Tissue-nonspecific alkaline phosphatase promotes axonal growth of hippocampal neurons

Axonal growth is essential for establishing neuronal circuits during brain development and for regenerative processes in the adult brain. Unfortunately, the extracellular signals controlling axonal growth are poorly understood. Here we report that a reduction in extracellular ATP levels by tissue-nonspecific alkaline phosphatase (TNAP) is essential for the development of neuritic processes by cultured hippocampal neurons. Selective blockade of TNAP activity with levamisole or specific TNAP knockdown with short hairpin RNA interference inhibited the growth and branching of principal axons, whereas addition of alkaline phosphatase (ALP) promoted axonal growth. Neither activation nor inhibition of adenosine receptors affected the axonal growth, excluding the contribution of extracellular adenosine as a potential hydrolysis product of extracellular ATP to the TNAP-mediated effects. TNAP was colocalized at axonal growth cones with ionotropic ATP receptors (P2X7 receptor), whose activation inhibited axonal growth. Additional analyses suggested a close functional interrelation of TNAP and P2X7 receptors whereby TNAP prevents P2X7 receptor activation by hydrolyzing ATP in the immediate environment of the receptor. Furthermore inhibition of P2X7 receptor reduced TNAP expression, whereas addition of ALP enhanced P2X7 receptor expression. Our results demonstrate that TNAP, regulating both ligand availability and protein expression of P2X7 receptor, is essential for axonal development

P2X7 receptor in epilepsy; role in pathophysiology and potential targeting for seizure control.

The P2X7 receptor is an ATP-gated non-selective cation-permeable ionotropic receptor selectively expressed in neurons and glia in the brain. Activation of the P2X7 receptor has been found to modulate neuronal excitability in the hippocampus and it has also been linked to microglia activation and neuroinflammatory responses. Accordingly, interest developed on the P2X7 receptor in disorders of the nervous system, including epilepsy. Studies show that expression of the P2X7 receptor is elevated in damaged regions of the brain after prolonged seizures (status epilepticus) in both neurons and glia. P2X7 receptor expression is also increased in the hippocampus in experimental epilepsy. Recent data show that mice lacking the P2X7 receptor display altered susceptibility to status epilepticus and that drugs targeting the P2X7 receptor have potent anticonvulsant effects. Together, this suggests that P2X7 receptor ligands may be useful adjunctive treatments for refractory status epilepticus or perhaps pharmacoresistant epilepsy. This review summarizes the evidence of P2X7 receptor involvement in the pathophysiology of epilepsy and the potential of drugs targeting this receptor for seizure control

Functional characterization of P2Y1 and P2X4 receptors in human neuroblastoma SK-N-MC cells

Nucleotides are important signalling molecules in both the peripheral and central nervous system. However, the in vitro study of their receptors can be hampered by the heterogeneity of primary neuronal cultures. The use of clonal neuroblastoma cell lines allows to circumvent this difficulty, so these lines are often used as a model to analyze the properties, regulation and physiological role of nucleotide receptors in neural tissues. Expression studies indicated the presence of P2Y1, P2Y6, P2Y11, P2Y13, P2X1, P2X4, P2X5, P2X6 and P2X7 proteins in SK-N-MC cells. Functional analyses showed transient [Ca2+]i increases upon application of ADP, 2-MeSADP or ADPβS. Responses to these agonists seem to be mediated by a P2Y1 receptor, as demonstrated by the almost complete blockade exerted by the P2Y1-selective antagonist MRS2179. ATP was also able to induce [Ca2+]i increases in SK-N-MC cells. Responses to ATP were partially blocked by MRS2179 and the P2X antagonist TNP-ATP, thus suggesting that ATP can interact with two different P2 receptors: a P2Y1 receptor, inhibited by MRS2179, and a TNP-ATP sensitive P2X receptor. To characterize the P2X receptor responsible for the MRS2179-resistant component of the ATP response, we analyze the effect of several P2X agonists on [Ca2+]i. Cells did not show responses to either α,β-meATP or BzATP, although [Ca2+]i increases could be observed when cells were challenged with CTP. Both the response to CTP and the MRS2179-resistant component of ATP response were potentiated by ivermectin. Such pharmacological profile is consistent with the presence of a functional P2X4 receptor in SK-N-MC cell lineLos nucleótidos son importantes moléculas señalizadoras en el sistema nervioso. El estudio in vitro de sus receptores puede verse obstaculizado por la heterogeneidad de los cultivos neuronales. El uso de líneas celulares de neuroblastoma permite eludir esta dificultad y dichas líneas se utilizan frecuentemente como un modelo con el que analizar las propiedades, regulación y función de los receptores de nucleótidos en tejidos neurales. Estudios de expresión indicaron la presencia de proteínas P2Y1, P2Y6, P2Y11, P2Y13, P2X1, P2X4, P2X5, P2X6 y P2X7 en las células SK-N-MC. Análisis funcionales mostraron incrementos transitorios de [Ca2+]i tras la aplicación de ADP, 2- MeSADP o ADPβS, respuestas que parecen estar mediadas a través un receptor P2Y1, como se pone de manifiesto por el bloqueo casi total ejercido por el antagonista selectivo P2Y1, MRS2179. El ATP también indujo incrementos de [Ca2+]i en las células SK-N-MC, siendo su respuesta parcialmente bloqueada por MRS2179 y por el antagonista P2X TNP-ATP, lo que sugiere que el ATP puede interactuar con dos receptores P2 diferentes: un receptor P2Y1, inhibido por MRS2179, y un receptor P2X sensible a TNP-ATP. Se caracterizó el receptor P2X analizando el efecto de varios agonistas en la [Ca2+]i. Ninguna célula mostró respuestas a αβ- meATP o BzATP, aunque se observaron incrementos de [Ca2+]i cuando las células fueron estimuladas con CTP. Tanto la respuesta a CTP como el componente de la respuesta a ATP resistente a MRS2179, se potenciaron en presencia de ivermectina. Todos estos datos sugieren la presencia de un receptor P2X4 funcional en las células SK-N-MCThis work was supported by research grants from Ministerio de Ciencia e Innovación (BFU2014-53654-P) and Comunidad de Madrid (S2013/ICE-2958 BRADECM

