Axodendritic fibres of mouse cerebellar granule neurons exhibit a diversity of functional P2X receptors

Sánchez-Nogueiro, Jesús; Marín-García, Patricia; León, David; León-Otegui, Míriam; Salas, Elvira; Gómez‐Villafuertes, Rosa; Gualix, Javier; Miras-Portugal, Marı́a Teresa

doi:10.1016/j.neuint.2009.06.009

Cited by 12 publications

(4 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The expression of a variety of functional P2Y and P2X nucleotide receptors in primary cultures of cerebellar granule cells was already reported ( Hervas et al, 2003 , 2005 ; Leon et al, 2006 ; Sanchez-Nogueiro et al, 2009 , 2014 ). In order to investigate the presence of native VNUT in granule neurons, western blot and immunofluorescence studies were performed.…”

Section: Resultsmentioning

confidence: 82%

“…Primary cultures of cerebellar granule neurons are widely used to study neurotransmission, as these cells can develop the morphological, biochemical and electrophysiological characteristics of mature neurons in vitro . Indeed, they express a variety of functional P2Y and P2X nucleotide receptors, making them a suitable model to study the purinergic system ( Hervas et al, 2003 , 2005 ; Leon et al, 2006 ; Sanchez-Nogueiro et al, 2009 , 2014 ).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

et al. 2017

Self Cite

View full text Add to dashboard Cite

Adenosine triphosphate (ATP) is an important extracellular neurotransmitter that participates in several critical processes like cell differentiation, neuroprotection or axon guidance. Prior to its exocytosis, ATP must be stored in secretory vesicles, a process that is mediated by the Vesicular Nucleotide Transporter (VNUT). This transporter has been identified as the product of the SLC17A9 gene and it is prominently expressed in discrete brain areas, including the cerebellum. The main population of cerebellar neurons, the glutamatergic granule neurons, depends on purinergic signaling to trigger neuroprotective responses. However, while nucleotide receptors like P2X7 and P2Y13 are known to be involved in neuroprotection, the mechanisms that regulate ATP release in relation to such events are less clearly understood. In this work, we demonstrate that cerebellar granule cells express a functional VNUT that is involved in the regulation of ATP exocytosis. Numerous vesicles loaded with this nucleotide can be detected in these granule cells and are staining by the fluorescent ATP-marker, quinacrine. High potassium stimulation reduces quinacrine fluorescence in granule cells, indicating they release ATP via calcium dependent exocytosis. Specific subcellular markers were used to assess the localization of VNUT in granule cells, and the transporter was detected in both the axonal and somatodendritic compartments, most predominantly in the latter. However, co-localization with the specific lysosomal marker LAMP-1 indicated that VNUT can also be found in non-synaptic vesicles, such as lysosomes. Interestingly, the weak co-localization between VNUT and VGLUT1 suggests that the ATP and glutamate vesicle pools are segregated, as also observed in the cerebellar cortex. During post-natal cerebellar development, VNUT is found in granule cell precursors, co-localizing with markers of immature cells like doublecortin, suggesting that this transporter may be implicated in the initial stages of granule cell development.

show abstract

Section: Resultsmentioning

confidence: 82%

Section: Introductionmentioning

confidence: 99%

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

et al. 2017

Self Cite

View full text Add to dashboard Cite

show abstract

“…High concentrations of ivermectin were generally toxic to the fish, while concentrations below 250 nM caused hair cell death without overt morbidity; larvae swam normally and there was no significant increase in the overall number of apoptotic cells. In vertebrates, ivermectin primarily targets GABA receptors but can also act on glycinergic and purinergic receptors ( Soderlund et al, 1987 ; Sánchez-Nogueiro et al, 2009 ; Plested, 2016 ). By agonizing these targets, ivermectin generally acts to increase neural inhibition, consistent with literature reporting ivermectin neurotoxicity in rodents and other animals ( Trailovic and Nedeljkovic, 2011 ; Salam et al, 2022 ).…”

Section: Discussionmentioning

confidence: 99%

Putative COVID-19 therapies imatinib, lopinavir, ritonavir, and ivermectin cause hair cell damage: A targeted screen in the zebrafish lateral line

Coffin

Dale

Doppenberg

et al. 2022

Front. Cell. Neurosci.

