more detailed knowledge of cortical circuitry encoding somatosensory information. In summary, the present thesis brings both complementary findings and new intriguing insights on how our nervous system transmits somatosensory stimuli from different modalities, paving basic knowledge on the mechanisms that build pain and itch as distinct percepts.
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| 16. |
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| 17. |
- Gao, Tianle, et al.
(författare)
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The Neuropeptide Y System Regulates Both Mechanical and Histaminergic Itch
- 2018
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Ingår i: Journal of Investigative Dermatology. - : ELSEVIER SCIENCE INC. - 0022-202X .- 1523-1747. ; 138:11, s. 2405-2411
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Tidskriftsartikel (refereegranskat)abstract
- Itch is a somatosensory modality that serves to alert an organism to harmful elements removable by scratching, such as parasites and chemical irritants. Recently, ablation or silencing of neuropeptide Y (NPY)-expressing spinal interneurons was reported to selectively enhance mechanical itch, whereas chemical itch was unaffected. We examined the effect of activating the NPY/Y-1 receptor system on scratch behavior in mice. We found that intrathecal administration of the Y-1 agonist [Leu(31), Pro(34)]-NPY (LP-NPY) attenuated itch behavior induced by application of 0.07 g von Frey filament in the nape of the neck compared with saline treatment, indicating that activation of the spinal NPY/Y-1 system dampens mechanical itch. However, intrathecal administration of LP-NPY also attenuated chemically induced scratching provoked by intradermal application of histamine or the mast cell degranulator 48/80 (histaminergic itch), and the latter effect could be reversed by administration of the Y-1 antagonist BIBO3304. Intrathecal application of the native nonselective agonist NPY also attenuated histamine or 48/80-induced scratching. Our analyses emphasize the importance of including additional quantitative parameters to characterize the full spectrum of itch behavior and show that the NPY/Y-1 system dampens both mechanically and chemically induced scratching and hence is shared by the two submodalities of itch.
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| 18. |
- Haitina, Tatjana, et al.
(författare)
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Cloning, tissue distribution, pharmacology and three-dimensional modelling of melanocortin receptors 4 and 5 in rainbow trout suggest close evolutionary relationship of these subtypes
- 2004
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Ingår i: Biochemical Journal. - 0264-6021 .- 1470-8728. ; 380:2, s. 475-486
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Tidskriftsartikel (refereegranskat)abstract
- The rainbow trout (Oncorhynchus mykiss) is one of the most widely used fish species in aquaculture and physiological research. In the present paper, we report the first cloning, 3D (three-dimensional) modelling, pharmacological characterization and tissue distribution of two melanocortin (MC) receptors in rainbow trout. Phylogenetic analysis indicates that these receptors are orthologues of the human MC4 and MC5 receptors. We created 3D molecular models of these rainbow trout receptors and their human counterparts. These models suggest greater divergence between the two human receptors than between their rainbow trout counterparts. The pharmacological analyses demonstrated that ACTH (adrenocorticotropic hormone) had surprisingly high affinity for the rainbow trout MC4 and MC5 receptors, whereas alpha-, beta- and gamma-MSH (melanocyte-stimulating hormone) had lower affinity. In second-messenger studies, the cyclic MSH analogues MTII and SHU9119 acted as potent agonist and antagonist respectively at the rainbow trout MC4 receptor, indicating that these ligands are suitable for physiological studies in rainbow trout. Interestingly, we found that the rainbow trout MC4 receptor has a natural high-affinity binding site for zinc ions (0.5 microM) indicating that zinc may play an evolutionary conserved role at this receptor. Reverse transcription PCR indicates that the rainbow trout receptors are expressed both in peripheral tissues and in the central nervous system, including the telencephalon, optic tectum and hypothalamus. Overall, this analysis indicates that the rainbow trout MC4 and MC5 receptors have more in common than their mammalian counterparts, which may suggest that these two receptors have a closer evolutionary relationship than the other MC receptor subtypes.
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| 19. |
- Haring, Martin, et al.
(författare)
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Neuronal atlas of the dorsal horn defines its architecture and links sensory input to transcriptional cell types
- 2018
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Ingår i: Nature Neuroscience. - : NATURE PUBLISHING GROUP. - 1097-6256 .- 1546-1726. ; 21:6, s. 869-880
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Tidskriftsartikel (refereegranskat)abstract
- The dorsal horn of the spinal cord is critical to processing distinct modalities of noxious and innocuous sensation, but little is known of the neuronal subtypes involved, hampering efforts to deduce principles governing somatic sensation. Here we used single-cell RNA sequencing to classify sensory neurons in the mouse dorsal horn. We identified 15 inhibitory and 15 excitatory molecular subtypes of neurons, equaling the complexity in cerebral cortex. Validating our classification scheme in vivo and matching cell types to anatomy of the dorsal horn by spatial transcriptomics reveals laminar enrichment for each of the cell types. Neuron types, when combined, define a multilayered organization with like neurons layered together. Employing our scheme, we find that heat and cold stimuli activate discrete sets of both excitatory and inhibitory neuron types. This work provides a systematic and comprehensive molecular classification of spinal cord sensory neurons, enabling functional interrogation of sensory processing.
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| 20. |
- Iglesias Gonzalez, Ana Belen, et al.