P2Y receptors (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

P2Y receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on P2Y Receptors [3, 5]) are activated by the endogenous ligands ATP, ADP, uridine triphosphate, uridine diphosphate and UDP-glucose. The relationship of many of the cloned receptors to endogenously expressed receptors is not yet established and so it might be appropriate to use wording such as 'uridine triphosphate-preferring (or ATP-, etc.) P2Y receptor' or 'P2Y1-like', etc., until further, as yet undefined, corroborative criteria can be applied [46, 109, 187, 375, 388].Clinically used drugs acting on these receptors include the dinucleoside polyphosphate diquafosol, agonist of the P2Y2 receptor subtype, approved in Japan for the management of dry eye disease [236], and the P2Y12 receptor antagonists prasugrel, ticagrelor and cangrelor, all approved as antiplatelet drugs [52, 316]

P2Y receptors in GtoPdb v.2023.1

P2Y receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on P2Y Receptors [3, 5, 189]) are activated by the endogenous ligands ATP, ADP, UTP, UDP, UDP-glucose and adenosine. The eight mammalian P2Y receptors are activated by distinct nucleotides: P2Y1, P2Y11, P2Y12 and P2Y13 are activated by adenosine-nucleotides; P2Y2, P2Y4 can be activated by both adenosine and uridine nucleotides, with some species-specific differences; P2Y6 is mainly activated by UDP; P2Y14 is preferentially activated by sugar-uracil nucleotides. The missing numbers in the receptor nomenclature refer either to non-mammalian orthologs or receptors having some sequence homology to P2Y receptors but for which there is no functional evidence of responsiveness to nucleotides [380]. Based on their G protein coupling P2Y receptors can be divided into two subfamilies: P2Y1, P2Y2, P2Y4, P2Y6 and P2Y11 receptors couple via Gq proteins to stimulate phospholipase C followed by increases in inositol phosphates and mobilization of Ca2+ from intracellular stores. P2Y11 receptors couple in addition to Gs proteins followed by increased adenylate cyclase activity. In contrast, P2Y12, P2Y13, and P2Y14 receptors signal primarily through activation of Gi proteins and inhibition of adenylate cyclase activity or control of ion channel activity [380]. Clinically used drugs acting on these receptors include the dinucleoside polyphosphate diquafosol, agonist of the P2Y2 receptor subtype, approved in Japan and South Korea for the management of dry eye disease [238], and the P2Y12 receptor antagonists prasugrel, ticagrelor and cangrelor, all approved as antiplatelet drugs [52, 320]

P2Y receptors in GtoPdb v.2021.3

P2Y receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on P2Y Receptors [3, 5, 192]) are activated by the endogenous ligands ATP, ADP, uridine triphosphate, uridine diphosphate and UDP-glucose. The relationship of many of the cloned receptors to endogenously expressed receptors is not yet established and so it might be appropriate to use wording such as 'uridine triphosphate-preferring (or ATP-, etc.) P2Y receptor' or 'P2Y1-like', etc., until further, as yet undefined, corroborative criteria can be applied [47, 110, 190, 383, 396]. Clinically used drugs acting on these receptors include the dinucleoside polyphosphate diquafosol, agonist of the P2Y2 receptor subtype, approved in Japan for the management of dry eye disease [241], and the P2Y12 receptor antagonists prasugrel, ticagrelor and cangrelor, all approved as antiplatelet drugs [53, 323]

Changes in P2Y Purinergic Receptor Expression in the Ciliary Body in a Murine Model of Glaucoma

Glaucoma is a neuropathology, often accompanied by an elevated intraocular pressure (IOP), which can lead to blindness. Since DBA/2J mice develop glaucoma, several studies of the physiopathology of glaucoma have been reported in this animal model. It is also known that purinergic receptors are involved in the pathology of glaucoma by controlling aqueous humor production and drainage and therefore controlling IOP. There are no studies on purinergic receptors in the DBA/2J model of glaucoma and their relation to the development of the pathology, so the aim of this study was to make an approach to the purinergic mechanisms involved in glaucoma. All the experiments were performed using DBA/2J and C57BL/6J mice and investigating P2Y1, P2Y2, and P2Y6 receptors. IOP measurements were made with a non-invasive rebound tonometer, and animals were instilled with diadenosine tetraphosphate (Ap4A) and the corresponding purinergic antagonists in order to see their effects on IOP. The expression of mRNA for P2Y1, P2Y2, and P2Y6 purinergic receptors was carried out by quantitative real-time PCR. Additionally, P2Y-receptor expression was performed by immunohistochemical techniques carried out on the ciliary processes. The results showed that IOP decreases when Ap4A was instilled and that the expressions of the analyzed purinergic receptors were stable throughout all the ages under study in the C57BL/6J mice (control mice). On the other hand, there were significant changes in the purinergic receptor expression in DBA/2J suggesting that elevated IOP in these animals could be related to an increase of P2Y2 expression and a decrease in P2Y1 receptors