View full text Add to dashboard Cite

The biomedical community is rapidly developing COVID-19 drugs to bring much-need therapies to market, with over 900 drugs and drug combinations currently in clinical trials. While this pace of drug development is necessary, the risk of producing therapies with significant side-effects is also increased. One likely side-effect of some COVID-19 drugs is hearing loss, yet hearing is not assessed during preclinical development or clinical trials. We used the zebrafish lateral line, an established model for drug-induced sensory hair cell damage, to assess the ototoxic potential of seven drugs in clinical trials for treatment of COVID-19. We found that ivermectin, lopinavir, imatinib, and ritonavir were significantly toxic to lateral line hair cells. By contrast, the approved COVID-19 therapies dexamethasone and remdesivir did not cause damage. We also did not observe damage from the antibiotic azithromycin. Neither lopinavir nor ritonavir altered the number of pre-synaptic ribbons per surviving hair cell, while there was an increase in ribbons following imatinib or ivermectin exposure. Damage from lopinavir, imatinib, and ivermectin was specific to hair cells, with no overall cytotoxicity noted following TUNEL labeling. Ritonavir may be generally cytotoxic, as determined by an increase in the number of TUNEL-positive non-hair cells following ritonavir exposure. Pharmacological inhibition of the mechanotransduction (MET) channel attenuated damage caused by lopinavir and ritonavir but did not alter imatinib or ivermectin toxicity. These results suggest that lopinavir and ritonavir may enter hair cells through the MET channel, similar to known ototoxins such as aminoglycoside antibiotics. Finally, we asked if ivermectin was ototoxic to rats in vivo. While ivermectin is not recommended by the FDA for treating COVID-19, many people have chosen to take ivermectin without a doctor’s guidance, often with serious side-effects. Rats received daily subcutaneous injections for 10 days with a clinically relevant ivermectin dose (0.2 mg/kg). In contrast to our zebrafish assays, ivermectin did not cause ototoxicity in rats. Our research suggests that some drugs in clinical trials for COVID-19 may be ototoxic. This work can help identify drugs with the fewest side-effects and determine which therapies warrant audiometric monitoring.

show abstract

“…Real-time PCR was conducted with SYBR Green (Roche) on a MyiQ Real-Time PCR System (Bio-Rad). The TaqMan probes for mouse EP1, EP2, EP3, EP4, P2Y2, and P2Y4 used in this study were purchased from Applied Biosystems and experiments for validation of amplification efficiency were performed for each TaqMan probe set [28, 29]. PCR thermocycling parameters were 95°C for 10 min, 40 cycles of 95°C for 15 s, and 60°C for 1 min.…”

Section: Methodsmentioning

confidence: 99%

Sustained Release of Prostaglandin E₂in Fibroblasts Expressing Ectopically Cyclooxygenase 2 Impairs P2Y-Dependent Ca²⁺-Mobilization

Pimentel-Santillana

Través

Pérez-Sen

et al. 2014

Mediators of Inflammation

Self Cite

View full text Add to dashboard Cite

The nucleotide uridine trisphosphate (UTP) released to the extracellular milieu acts as a signaling molecule via activation of specific pyrimidine receptors (P2Y). P2Y receptors are G protein-coupled receptors expressed in many cell types. These receptors mediate several cell responses and they are involved in intracellular calcium mobilization. We investigated the role of the prostanoid PGE2 in P2Y signaling in mouse embryonic fibroblasts (MEFs), since these cells are involved in different ontogenic and physiopathological processes, among them is tissue repair following proinflammatory activation. Interestingly, Ca2+-mobilization induced by UTP-dependent P2Y activation was reduced by PGE2 when this prostanoid was produced by MEFs transfected with COX-2 or when PGE2 was added exogenously to the culture medium. This Ca2+-mobilization was important for the activation of different metabolic pathways in fibroblasts. Moreover, inhibition of COX-2 with selective coxibs prevented UTP-dependent P2Y activation in these cells. The inhibition of P2Y responses by PGE2 involves the activation of PKCs and PKD, a response that can be suppressed after pharmacological inhibition of these protein kinases. In addition to this, PGE2 reduces the fibroblast migration induced by P2Y-agonists such as UTP. Taken together, these data demonstrate that PGE2 is involved in the regulation of P2Y signaling in these cells.

show abstract

Axodendritic fibres of mouse cerebellar granule neurons exhibit a diversity of functional P2X receptors

Cited by 12 publications

References 40 publications

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

Putative COVID-19 therapies imatinib, lopinavir, ritonavir, and ivermectin cause hair cell damage: A targeted screen in the zebrafish lateral line

Sustained Release of Prostaglandin E₂in Fibroblasts Expressing Ectopically Cyclooxygenase 2 Impairs P2Y-Dependent Ca²⁺-Mobilization

Contact Info

Product

Resources

About

Axodendritic fibres of mouse cerebellar granule neurons exhibit a diversity of functional P2X receptors

Cited by 12 publications

References 40 publications

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

Putative COVID-19 therapies imatinib, lopinavir, ritonavir, and ivermectin cause hair cell damage: A targeted screen in the zebrafish lateral line

Sustained Release of Prostaglandin E2in Fibroblasts Expressing Ectopically Cyclooxygenase 2 Impairs P2Y-Dependent Ca2+-Mobilization

Contact Info

Product

Resources

About

Sustained Release of Prostaglandin E₂in Fibroblasts Expressing Ectopically Cyclooxygenase 2 Impairs P2Y-Dependent Ca²⁺-Mobilization