(författare)
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Single cell transcriptomic analysis of spinal Dmrt3 neurons in zebrafish and mouse identifies distinct subtypes and reveal novel subpopulations within the dI6 domain
- 2021
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Ingår i: Frontiers in Cellular Neuroscience. - : Frontiers Media S.A.. - 1662-5102. ; 15
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Tidskriftsartikel (refereegranskat)abstract
- The spinal locomotor network is frequently used for studies into how neuronal circuitsare formed and how cellular activity shape behavioral patterns. A population of dI6interneurons, marked by the Doublesex and mab-3 related transcription factor 3(Dmrt3), has been shown to participate in the coordination of locomotion and gaitsin horses, mice and zebrafish. Analyses of Dmrt3 neurons based on morphology,functionality and the expression of transcription factors have identified differentsubtypes. Here we analyzed the transcriptomes of individual cells belonging to theDmrt3 lineage from zebrafish and mice to unravel the molecular code that underliestheir subfunctionalization. Indeed, clustering of Dmrt3 neurons based on their geneexpression verified known subtypes and revealed novel populations expressing uniquemarkers. Differences in birth order, differential expression of axon guidance genes,neurotransmitters, and their receptors, as well as genes affecting electrophysiologicalproperties, were identified as factors likely underlying diversity. In addition, thecomparison between fish and mice populations offers insights into the evolutionarydriven subspecialization concomitant with the emergence of limbed locomotion
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| 21. |
- Jakobsson, Jon E. T., 1988-
(författare)
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Deciphering neural networks in the somatosensory system using single-cell transcriptomics and rabies tracing
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Itch has evolved to protect us from malicious parasites keen to suck our blood or lay their eggs in our skin. We can detect both the movement of these parasites and the toxins they introduce with specialized neurons called pruriceptors. When we feel an itch, we get a desire to scratch it. Scratching an itch eases the itch sensation, and this is thought to be regulated by neuronal circuits in the spinal cord. This reactive aspect of itch makes is an interesting system to study as it involves both sensory and motor circuitry. The spinal cord hosts a vast number of different neuronal cell types, and better understanding of these are needed to efficiently delineate the circuitry between them. To find these cell types, we sequence the transcriptome of thousands of individual neurons in the dorsal horn of the spinal cord and identified 15 excitatory and 15 inhibitory neuronal populations (Paper I). Furthermore, we found that cell types expressing neuropeptide Y (NPY) contributed to the inhibition of chemically induced itch via the NPY receptor 2 (Paper II) and inhibition of somatostatin-induced itch via NPY receptor 1 (Paper III). We are currently mapping the neurons presynaptic to the NPY neurons using a Npy-Cre mouse line combined with monosynaptic rabies tracing and find inputs from the dorsal root ganglions, the spinal cord, and the brain (Paper IV). To help decipher circuit connectivity, we developed a method that links cell types expressing matching ligand and receptor pairs in single cell RNA-sequencing (scRNA-seq) datasets (Paper V). We furthermore used scRNA-seq to identify differences and similarities of locomotor circuitry related cells expressing doublesex and mab-3 related transcription factor 3 in zebrafish and mouse (Paper VI).In this thesis, we used a combination of powerful and novel tools to investigate questions that were previously difficult to address. It is my belief that spatial transcriptomics, now poised with the knowledge gained from scRNA-seq, will transform how we think about cell types in the central nervous system, since the location of a neuron is critical for its role in a circuit.
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| 22. |
- Jakobsson, Jon E. T., et al.
(författare)
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Neuropeptide Y in itch regulation
- 2019
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Ingår i: Neuropeptides. - : Elsevier BV. - 0143-4179 .- 1532-2785. ; 78
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Forskningsöversikt (refereegranskat)abstract
- Itch is a somatosensory sensation that informs the organism about the presence of potentially harmful substances or parasites, and initiates scratching to remove the threat. Itch-inducing (pruritogenic) substances activate primary afferent neurons in the skin through interactions with specific receptors that converts the stimulus into an electrical signal. These signals are conveyed to the dorsal horn of the spinal cord through the release of neurotransmitters such as natriuretic polypeptide b and somatostatin, leading to an integrated response within a complex spinal inteneuronal network. A large sub-population of somatostatin-expressing spinal interneurons also carry the Neuropeptide Y (NPY) Y1 receptor, indicating that NPY and somatostatin partly regulate the same neuronal pathway. This review focuses on recent findings regarding the role of the NPY/Y1 and somatostatin/SST2A receptor in itch, and also presents data integrating the two neurotransmitter systems.
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| 23. |
- Jakobsson, Jon E. T., et al.
(författare)
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scConnect : a method for exploratory analysis of cell–cell communication based on single-cell RNA-sequencing data
- 2021
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Ingår i: Bioinformatics. - : Oxford University Press. - 1367-4803 .- 1367-4811 .- 1460-2059. ; 37:20, s. 3501-3508
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Tidskriftsartikel (refereegranskat)abstract
- MotivationCell to cell communication is critical for all multicellular organisms, and single-cell sequencing facilitates the construction of full connectivity graphs between cell types in tissues. Such complex data structures demand novel analysis methods and tools for exploratory analysis.ResultsWe propose a method to predict the putative ligand–receptor interactions between cell types from single-cell RNA-sequencing data. This is achieved by inferring and incorporating interactions in a multi-directional graph, thereby enabling contextual exploratory analysis. We demonstrate that our approach can detect common and specific interactions between cell types in mouse brain and human tumors, and that these interactions fit with expected outcomes. These interactions also include predictions made with molecular ligands integrating information from several types of genes necessary for ligand production and transport. Our implementation is general and can be appended to any transcriptome analysis pipeline to provide unbiased hypothesis generation regarding ligand to receptor interactions between cell populations or for network analysis in silico.Availability and implementationscConnect is open source and available as a Python package at https://github.com/JonETJakobsson/scConnect. scConnect is directly compatible with Scanpy scRNA-sequencing pipelines.
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| 24. |
- Lagerström, Malin C., et al.
(författare)
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A sensory subpopulation depends on vesicular glutamate transporter 2 for mechanical pain, and together with substance P, inflammatory pain
- 2011
-
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 108:14, s. 5789-5794
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Tidskriftsartikel (refereegranskat)abstract
- Ablating or functionally compromising sets of sensory neurons has provided important insights into peripheral modality-specific wiring in the somatosensory system. Inflammatory hyperalgesia, cold pain, and noxious mechanosensation have all been shown to depend upon Na(v)1.8-positive sensory neurons. The release of fast-acting neurotransmitters, such as glutamate, and more slowly released neuropeptides, such as substance P (SP), contribute to the diversified responses to external stimuli. Here we show that deleting Vglut2 in Na(v)1.8(Cre)-positive neurons compromised mechanical pain and NGF-induced thermal hyperalgesia, whereas tactile-evoked sensation, thermal, formalin-evoked, and chronic neuropathic pain were normal. However, when Vglut2(f/f); Na(v)1.8(Cre) mice were injected with a SP antagonist before the formalin test, the second phase pain response was nearly completely abolished, whereas in control mice, the pain response was unaffected. Our results suggest that VGLUT2-dependent signaling originating from Na(v)1.8-positive neurons is a principal sensing mechanism for mechanical pain and, together with SP, inflammatory pain. These data define sets of primary afferents associated with specific modalities and provide useful genetic tools with which to analyze the pathways that are activated by functionally distinct neuronal populations and transmitters.
