Patients with liver diseases often suffer from chronic itch, yet the pruritogen(s) and receptor(s) remain largely elusive. Here, we identify bile acids as natural ligands for MRGPRX4. MRGPRX4 is ...expressed in human dorsal root ganglion (hDRG) neurons and co-expresses with itch receptor HRH1. Bile acids elicited Ca
responses in cultured hDRG neurons, and bile acids or a MRGPRX4 specific agonist induced itch in human subjects. However, a specific agonist for another bile acid receptor TGR5 failed to induce itch in human subjects and we find that human TGR5 is not expressed in hDRG neurons. Finally, we show positive correlation between cholestatic itch and plasma bile acids level in itchy patients and the elevated bile acids is sufficient to activate MRGPRX4. Taken together, our data strongly suggest that MRGPRX4 is a novel bile acid receptor that likely underlies cholestatic itch in human, providing a promising new drug target for anti-itch therapies.
Glial reaction is a common feature of neurodegenerative diseases. Recent studies have suggested that reactive astrocytes gain neurotoxic properties, but exactly how reactive astrocytes contribute to ...neurotoxicity remains to be determined. Here, we identify lipocalin 2 (lcn2) as an inducible factor that is secreted by reactive astrocytes and that is selectively toxic to neurons. We show that lcn2 is induced in reactive astrocytes in transgenic rats with neuronal expression of mutant human TAR DNA-binding protein 43 (TDP-43) or RNA-binding protein fused in sarcoma (FUS). Therefore, lcn2 is induced in activated astrocytes in response to neurodegeneration, but its induction is independent of TDP-43 or FUS expression in astrocytes. We found that synthetic lcn2 is cytotoxic to primary neurons in a dose-dependent manner, but is innocuous to astrocytes, microglia, and oligodendrocytes. Lcn2 toxicity is increased in neurons that express a disease gene, such as mutant FUS or TDP-43. Conditioned medium from rat brain slice cultures with neuronal expression of mutant TDP-43 contains abundant lcn2 and is toxic to primary neurons as well as neurons in cultured brain slice from WT rats. Partial depletion of lcn2 by immunoprecipitation reduced conditioned medium-mediated neurotoxicity. Our data indicate that reactive astrocytes secrete lcn2, which is a potent neurotoxic mediator.
Mutation of Tar DNA‐binding protein 43 (TDP‐43) is linked to amyotrophic lateral sclerosis. Although astrocytes have important roles in neuron function and survival, their potential contribution to ...TDP‐43 pathogenesis is unclear. Here, we created novel lines of transgenic rats that express a mutant form of human TDP‐43 (M337V substitution) restricted to astrocytes. Selective expression of mutant TDP‐43 in astrocytes caused a progressive loss of motor neurons and the denervation atrophy of skeletal muscles, resulting in progressive paralysis. The spinal cord of transgenic rats also exhibited a progressive depletion of the astroglial glutamate transporters GLT‐1 and GLAST. Astrocytic expression of mutant TDP‐43 led to activation of astrocytes and microglia, with an induction of the neurotoxic factor Lcn2 in reactive astrocytes that was independent of TDP‐43 expression. These results indicate that mutant TDP‐43 in astrocytes is sufficient to cause non‐cell‐autonomous death of motor neurons. This motor neuron death likely involves deficiency in neuroprotective genes and induction of neurotoxic genes in astrocytes.
The debated role of astrocytes in amyotrophic lateral sclerosis (ALS) gains additional support from neurodegenerative phenotypes caused by astrocyte‐restricted expression of mutant TDP‐43 in transgenic rats.
Whether glutamate or itch-selective neurotransmitters are used to confer itch specificity is still under debate. We focused on an itch-selective population of primary afferents expressing MRGPRA3, ...which highly expresses Vglut2 and the neuropeptide neuromedin B (Nmb), to investigate this question. Optogenetic stimulation of MRGPRA3+ afferents triggers scratching and other itch-related avoidance behaviors. Using a combination of optogenetics, spinal cord slice recordings, Vglut2 conditional knockout mice, and behavior assays, we showed that glutamate is essential for MRGPRA3+ afferents to transmit itch. We further demonstrated that MRGPRA3+ afferents form monosynaptic connections with both NMBR+ and NMBR− neurons and that NMB and glutamate together can enhance the activity of NMBR+ spinal DH neurons. Moreover, Nmb in MRGPRA3+ afferents and NMBR+ DH neurons are required for chloroquine-induced scratching. Together, our results establish a new model in which glutamate is an essential neurotransmitter in primary afferents for itch transmission, whereas NMB signaling enhances its activities.
