Expression of Cholinergic Markers and Characterization of Splice Variants during Ontogenesis of Rat Dorsal Root Ganglia Neurons

Dorsal root ganglia (DRG) neurons synthesize acetylcholine (ACh), in addition to their peptidergic nature. They also release ACh and are cholinoceptive, as they express cholinergic receptors. During gangliogenesis, ACh plays an important role in neuronal differentiation, modulating neuritic outgrowth and neurospecific gene expression. Starting from these data, we studied the expression of choline acetyltransferase (ChAT) and vesicular ACh transporter (VAChT) expression in rat DRG neurons. ChAT and VAChT genes are arranged in a “cholinergic locus”, and several splice variants have been described. Using selective primers, we characterized splice variants of these cholinergic markers, demonstrating that rat DRGs express R1, R2, M, and N variants for ChAT and V1, V2, R1, and R2 splice variants for VAChT. Moreover, by RT-PCR analysis, we observed a progressive decrease in ChAT and VAChT transcripts from the late embryonic developmental stage (E18) to postnatal P2 and P15 and in the adult DRG. Interestingly, Western blot analyses and activity assays demonstrated that ChAT levels significantly increased during DRG ontogenesis. The modulated expression of different ChAT and VAChT splice variants during development suggests a possible differential regulation of cholinergic marker expression in sensory neurons and confirms multiple roles for ACh in DRG neurons, both in the embryo stage and postnatally.

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Suppression of KCNQ/M Potassium Channel in Dorsal Root Ganglia Neurons Contributes to the Development of Osteoarthritic Pain

Pharmacology ◽

10.1159/000496422 ◽

2019 ◽

Vol 103 (5-6) ◽

pp. 257-262 ◽

Cited By ~ 3

Author(s):

Fan Zhang ◽

Yani Liu ◽

Dandan Zhang ◽

Xizhenzi Fan ◽

Decheng Shao ◽

...

Keyword(s):

Dorsal Root Ganglia ◽

Dorsal Root ◽

Strong Impact ◽

Drg Neurons ◽

M Current ◽

Mechanical Threshold ◽

Dorsal Root Ganglia Neurons ◽

M Channels ◽

Osteoarthritic Pain

Osteoarthritic pain has a strong impact on patients’ quality of life. Understanding the pathogenic mechanisms underlying osteoarthritic pain will likely lead to the development of more effective treatments. In the present study of osteoarthritic model rats, we observed a reduction of M-current density and a remarkable decrease in the levels of KCNQ2 and KCNQ3 proteins and mRNAs in dorsal root ganglia (DRG) neurons, which were associated with hyperalgesic behaviors. The activation of KCNQ/M channels with flupirtine significantly increased the mechanical threshold and prolonged the withdrawal latency of osteoarthritic model rats at 3–14 days after model induction, and all effects of flupirtine were blocked by KCNQ/M-channel antagonist, XE-991. Together, these results indicate that suppression of KCNQ/M channels in primary DRG neurons plays a crucial role in the development of osteoarthritic pain.

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Two TTX-resistant Na+ currents in mouse colonic dorsal root ganglia neurons and their role in colitis-induced hyperexcitability

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00154.2004 ◽

2004 ◽

Vol 287 (4) ◽

pp. G845-G855 ◽

Cited By ~ 113

Author(s):

Michael J. Beyak ◽

Noor Ramji ◽

Karmen M. Krol ◽

Michael D. Kawaja ◽

Stephen J. Vanner

Keyword(s):

Sensory Neurons ◽

Dorsal Root Ganglia ◽

Dorsal Root ◽

Neuronal Excitability ◽

Distal Colon ◽

Inactivation Kinetics ◽

Drg Neurons ◽

Trinitrobenzenesulfonic Acid ◽

Na Currents ◽

Dorsal Root Ganglia Neurons

The composition of Na+ currents in dorsal root ganglia (DRG) neurons depends on their neuronal phenotype and innervation target. Two TTX-resistant (TTX-R) Na+ currents [voltage-gated Na channels (Na v)] have been described in small DRG neurons; one with slow inactivation kinetics (Na v1.8) and the other with persistent kinetics (Na v1.9), and their modulation has been implicated in inflammatory pain. This has not been studied in neurons projecting to the colon. This study examined the relative importance of these currents in inflammation-induced changes in a mouse model of inflammatory bowel disease. Colonic sensory neurons were retrogradely labeled, and colitis was induced by instillation of trinitrobenzenesulfonic acid (TNBS) into the lumen of the distal colon. Seven to ten days later, immunohistochemical properties were characterized in controls, and whole cell recordings were obtained from small (<40 pF) labeled DRG neurons from control and TNBS animals. Most neurons exhibited both fast TTX-sensitive (TTX-S)- and slow TTX-R-inactivating Na+ currents, but persistent TTX-R currents were uncommon (<15%). Most labeled neurons were CGRP (79%), tyrosine kinase A (trkA) (84%) immunoreactive, but only a small minority bind IB4 (14%). TNBS-colitis caused ulceration, thickening of the colon and significantly increased neuronal excitability. The slow TTX-R-inactivating Na current density (Na v1.8) was significantly increased, but other Na currents were unaffected. Most small mouse colonic sensory neurons are CGRP, trkA immunoreactive, but not isolectin B4 reactive and exhibit fast TTX-S, slow TTX-R, but not persistent TTX-R Na+ currents. Colitis-induced hyperexcitability is associated with increased slow TTX-R (Na v1.8) Na+ current. Together, these findings suggest that colitis alters trkA-positive neurons to preferentially increase slow TTX-R Na+ (Na v1.8) currents.