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| 25. |
- Lagerström, Malin C, 1977-
(författare)
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Classification, Evolution, Pharmacology and Structure of G protein-coupled Receptors
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- G protein-coupled receptors (GPCR) are integral membrane proteins with seven α-helices that translate a remarkable diversity of signals into cellular responses. The superfamily of GPCRs is among the largest and most diverse protein families in vertebrates.We have searched the human genome for GPCRs and show that the family includes approximately 800 proteins, which can divided into five main families; Glutamate, Rhodopsin, Adhesion, Frizzled/Taste2 and Secretin. This study represents one of the first overall road maps of the GPCR family in a mammalian genome. Moreover, we identified eight novel members of the human Adhesion family which are characterized by long N-termini with various domains. We also investigated the GPCR repertoire of the chicken genome, where we manually verified a total of 557 chicken GPCRs. We detected several specific expansions and deletions that may reflect some of the functional differences between human and chicken.Substantial effort has been made over the years to find compounds that can bind and activate the melanocortin 4 receptor (MC4R). This receptor is involved in food intake and is thus an important target for antiobesity drugs. We used site-directed mutagenesis to insert micromolar affinity binding sites for zinc between transmembrane (TM) regions 2 and 3. We generated a molecular model of the human MC4R which suggests that a rotation of TM3 is important for activation of the MC4R.Furthermore, we present seven new vertebrate prolactin releasing hormone receptors (PRLHRs) and show that they form two separate subtypes, PRLHR1 and PRLHR2. We performed a pharmacological characterization of the human PRLHR which showed that the receptor can bind neuropeptide Y (NPY) related ligands. We propose that an ancestral PRLH peptide has coevolved with a redundant NPY binding receptor, which then became PRLHR. This suggests how gene duplication events can lead to novel peptide ligand/receptor interactions and hence spur the evolution of new physiological functions.
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| 26. |
- Lagerström, Malin C., et al.
(författare)
-
High affinity agonistic metal ion binding sites within the melanocortin 4 receptor illustrate conformational change of transmembrane region 3
- 2003
-
Ingår i: Journal of Biological Chemistry. - : American Society for Biochemistry and Molecular Biology. - 0021-9258 .- 1083-351X. ; 278:51, s. 51521-51526
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Tidskriftsartikel (refereegranskat)abstract
- We created a molecular model of the human melanocortin 4 receptor (MC4R) and introduced a series of His residues into the receptor protein to form metal ion binding sites. We were able to insert micromolar affinity binding sites for zinc between transmembrane region (TM) 2 and TM3 where the metal ion alone was able to activate this peptide binding G-protein-coupled receptor. The exact conformation of the metal ion interactions allowed us to predict the orientation of the helices, and remodeling of the receptor protein indicated that Glu100 and Ile104 in TM2 and Asp122 and Ile125 in TM3 are directed toward a putative area of activation of the receptor. The molecular model suggests that a rotation of TM3 may be important for activation of the MC4R. Previous models of G-protein-coupled receptors have suggested that unlocking of a stabilizing interaction between the DRY motif, in the cytosolic part of TM3, and TM6 is important for the activation process. We suggest that this unlocking process may be facilitated through creation of a new interaction between TM3 and TM2 in the MC4R.
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| 27. |
- Lagerström, Malin C., et al.
(författare)
-
Origin of the prolactin-releasing hormone (PRLH) receptors : Evidence of coevolution between PRLH and a redundant neuropeptide Y receptor during vertebrate evolution
- 2005
-
Ingår i: Genomics. - : Elsevier BV. - 0888-7543 .- 1089-8646. ; 85:6, s. 688-703
-
Tidskriftsartikel (refereegranskat)abstract
- We present seven new vertebrate homologs of the prolactin-releasing hormone receptor (PRLHR) and show that these are found as two separate subtypes, PRLHR1 and PRLHR2. Analysis of a number of vertebrate sequences using phylogeny, pharmacology, and paralogon analysis indicates that the PRLHRs are likely to share a common ancestry with the neuropeptide Y (NPY) receptors. Moreover, a micromolar level of NPY was able to bind and inhibit completely the PRLH-evoked response in PRLHR1-expressing cells. We suggest that an ancestral PRLH peptide started coevolving with a redundant NPY binding receptor, which then became PRLHR, approximately 500 million years ago. The PRLHR1 subtype was shown to have a relatively high evolutionary rate compared to receptors with fixed peptide preference, which could indicate a drastic change in binding preference, thus supporting this hypothesis. This report suggests how gene duplication events can lead to novel peptide ligand/receptor interactions and hence spur the evolution of new physiological functions.
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| 28. |
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| 29. |
- Lagerström, Malin C., et al.
(författare)
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Structural diversity of G protein-coupled receptors and significance for drug discovery
- 2008
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Ingår i: Nature reviews. Drug discovery. - : Springer Science and Business Media LLC. - 1474-1776 .- 1474-1784. ; 7:4, s. 339-357
-
Forskningsöversikt (refereegranskat)abstract
- G protein-coupled receptors (GPCRs) are the largest family of membrane-bound receptors and also the targets of many drugs. Understanding of the functional significance of the wide structural diversity of GPCRs has been aided considerably in recent years by the sequencing of the human genome and by structural studies, and has important implications for the future therapeutic potential of targeting this receptor family. This article aims to provide a comprehensive overview of the five main human GPCR families--Rhodopsin, Secretin, Adhesion, Glutamate and Frizzled/Taste2--with a focus on gene repertoire, general ligand preference, common and unique structural features, and the potential for future drug discovery.
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| 30. |
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| 31. |
- Lagerström, Malin C., et al.