•Itch-selective MRGPRA3+ afferents highly express Vglut2 and Nmb•Glutamate is an essential neurotransmitter in MRGPRA3+ afferents for itch signaling•MRGPRA3+ afferents form monosynaptic connections with NMBR+ dorsal horn neurons•Nmb in MRGPRA3+ afferents and NMBR+ dorsal horn neurons function in itch sensation
Cui et al. demonstrate that glutamate is required for itch transmission in MRGPRA3+ primary afferents, which also use the neuropeptide NMB to enhance signaling. Because both glutamate and Nmb are broadly available in primary afferents, this suggests that distinct neurotransmitters are not necessary to confer itch modality specificity.
Pathogenic mutation of ubiquilin 2 (UBQLN2) causes neurodegeneration in amyotrophic lateral sclerosis and frontotemporal lobar degeneration. How UBQLN2 mutations cause the diseases is not clear. ...While over‐expression of UBQLN2 with pathogenic mutation causes neuron death in rodent models, deletion of the Ubqln2 in rats has no effect on neuronal function. Previous findings in animal models suggest that UBQLN2 mutations cause the diseases mainly through a gain rather than a loss of functions. To examine whether the toxic gain in UBQLN2 mutation is related to the enhancement of UBQLN2 functions, we created new transgenic rats over‐expressing wild‐type human UBQLN2. Considering that human UBQLN2 may not function properly in the rat genome, we also created transgenic rats over‐expressing rat's own Ubqln2. When over‐expressed in rats, both human and rat wild‐type Ubqln2 caused neuronal death and spatial learning deficits, the pathologies that were indistinguishable from those observed in mutant UBQLN2 transgenic rats. Over‐expressed wild‐type UBQLN2 formed protein inclusions attracting the autophagy substrate sequestosome‐1 and the proteasome component 26S proteasome regulatory subunit 7. These findings suggest that excess UBQLN2 is toxic rather than protective to neurons and that the enhancement of UBQLN2 functions is involved in UBQLN2 pathogenesis.
Pathogenic mutation in ubiquilin 2 (UBQLN2) causes neurodegeneration in ALS and FTLD. Studies in rodent models suggest a gain of toxic function in mutant UBQLN2. We created new transgenic rats as a relevant model and examined whether enhancing wild‐type UBQLN2 expression is implicated in the pathogenesis of mutant UBQLN2. We observed that over‐expression of human or rat wild‐type Ubqln2 caused protein aggregation and neuronal death in transgenic rats. Our findings suggest that excess UBQLN2 is toxic rather than protective to neurons and that uncontrolled enhancement of UBQLN2 function is involved in UBQLN2 pathogenesis.
Read the Editorial Highlight for this article on page 159.
Pathogenic mutation in ubiquilin 2 (UBQLN2) causes neurodegeneration in ALS and FTLD. Studies in rodent models suggest a gain of toxic function in mutant UBQLN2. We created new transgenic rats as a relevant model and examined whether enhancing wild‐type UBQLN2 expression is implicated in the pathogenesis of mutant UBQLN2. We observed that over‐expression of human or rat wild‐type Ubqln2 caused protein aggregation and neuronal death in transgenic rats. Our findings suggest that excess UBQLN2 is toxic rather than protective to neurons and that uncontrolled enhancement of UBQLN2 function is involved in UBQLN2 pathogenesis.
Read the Editorial Highlight for this article on page 159.
The roles of Aβ low-threshold mechanoreceptors (LTMRs) in transmitting mechanical hyperalgesia and in alleviating chronic pain have been of great interest but remain contentious. Here we utilized ...intersectional genetic tools, optogenetics, and high-speed imaging to specifically examine functions of Split
labeled mouse Aβ-LTMRs in this regard. Genetic ablation of Split
-Aβ-LTMRs increased mechanical nociception but not thermosensation in both acute and chronic inflammatory pain conditions, indicating a modality-specific role in gating mechanical nociception. Local optogenetic activation of Split
-Aβ-LTMRs triggered nociception after tissue inflammation, whereas their broad activation at the dorsal column still alleviated mechanical hypersensitivity of chronic inflammation. Taking all data into consideration, we propose a model, in which Aβ-LTMRs play distinctive local and global roles in transmitting or alleviating mechanical hyperalgesia of chronic pain, respectively. Our model suggests a strategy of global activation plus local inhibition of Aβ-LTMRs for treating mechanical hyperalgesia.