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Calcitriol increases frataxin levels and restores altered markers in cell models of Friedreich Ataxia

10.1101/2020.04.09.034017 ◽

2020 ◽

Author(s):

E. Britti ◽

F. Delaspre ◽

M. Medina-Carbonero ◽

A. Sanz ◽

M. Llovera ◽

...

Keyword(s):

Dorsal Root Ganglia ◽

Dorsal Root ◽

Apoptotic Cell ◽

Primary Cultures ◽

Active Form ◽

Friedreich Ataxia ◽

Mitochondrial Protein ◽

Drg Neurons ◽

Dorsal Root Ganglia Neurons ◽

Neurite Degeneration

ABSTRACTFriedreich Ataxia (FA) is a neurodegenerative disease caused by the deficiency of frataxin, a mitochondrial protein. In primary cultures of dorsal root ganglia neurons, we showed that frataxin depletion resulted in decreased levels of the mitochondrial calcium exchanger NCLX, neurite degeneration and apoptotic cell death. Here we describe that frataxin-deficient dorsal root ganglia neurons display low levels of ferredoxin 1, a mitochondrial Fe/S cluster-containing protein that interacts with frataxin and, interestingly, is essential for the synthesis of calcitriol, the active form of vitamin D. We provide data that calcitriol supplementation, used at nanomolar concentrations, is able to reverse the molecular and cellular markers altered in DRG neurons. Calcitriol is able to recover both ferredoxin 1 and NCLX levels and restores mitochondrial membrane potential. Accordingly, apoptotic markers and neurite degeneration are reduced resulting in cell survival recovery with calcitriol supplementation. All these beneficial effects would be explained by the finding that calcitriol is able to increase the mature frataxin levels in both, frataxin-deficient DRG neurons and cardiomyocytes; remarkably, this increase also occurs in lymphoblastoid cell lines derived from FA patients. In conclusion, these results provide molecular bases to consider calcitriol for an easy and affordable therapeutic approach for FA patients.

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Calcitriol increases frataxin levels and restores mitochondrial function in cell models of Friedreich Ataxia

Biochemical Journal ◽

10.1042/bcj20200331 ◽

2020 ◽

Author(s):

Elena Britti ◽

Fabien Delaspre ◽

Arabela Sanz ◽

Marta Medina-Carbonero ◽

Marta Llovera ◽

...

Keyword(s):

Dorsal Root Ganglia ◽

Mitochondrial Function ◽

Dorsal Root ◽

Primary Cultures ◽

Active Form ◽

Friedreich Ataxia ◽

Mitochondrial Protein ◽

Drg Neurons ◽

Dorsal Root Ganglia Neurons ◽

Neurite Degeneration

Friedreich Ataxia (FA) is a neurodegenerative disease caused by the deficiency of frataxin, a mitochondrial protein. In primary cultures of dorsal root ganglia neurons, we showed that frataxin depletion resulted in decreased levels of the mitochondrial calcium exchanger NCLX, neurite degeneration and apoptotic cell death. Here we describe that frataxin-deficient dorsal root ganglia neurons display low levels of ferredoxin 1, a mitochondrial Fe/S cluster-containing protein that interacts with frataxin and, interestingly, is essential for the synthesis of calcitriol, the active form of vitamin D. We provide data that calcitriol supplementation, used at nanomolar concentrations, is able to reverse the molecular and cellular markers altered in DRG neurons. Calcitriol is able to recover both ferredoxin 1 and NCLX levels and restores mitochondrial membrane potential indicating an overall mitochondrial function improvement. Accordingly, reduction of apoptotic markers and neurite degeneration was observed and, as a result, cell survival was also recovered. All these beneficial effects would be explained by the finding that calcitriol is able to increase the mature frataxin levels in both, frataxin-deficient DRG neurons and cardiomyocytes; remarkably, this increase also occurs in lymphoblastoid cell lines derived from FA patients. In conclusion, these results provide molecular bases to consider calcitriol for an easy and affordable therapeutic approach for FA patients.

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