(författare)
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The evolutionary history and tissue mapping of GPR123 : specific CNS expression pattern predominantly in thalamic nuclei and regions containing large pyramidal cells
- 2007
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Ingår i: Journal of Neurochemistry. - : Wiley. - 0022-3042 .- 1471-4159. ; 100:4, s. 1129-1142
-
Tidskriftsartikel (refereegranskat)abstract
- The Adhesion family of G protein-coupled receptors (GPCRs) includes 33 receptors and is the second largest GPCR family. Most of these proteins are still orphans and fairly little is known of their tissue distribution and evolutionary context. We report the evolutionary history of the Adhesion family protein GPR123 as well as mapping of GPR123 mRNA expression in mouse and rat using in situ hybridization and real-time PCR, respectively. GPR123 was found to be well conserved within the vertebrate lineage, especially within the transmembrane regions and in the distal part of the cytoplasmic tail, containing a potential PDZ binding domain. The real-time PCR data indicates that GPR123 is predominantly expressed in CNS. The in situ data show high expression in thalamic nuclei and regions containing large pyramidal cells like cortex layers 5 and 6 and subiculum. Moreover, we found distinct expression in amygdala, hypothalamus, inferior olive and spinal cord. The CNS specific expression, together with the high sequence conservation between the vertebrate sequences investigated, indicate that GPR123 may have an important role in the regulation of neuronal signal transduction.
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| 32. |
- Lagerström, Malin C., et al.
(författare)
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The G protein-coupled receptor subset of the chicken genome
- 2006
-
Ingår i: PloS Computational Biology. - : Public Library of Science (PLoS). - 1553-734X .- 1553-7358. ; 2:6, s. e54-
-
Tidskriftsartikel (refereegranskat)abstract
- G protein-coupled receptors (GPCRs) are one of the largest families of proteins, and here we scan the recently sequenced chicken genome for GPCRs. We use a homology-based approach, utilizing comparisons with all human GPCRs, to detect and verify chicken GPCRs from translated genomic alignments and Genscan predictions. We present 557 manually curated sequences for GPCRs from the chicken genome, of which 455 were previously not annotated. More than 60% of the chicken Genscan gene predictions with a human ortholog needed curation, which drastically changed the average percentage identity between the human-chicken orthologous pairs (from 56.3% to 72.9%). Of the non-olfactory chicken GPCRs, 79% had a one-to-one orthologous relationship to a human GPCR. The Frizzled, Secretin, and subgroups of the Rhodopsin families have high proportions of orthologous pairs, although the percentage of amino acid identity varies. Other groups show large differences, such as the Adhesion family and GPCRs that bind exogenous ligands. The chicken has only three bitter Taste 2 receptors, and it also lacks an ortholog to human TAS1R2 (one of three GPCRs in the human genome in the Taste 1 receptor family [TAS1R]), implying that the chicken's ability and mode of detecting both bitter and sweet taste may differ from the human's. The chicken genome contains at least 229 olfactory receptors, and the majority of these (218) originate from a chicken-specific expansion. To our knowledge, this dataset of chicken GPCRs is the largest curated dataset from a single gene family from a non-mammalian vertebrate. Both the updated human GPCR dataset, as well the chicken GPCR dataset, are available for download.
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| 33. |
- Lagerström, Malin C., et al.
(författare)
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VGLUT2-Dependent Sensory Neurons in the TRPV1 Population Regulate Pain and Itch
- 2010
-
Ingår i: Neuron. - : Elsevier BV. - 0896-6273 .- 1097-4199. ; 68:3, s. 529-542
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Tidskriftsartikel (refereegranskat)abstract
- The natural response to itch sensation is to scratch, which relieves the itch through an unknown mechanism. Interaction between pain and itch has been frequently demonstrated, and the selectivity hypothesis of itch, based on data from electrophysiological and behavioral experiments, postulates the existence of primary pain afferents capable of repressing itch. Here, we demonstrate that deletion of vesicular glutamate transporter (VGLUT) 2 in a subpopulation of neurons partly overlapping with the vanilloid receptor (TRPV1) primary afferents resulted in a dramatic increase in itch behavior accompanied by a reduced responsiveness to thermal pain. The increased itch behavior was reduced by administration of antihistaminergic drugs and by genetic deletion of the gastrin-releasing peptide receptor, demonstrating a dependence on VGLUT2 to maintain normal levels of both histaminergic and nonhistaminergic itch. This study establishes that VGLUT2 is a major player in TRPV1 thermal nociception and also serves to regulate a normal itch response.
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| 34. |
- Ma, Haisha, et al.
(författare)
-
The Neuropeptide Y Y-2 Receptor Is Coexpressed with Nppb in Primary Afferent Neurons and Y-2 Activation Reduces Histaminergic and IL-31-Induced Itch
- 2020
-
Ingår i: Journal of Pharmacology and Experimental Therapeutics. - : AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS. - 0022-3565 .- 1521-0103. ; 372:1, s. 73-82
-
Tidskriftsartikel (refereegranskat)abstract
- Itch stimuli are detected by specialized primary afferents that convey the signal to the spinal cord, but how itch transmission is regulated is still not completely known. Here, we investigated the roles of the neuropeptide Y (NPY)/Y-2 receptor system on scratch behavior. The inhibitory Y-2 receptor is expressed on mouse primary afferents, and intrathecal administration of the Y-2 agonist peptide YY (PYY)(3-36) reduced scratch episode frequency and duration induced by compound 48/80, an effect that could be reversed by intrathecal preadministration of the Y-2 antagonist BIIE0246. Also, scratch episode duration induced by histamine could be reduced by PYY3-36. In contrast, scratch behavior induced by alpha-methyl-5HT, protease-activated receptor-2-activating peptide SLIGRL, chloroquine, topical dust mite extract, or mechanical itch induced by von Frey filaments was unaffected by stimulation of Y2. Primary afferent neurons expressing the Npy2r gene were found to coexpress itch-associated markers such as natriuretic peptide precursor b, oncostatin M receptor, and interleukin (IL) 31 receptor A. Accordingly, intrathecal PYY3-36 reduced the scratch behavior induced by IL-31. Our findings imply that the NPY/Y-2 system reduces histaminergic and IL-31-associated itch through presynaptic inhibition of a subpopulation of itch-associated primary afferents. SIGNIFICANCE STATEMENT The spinal neuropeptide Y system dampens scratching behavior induced by histaminergic compounds and interleukin 31, a cytokine involved in atopic dermatitis, through interactions with the Y-2 receptor. The Y-2 receptor is expressed by primary afferent neurons that are rich in itch-associated neurotransmitters and receptors such as somatostatin, natriuretic peptide precursor b, and interleukin 31 receptors.