A dominant mutation in hnRNPA1 causes amyotrophic lateral sclerosis (ALS), but it is not known whether this mutation leads to motor neuron death through increased or decreased function. To elucidate ...the relationship between pathogenic hnRNPA1 mutation and its native function, we created novel transgenic rats that overexpressed wildtype rat hnRNPA1 exclusively in motor neurons. This targeted expression of wildtype hnRNPA1 caused severe motor neuron loss and subsequent denervation muscle atrophy in transgenic rats that recapitulated the characteristics of ALS. These findings demonstrate that the augmentation of hnRNPA1 expression suffices to trigger motor neuron degeneration and the manifestation of ALS-like phenotypes. It is reasonable to infer that an amplification of an as-yet undetermined hnRNPA1 function plays a pivotal role in the pathogenesis of familial ALS caused by pathogenic hnRNPA1 mutation.
Ubiquitin-positive inclusion containing Fused in Sarcoma (FUS) defines a new subtype of frontotemporal lobar degeneration (FTLD). FTLD is characterized by progressive alteration in cognitions and it ...preferentially affects the superficial layers of frontotemporal cortex. Mutation of FUS is linked to amyotrophic lateral sclerosis and to motor neuron disease with FTLD. To examine FUS pathology in FTLD, we developed the first mammalian animal model expressing human FUS with pathogenic mutation and developing progressive loss of memory. In FUS transgenic rats, ubiquitin aggregation and FUS mislocalization were developed primarily in the entorhinal cortex of temporal lobe, particularly in the superficial layers of affected cortex. Overexpression of mutant FUS led to Golgi fragmentation and mitochondrion aggregation. Intriguingly, aggregated ubiquitin was not colocalized with either fragmented Golgi or aggregated mitochondria, and neurons with ubiquitin aggregates were deprived of endogenous TDP-43. Agonists of peroxisome proliferator-activated receptor gamma (PPAR-γ) possess anti-glial inflammation effects and are also shown to preserve the dendrite and dendritic spines of cortical neurons in culture. Here we show that rosiglitazone, a PPAR-γ agonist, rescued the dendrites and dendritic spines of neurons from FUS toxicity and preserved rats' spatial memory. Our FUS transgenic rats would be useful to the mechanistic study of cortical dementia in FTLD. As rosiglitazone is clinically used to treat diabetes, our results would encourage immediate application of PPAR-γ agonists in treating patients with cortical dementia.
Protein inclusion is a prominent feature of neurodegenerative diseases including frontotemporal lobar degeneration (FTLD) that is characterized by the presence of ubiquitinated TDP‐43 inclusion. ...Presence of protein inclusions indicates an interruption to protein degradation machinery or the overload of misfolded proteins. In response to the increase in misfolded proteins, cells usually initiate a mechanism called unfolded protein response (UPR) to reduce misfolded proteins in the lumen of endoplasmic reticules. Here, we examined the effects of mutant TDP‐43 on the UPR in transgenic rats that express mutant human TDP‐43 restrictedly in the neurons of the forebrain. Over‐expression of mutant TDP‐43 in rats caused prominent aggregation of ubiquitin and remarkable fragmentation of Golgi complexes prior to neuronal loss. While ubiquitin aggregates and Golgi fragments were accumulating, neurons expressing mutant TDP‐43 failed to up‐regulate chaperones residing in the endoplasmic reticules and failed to initiate the UPR. Prior to ubiquitin aggregation and Golgi fragmentation, neurons were depleted of X‐box‐binding protein 1 (XBP1), a key player of UPR machinery. Although it remains to determine how mutation of TDP‐43 leads to the failure of the UPR, our data demonstrate that failure of the UPR is implicated in TDP‐43 pathogenesis.
Electromicroscopy reveals that ER is severely vacuolated in neurons while mitochondrial morphology is largely intact. ER likely is the preferable targets of degeneration caused by mutation in TDP‐43.
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