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| 35. |
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| 36. |
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| 37. |
- Magnúsdóttir, Elín Ingibjörg, et al.
(författare)
-
Mouse connective tissue mast cell proteases tryptase and carboxypeptidase A3 play protective roles in itch induced by endothelin-1
- 2020
-
Ingår i: Journal of Neuroinflammation. - : BMC. - 1742-2094. ; 17
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Itch is an unpleasant sensation that can be debilitating, especially if it is chronic and of non-histaminergic origin, as treatment options are limited. Endothelin-1 (ET-1) is a potent endogenous vasoconstrictor that also has the ability to induce a burning, non-histaminergic pruritus when exogenously administered, by activating the endothelin A receptor (ETAR) on primary afferents. ET-1 is released endogenously by several cell-types found in the skin, including macrophages and keratinocytes. Mast cells express ET(A)Rs and can thereby be degranulated by ET-1, and mast cell proteases chymase and carboxypeptidase A3 (CPA3) are known to either generate or degrade ET-1, respectively, suggesting a role for mast cell proteases in the regulation of ET-1-induced itch. The mouse mast cell proteases (mMCPs) mMCP4 (chymase), mMCP6 (tryptase), and CPA3 are found in connective tissue type mast cells and are the closest functional homologs to human mast cell proteases, but little is known about their role in endothelin-induced itch.Methods: In this study, we evaluated the effects of mast cell protease deficiency on scratching behavior induced by ET-1. To investigate this, mMCP knock-out and transgenic mice were injected intradermally with ET-1 and their scratching behavior was recorded and analyzed.Results: CPA3-deficient mice and mice lacking all three proteases demonstrated highly elevated levels of scratching behavior compared with wild-type controls. A modest increase in the number of scratching bouts was also seen in mMCP6-deficient mice, while mMCP4-deficiency did not have any effect.Conclusion: Altogether, these findings identify a prominent role for the mast cell proteases, in particular CPA3, in the protection against itch induced by ET-1.
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| 38. |
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| 39. |
- Magnúsdóttir, Elín Ingibjörg, et al.
(författare)
-
Mouse mast cells and mast cell proteases do not play a significant role in acute tissue injury pain induced by formalin
- 2018
-
Ingår i: Molecular Pain. - : SAGE Publications. - 1744-8069. ; 14
-
Tidskriftsartikel (refereegranskat)abstract
- Subcutaneous formalin injections are used as a model for tissue injury-induced pain where formalin induces pain and inflammation indirectly by crosslinking proteins and directly through activation of the transient receptor potential A1 receptor on primary afferents. Activation of primary afferents leads to both central and peripheral release of neurotransmitters. Mast cells are found in close proximity to peripheral sensory nerve endings and express receptors for neurotransmitters released by the primary afferents, contributing to the neuro/immune interface. Mast cell proteases are found in large quantities within mast cell granules and are released continuously in small amounts and upon mast cell activation. They have a wide repertoire of proposed substrates, including Substance P and calcitonin gene-related peptide, but knowledge of their in vivo function is limited. We evaluated the role of mouse mast cell proteases (mMCPs) in tissue injury pain responses induced by formalin, using transgenic mice lacking either mMCP4, mMCP6, or carboxypeptidase A3 (CPA3), or mast cells in their entirety. Further, we investigated the role of mast cells in heat hypersensitivity following a nerve growth factor injection. No statistical difference was observed between the respective mast cell protease knockout lines and wild-type controls in the formalin test. Mast cell deficiency did not have an effect on formalin-induced nociceptive responses nor nerve growth factor-induced heat hypersensitivity. Our data thus show that mMCP4, mMCP6, and CPA3 as well as mast cells as a whole, do not play a significant role in the pain responses associated with acute tissue injury and inflammation in the formalin test. Our data also indicate that mast cells are not essential to heat hypersensitivity induced by nerve growth factor.
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| 40. |
- Magnúsdóttir, Elín Ingibjörg
(författare)
-
Peripheral Regulation of Pain and Itch
- 2019
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Pain and itch are diverse sensory modalities, transmitted by the somatosensory nervous system. Stimuli such as heat, cold, mechanical pain and itch can be transmitted by different neuronal populations, which show considerable overlap with regards to sensory activation. Moreover, the immune and nervous systems can be involved in extensive crosstalk in the periphery when reacting to these stimuli. With recent advances in genetic engineering, we now have the possibility to study the contribution of distinct neuron types, neurotransmitters and other mediators in vivo by using gene knock-out mice. The neuropeptide calcitonin gene-related peptide (CGRP) and the ion channel transient receptor potential cation channel subfamily V member 1 (TRPV1) have both been implicated in pain and itch transmission. In Paper I, the Cre-LoxP system was used to specifically remove CGRPα from the primary afferent population that expresses TRPV1. CGRPα-mCherrylx/lx;Trpv1-Cre mice had attenuated responses to visceral pain induced by acid, while mechanosensitivity of the colon and somatic pain sensation remained unaffected.Mast cell proteases (MCPs) are stored in high quantities within mast cell (MC) granules and have been linked to both protective and pro-inflammatory properties, but little is known about their exact roles in vivo. In Papers II, IV and V, we used knock-out mice to investigate the contribution of MCs and their MCPs (the chymase mMCP4, tryptase mMCP6 and carboxypeptidase CPA3) in pain resulting from tissue injury, inflammation-induced heat hypersensitivity and different types of itch. Surprisingly, we found that neither MCPs nor MCs were essential for the pain behavior tested (Paper II). Our data indicate that mMCP6 and CPA3 have a protective role in scratching behavior induced by the peptide endothelin-1 (ET-1; Paper IV) and in scratching induced by the MC degranulator compound 48/80 (Paper V), but no differences were observed with the other pruritogens histamine, chloroquine or SLIGRL.In Paper III, we saw that a novel single-stranded oligonucleotide (ssON) attenuated compound 48-induced scratching in BALB/c mice by blocking MC degranulation. ssON could also block degranulation in human MC in vitro and we determined that this was due to ssON interfering with Mas-related G protein-coupled receptor X2 (MRGPRX2), a receptor involved in non-allergic MC degranulation.By better understanding the contribution of individual components of the nervous and immune systems in pain and itch, we hopefully increase the possibilities of developing better treatments for burdensome pain- and itch-related disorders in the future.
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| 41. |
- Nordström, Karl J V, et al.
(författare)
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The Secretin GPCRs descended from the family of Adhesion GPCRs
- 2009
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Ingår i: Molecular biology and evolution. - : Oxford University Press (OUP). - 0737-4038 .- 1537-1719. ; 26:1, s. 71-84
-
Tidskriftsartikel (refereegranskat)abstract
- The Adhesion G-protein-coupled receptors (GPCRs) are the most complex gene family among GPCRs with large genomic size, multiple introns, and a fascinating flora of functional domains, though the evolutionary origin of this family has been obscure. Here we studied the evolution of all class B (7tm2)-related genes, including the Adhesion, Secretin, and Methuselah families of GPCRs with a focus on nine genomes. We found that the cnidarian genome of Nematostella vectensis has a remarkably rich set of Adhesion GPCRs with a broad repertoire of N-terminal domains although this genome did not have any Secretin GPCRs. Moreover, the single-celled and colony-forming eukaryotes Monosiga brevicollis and Dictyostelium discoideum contain Adhesion-like GPCRs although these genomes do not have any Secretin GPCRs suggesting that the Adhesion types of GPCRs are the most ancient among class B GPCRs. Phylogenetic analysis found Adhesion group V (that contains GPR133 and GPR144) to be the closest relative to the Secretin family in the Adhesion family. Moreover, Adhesion group V sequences in N. vectensis share the same splice site setup as the Secretin GPCRs. Additionally, one of the most conserved motifs in the entire Secretin family is only found in group V of the Adhesion family. We suggest therefore that the Secretin family of GPCRs could have descended from group V Adhesion GPCRs. We found a set of unique Adhesion-like GPCRs in N. vectensis that have long N-termini containing one Somatomedin B domain each, which is a domain configuration similar to that of a set of Adhesion-like GPCRs found in Branchiostoma floridae. These sequences show slight similarities to Methuselah sequences found in insects. The extended class B GPCRs have a very complex evolutionary history with several species-specific expansions, and we identified at least 31 unique N-terminal domains originating from other protein classes. The overall N-terminal domain structure, however, concurs with the phylogenetic analysis of the transmembrane domains, thus enabling us to track the origin of most of the subgroups.
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| 42. |
- Pickering, Chris, et al.
(författare)
-
The Adhesion GPCR GPR125 is specifically expressed in the choroid plexus and is upregulated following brain injury
- 2008
-
Ingår i: BMC Neuroscience. - : Springer Science and Business Media LLC. - 1471-2202. ; 9, s. 97-
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUNDGPR125 belongs to the family of Adhesion G protein-coupled receptors (GPCRs). A single copy of GPR125 was found in many vertebrate genomes. We also identified a Drosophila sequence, DmCG15744, which shares a common ancestor with the entire Group III of Adhesion GPCRs, and also contains Ig, LRR and HBD domains which were observed in mammalian GPR125.RESULTSWe found specific expression of GPR125 in cells of the choroid plexus using in situ hybridization and protein-specific antibodies and combined in situ/immunohistochemistry co-localization using cytokeratin, a marker specific for epithelial cells. Induction of inflammation by LPS did not change GPR125 expression. However, GPR125 expression was transiently increased (almost 2-fold) at 4 h after traumatic brain injury (TBI) followed by a decrease (approximately 4-fold) from 2 days onwards in the choroid plexus as well as increased expression (2-fold) in the hippocampus that was delayed until 1 day after injury.CONCLUSIONThese findings suggest that GPR125 plays a functional role in choroidal and hippocampal response to injury.
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| 43. |
- Rogoz, Katarzyna, et al.
(författare)
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Glutamate, Substance P, and Calcitonin Gene-Related Peptide Cooperate in Inflammation-Induced Heat Hyperalgesia
- 2014
-
Ingår i: Molecular Pharmacology. - : American Society for Pharmacology & Experimental Therapeutics (ASPET). - 0026-895X .- 1521-0111. ; 85:2, s. 322-334
-
Tidskriftsartikel (refereegranskat)abstract
- The transient receptor potential cation channel subfamily V member 1 (TRPV1) is known as a thermosensor and integrator of inflammation-induced hyperalgesia. TRPV1 is expressed in a sub-population of primary afferent neurons that express several different neurotransmitters. The role of the TRPV1 channel in the development of hyperalgesia is established, but the role of the neurotransmitter glutamate, used partially by the same neuronal population and thus probably mediating the response, is still under investigation. We have used a Trpv1-Cre mouse line in which we either ablated Trpv1-Cre expressing neurons or induced vesicular glutamate transporter 2 (Vglut2) deficiency in Trpv1-Cre expressing neurons and investigated specific states of hyperalgesia after persistent inflammation. Furthermore, by pharmacologic inhibition of substance P (SP) or calcitonin gene-related peptide (CGRP) signaling in Vglut2-deficient mice, we also evaluated the contribution of SP or CGRP to inflammation-induced hyperalgesia, with or without the presence of vesicular glutamate transporter 2 (VGLUT2)-mediated glutamatergic transmission in Trpv1-Cre neurons. This examination, together with c-Fos analyses, showed that VGLUT2-mediated glutamatergic transmission in Trpv1-Cre afferents together with SP or CGRP is essential for the development of the heat hyperalgesia associated with persistent inflammation. Additionally, SP-, CGRP-, and VGLUT2-mediated transmission together were found to play a role in the development of mechanical hyperalgesia after persistent inflammation.
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| 44. |
- Rogóz, Katarzyna, et al.
(författare)
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Multimodal Use of Calcitonin Gene-Related Peptide and Substance P in Itch and Acute Pain Uncovered by the Elimination of Vesicular Glutamate Transporter 2 from Transient Receptor Potential Cation Channel Subfamily V Member 1 Neurons
- 2014
-
Ingår i: Journal of Neuroscience. - 0270-6474 .- 1529-2401. ; 34:42, s. 14055-14068
-
Tidskriftsartikel (refereegranskat)abstract
- Primary afferents are known to use glutamate as their principal fast neurotransmitter. However, it has become increasingly clear that peptides have an influential role in both mediating and modulating sensory transmission. Here we describe the transmission accounting for different acute pain states and itch transmitted via the transient receptor potential cation channel subfamily V member 1 (TRPV1) population by either ablating Trpv1-Cre-expressing neurons or inducing vesicular glutamate transporter 2 (VGLUT2) deficiency in Trpv1-Cre-expressing neurons. Furthermore, by pharmacological inhibition of substance P or calcitonin gene-related peptide (CGRP) signaling in Vglut2-deficient mice, we evaluated the contribution of substance P or CGRP to these sensory modulations, with or without the presence of VGLUT2-mediated glutamatergic transmission in Trpv1-Cre neurons. This examination, together with c-Fos analyses, showed that glutamate via VGLUT2 in the Trpv1-Cre population together with substance P mediate acute cold pain, whereas glutamate together with CGRP mediate noxious heat. Moreover, we demonstrate that glutamate together with both substance P and CGRP mediate tissue-injury associated pain. We further show that itch, regulated by the VGLUT2-mediated transmission via the Trpv1-Cre population, depends on CGRP and gastrin-releasing peptide receptor (GRPR) transmission because pharmacological blockade of the CGRP or GRPR pathway, or genetic ablation of Grpr, led to a drastically attenuated itch. Our study reveals how different neurotransmitters combined can cooperate with each other to transmit or regulate various acute sensations, including itch.
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| 45. |
- Rogóz, Katarzyna, et al.
(författare)
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VGLUT2 controls heat and punctuate hyperalgesia associated with nerve injury via TRPV1-Cre primary afferents
- 2015
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:1
-
Tidskriftsartikel (refereegranskat)abstract
- Nerve injury induces a state of prolonged thermal and mechanical hypersensitivity in the innervated area, causing distress in affected individuals. Nerve injury-induced hypersensitivity is partially due to increased activity and thereby sustained release of neurotransmitters from the injured fibers. Glutamate, a prominent neurotransmitter in primary afferents, plays a major role in development of hypersensitivity. Glutamate is packed in vesicles by vesicular glutamate transporters (VGLUTs) to enable controlled release upon depolarization. While a role for peripheral VGLUTs in nerve injury-induced pain is established, their contribution in specific peripheral neuronal populations is unresolved. We investigated the role of VGLUT2, expressed by transient receptor potential vanilloid (TRPV1) fibers, in nerve injury-induced hypersensitivity. Our data shows that removal of Vglut2 from Trpv1-Cre neurons using transgenic mice abolished both heat and punctuate hyperalgesia associated with nerve injury. In contrast, the development of cold hypersensitivity after nerve injury was unaltered. Here, we show that, VGLUT2-mediated glutamatergic transmission from Trpv1-Cre neurons selectively mediates heat and mechanical hypersensitivity associated with nerve injury. Our data clarifies the role of the Trpv1-Cre population and the dependence of VGLUT2-mediated glutamatergic transmission in nerve injury-induced hyperalgesia.
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| 46. |
- Rogoz, Katarzyna, et al.
(författare)
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VGLUT2-dependent glutamatergic transmission in primary afferents is required for intact nociception in both acute and persistent pain modalities
- 2012
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Ingår i: Pain. - : Ovid Technologies (Wolters Kluwer Health). - 0304-3959 .- 1872-6623. ; 153:7, s. 1525-1536
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Tidskriftsartikel (refereegranskat)abstract
- Glutamate is an essential transmitter in pain pathways. However, its broad usage in the central and peripheral nervous system prevents us from designing efficient glutamate-based pain therapies without causing harmful side effects. The discovery of vesicular glutamate transporters (VGLUT1-3) has been a crucial step in describing specific glutamatergic neuronal subpopulations and glutamate-dependent pain pathways. To assess the role of VGLUT2-mediated glutamatergic contribution to pain transmission from the entire primary sensory population, we crossed our Vglut2(f/f) line with the Ht-Pa-Cre line. Such Vglut2-deficient mice showed significantly decreased, but not completely absent, acute nociceptive responses. The animals were less prone to develop an inflammatory-related state of pain and were, in the partial sciatic nerve ligation chronic pain model, much less hypersensitive to mechanical stimuli and did not develop cold allodynia or heat hyperalgesia. To take advantage of this neuropathic pain-resistant model, we analyzed Vglut2-dependent transcriptional changes in the dorsal spinal cord after nerve injury, which revealed several novel candidate target genes potentially relevant for the development of neuropathic pain therapeutics. Taken together, we conclude that VGLUT2 is a major mediator of nociception in primary afferents, implying that glutamate is the key somatosensory neurotransmitter.
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| 47. |
- Spencer, Nick J., et al.
(författare)
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CGRP alpha within the Trpv1-Cre population contributes to visceral nociception
- 2018
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Ingår i: American Journal of Physiology - Gastrointestinal and Liver Physiology. - : AMER PHYSIOLOGICAL SOC. - 0193-1857 .- 1522-1547. ; 314:2, s. G188-G200
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Tidskriftsartikel (refereegranskat)abstract
- The role of calcitonin gene-related peptide (CGRP) in visceral and somatic nociception is incompletely understood. CGRP alpha is highly expressed in sensory neurons of dorsal root ganglia and particularly in neurons that also express the transient receptor potential cation channel subfamily V member 1 (Trpv1). Therefore, we investigated changes in visceral and somatic nociception following deletion of CGRP alpha from the Trpv1-Cre population using the Cre/lox system. In control mice, acetic acid injection (0.6%, ip) caused significant immobility (time stationary), an established indicator of visceral pain. In CGRP alpha-mCherry(lx/lx); Trpv1-Cre mice, the duration of immobility was significantly less than controls, and the distance CGRP alpha-mCherry(lx/lx); Trpv1-Cre mice traveled over 20 min following acetic acid was significantly greater than controls. However, following acetic acid injection, there was no difference between genotypes in the writhing reflex, number of abdominal licks, or forepaw wipes of the cheek. CGRP alpha-mCherry(lx/lx); Trpv1-Cre mice developed more pronounced inflammation-induced heat hypersensitivity above baseline values compared with controls. However, analyses of noxious acute heat or cold transmission revealed no difference between genotypes. Also, odor avoidance test, odor preference test, and buried food test for olfaction revealed no differences between genotypes. Our findings suggest that CGRP alpha-mediated transmission within the Trpv1-Cre population plays a significant role in visceral nociceptive pathways underlying voluntary movement. Monitoring changes in movement over time is a sensitive parameter to identify differences in visceral nociception, compared with writhing reflexes, abdominal licks, or forepaw wipes of the cheek that were unaffected by deletion of CGRP alpha- from Trpv1-Cre population and likely utilize different mechanisms. NEW & NOTEWORTHY The neuropeptide calcitonin gene-related peptide (CGRP) is highly colocalized with transient receptor potential cation channel subfamily V member 1 (TRPV1)-expressing primary afferent neurons, but the functional role of CGRP alpha specifically in these neurons is unknown in pain processing from visceral and somatic afferents. We used cre-lox recombination to conditionally delete CGRP alpha from TRPV1-expressing neurons in mice. We show that CGRP alpha from within TRPV1-cre population plays an important role in visceral nociception but less so in somatic nociception.
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| 48. |
- Viereckel, Thomas, et al.
(författare)
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Midbrain Gene Screening Identifies a New Mesoaccumbal Glutamatergic Pathway and a Marker for Dopamine Cells Neuroprotected in Parkinson's Disease
- 2016
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- The ventral tegmental area (VTA) and substantia nigra pars compacta (SNc) of the midbrain are associated with Parkinson's disease (PD), schizophrenia, mood disorders and addiction. Based on the recently unraveled heterogeneity within the VTA and SNc, where glutamate, GABA and co-releasing neurons have been found to co-exist with the classical dopamine neurons, there is a compelling need for identification of gene expression patterns that represent this heterogeneity and that are of value for development of human therapies. Here, several unique gene expression patterns were identified in the mouse midbrain of which NeuroD6 and Grp were expressed within different dopaminergic subpopulations of the VTA, and TrpV1 within a small heterogeneous population. Optogenetics-coupled in vivo amperometry revealed a previously unknown glutamatergic mesoaccumbal pathway characterized by TrpV1-Cre-expression. Human GRP was strongly detected in non-melanized dopaminergic neurons within the SNc of both control and PD brains, suggesting GRP as a marker for neuroprotected neurons in PD. This study thus unravels markers for distinct subpopulations of neurons within the mouse and human midbrain, defines unique anatomical subregions within the VTA and exposes an entirely new glutamatergic pathway. Finally, both TRPV1 and GRP are implied in midbrain physiology of importance to neurological and neuropsychiatric disorders.
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| 49. |
- Wallén-Mackenzie, Åsa, et al.
(författare)
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Restricted cortical and amygdaloid removal of vesicular glutamate transporter 2 in preadolescent mice impacts dopaminergic activity and neuronal circuitry of higher brain function.
- 2009
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Ingår i: The Journal of neuroscience : the official journal of the Society for Neuroscience. - 1529-2401 .- 0270-6474. ; 29:7, s. 2238-51
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Tidskriftsartikel (refereegranskat)abstract
- A major challenge in neuroscience is to resolve the connection between gene functionality, neuronal circuits, and behavior. Most, if not all, neuronal circuits of the adult brain contain a glutamatergic component, the nature of which has been difficult to assess because of the vast cellular abundance of glutamate. In this study, we wanted to determine the role of a restricted subpopulation of glutamatergic neurons within the forebrain, the Vglut2-expressing neurons, in neuronal circuitry of higher brain function. Vglut2 expression was selectively deleted in the cortex, hippocampus, and amygdala of preadolescent mice, which resulted in increased locomotor activity, altered social dominance and risk assessment, decreased sensorimotor gating, and impaired long-term spatial memory. Presynaptic VGLUT2-positive terminals were lost in the cortex, striatum, nucleus accumbens, and hippocampus, and a downstream effect on dopamine binding site availability in the striatum was evident. A connection between the induced late-onset, chronic reduction of glutamatergic neurotransmission and dopamine signaling within the circuitry was further substantiated by a partial attenuation of the deficits in sensorimotor gating by the dopamine-stabilizing antipsychotic drug aripiprazole and an increased sensitivity to amphetamine. Somewhat surprisingly, given the restricted expression of Vglut2 in regions responsible for higher brain function, our analyses show that VGLUT2-mediated neurotransmission is required for certain aspects of cognitive, emotional, and social behavior. The present study provides support for the existence of a neurocircuitry that connects changes in VGLUT2-mediated neurotransmission to alterations in the dopaminergic system with schizophrenia-like behavioral deficits as a major outcome.
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| 50. |
- Weman, Hannah M., 1993-, et al.
(författare)
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Exon excision of the vesicular glutamate transporter 2 or vesicular inhibitory amino acid transporter from glycine receptor alpha 3-expressing neurons does not alter thermal hypersensitivity induced by interleukin 1 beta
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The glycinergic system serves a protective role in acute itch and pain transmission and further, in inflammatory- and nerve damage-induced hypersensitivity. The glycine receptor alpha 3 (GLRA3) subunit has previously been coupled to inflammatory, but not acute pain. Using the Glra3-Cre mouse line, we investigated the role of Glra3(+) neurons in heat hypersensitivity following administration of the pro-inflammatory and -nociceptive cytokine interleukin 1 beta (IL1β). Spatial transcriptional analysis revealed, in consistency with our previous results, that the Glra3(+) population constitutes of a major excitatory vesicular glutamate transporter 2 (Vglut2) and a minor vesicular inhibitory amino acid transporter (Viaat) expressing subpopulation. Administration of IL1β resulted in hypersensitivity to heat, however, the conditional global attenuation of the fast glutamatergic or inhibitory transmission in Glra3-Cre neurons did not affect the heat withdrawal response, suggesting that the Glra3(+)Vglut2(+) nor Glra3(+)Viaat(+) neurons are involved in interleukin 1 beta-induced heat hypersensitivity. Future experiments for this project will include chemogenetic experiments to overall investigate the hypersensitivity role of the lumbar Glra3-Cre population, as well as investigation of the sensory involvement in nerve growth factor beta (NGFβ)-induced thermal hyperalgesia.
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