Purinergic P2X receptors (P2X) are ATP-gated ion channels that are broadly expressed in the brain, particularly in the hypothalamus. As ionic channels with high permeability to calcium, P2X play an important and active role in neural functions. The hypothalamus contains a number of small nuclei with many molecularly defined types of peptidergic neurons that affect a wide range of physiological functions, including water balance, blood pressure, metabolism, food intake, circadian rhythm, childbirth and breastfeeding, growth, stress, body temperature, and multiple behaviors. P2X are expressed in hypothalamic neurons, astrocytes, tanycytes, and microvessels. This review focuses on cell-type specific expression of P2X in the most important hypothalamic nuclei, such as the supraoptic nucleus (SON), paraventricular nucleus (PVN), suprachiasmatic nucleus (SCN), anteroventral periventricular nucleus (AVPV), anterior hypothalamic nucleus (AHN), arcuate nucleus (ARC), ventromedial hypothalamic nucleus (VMH), dorsomedial hypothalamic nucleus (DMH), tuberomammillary nucleus (TMN), and lateral hypothalamic area (LHA).> The review also notes the possible role of P2X and extracellular ATP in specific hypothalamic functions. The literature summarized here shows that purinergic signaling is involved in the control of the hypothalamic-pituitary endocrine system, the hypothalamic-neurohypophysial system, the circadian systems and nonendocrine hypothalamic functions.

• Klíčová slova
• P2X, arcuate nucleus (ARC), extracellular ATP, hypothalamus, paraventricular nucleus (PVN), suprachiasmatic nucleus (SCN), supraoptic nucleus (SON),
• MeSH
• adenosintrifosfát metabolismus   MeSH
• astrocyty metabolismus   MeSH
• hypothalamus * metabolismus   cytologie   MeSH
• lidé MeSH
• neurony metabolismus   MeSH
• purinergní receptory P2X * metabolismus   genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• adenosintrifosfát MeSH
• purinergní receptory P2X * MeSH

Purinergic P2X receptors (P2XR), activated by extracellular adenosine 5'-triphosphate (ATP), represent a specific type of ligand-gated ion channels. They form functional trimeric homomers or heteromers which are nonselectively cation-permeable after receptor activation. P2X receptors are widely expressed in excitable and nonexcitable tissues and are involved in many physiological and pathophysiological processes such as platelet aggregation, contraction of smooth muscle, immune responses, cell proliferation and apoptosis or neurotransmission. In mammals, seven P2X subunits (P2X1-P2X7) have been identified. They differ mainly in distribution, pharmacological profile and kinetics of ATP-induced responses. The subtype P2X7 is the most specific in the P2X family and widely differs from other P2X subtypes.

• MeSH
• adenosintrifosfát metabolismus   MeSH
• lidé MeSH
• molekulární modely MeSH
• podjednotky proteinů MeSH
• purinergní receptory P2X7 chemie   metabolismus   MeSH
• savci MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• adenosintrifosfát MeSH
• podjednotky proteinů MeSH
• purinergní receptory P2X7 MeSH

Purinergic P2X receptors represent a novel structural type of ligand-gated ion channels activated by extracellular ATP. So far, seven P2X receptor subunits have been found in excitable as well as non-excitable tissues. Little is known about their structure, mechanism of channel opening, localization, and role in the central nervous system. The aim of this work is to summarize recent investigations and describe our contribution to elucidating the structure of the ATP binding site and transmembrane domains of the P2X receptor, we also discuss the expression and physiological roles played by the ATP and P2X receptors in the anterior pituitary and hypothalamus.

• MeSH
• adenosintrifosfát metabolismus   MeSH
• gonadotropní buňky metabolismus   MeSH
• hypofýza metabolismus   MeSH
• hypothalamus metabolismus   MeSH
• ivermektin chemie   farmakologie   MeSH
• kvarterní struktura proteinů MeSH
• lidé MeSH
• molekulární modely MeSH
• neuroglie metabolismus   MeSH
• neurony metabolismus   MeSH
• purinergní receptory P2 chemie   účinky léků   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• terciární struktura proteinů MeSH
• vazba proteinů MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• adenosintrifosfát MeSH
• ivermektin MeSH
• purinergní receptory P2 MeSH

Neurosteroids are steroids synthesized de novo in the brain from cholesterol in an independent manner from peripheral steroid sources. The term "neuroactive steroid" includes all steroids independent of their origin, and newly synthesized analogs of neurosteroids that modify neuronal activities. In vivo application of neuroactive steroids induces potent anxiolytic, antidepressant, anticonvulsant, sedative, analgesic and amnesic effects, mainly through interaction with the γ-aminobutyric acid type-A receptor (GABAAR). However, neuroactive steroids also act as positive or negative allosteric regulators on several ligand-gated channels including N-methyl-d-aspartate receptors (NMDARs), nicotinic acetylcholine receptors (nAChRs) and ATP-gated purinergic P2X receptors. Seven different P2X subunits (P2X1-7) can assemble to form homotrimeric or heterotrimeric ion channels permeable for monovalent cations and calcium. Among them, P2X2, P2X4, and P2X7 are the most abundant within the brain and can be regulated by neurosteroids. Transmembrane domains are necessary for neurosteroid binding, however, no generic motif of amino acids can accurately predict the neurosteroid binding site for any of the ligand-gated ion channels including P2X. Here, we will review what is currently known about the modulation of rat and human P2X by neuroactive steroids and the possible structural determinants underlying neurosteroid-induced potentiation and inhibition of the P2X2 and P2X4 receptors. This article is part of the Special Issue on "Purinergic Signaling: 50 years".

• MeSH
• adenosintrifosfát metabolismus   MeSH
• iontové kanály řízené ligandy * metabolismus   MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• mozek metabolismus   MeSH
• neurosteroidy * MeSH
• purinergní receptory P2X metabolismus   MeSH
• purinergní receptory P2X2 metabolismus   MeSH
• vazebná místa MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• adenosintrifosfát MeSH
• iontové kanály řízené ligandy * MeSH
• neurosteroidy * MeSH
• purinergní receptory P2X MeSH
• purinergní receptory P2X2 MeSH

The family of ATP-gated purinergic P2X receptors comprises seven bunits (P2X1-7) that are unevenly distributed in the central and peripheral nervous systems as well as other organs. Endogenous modulators of P2X receptors are phospholipids, steroids and neurosteroids. Here, we analyzed whether bile acids, which are natural products derived from cholesterol, affect P2X receptor activity. We examined the effects of primary and secondary bile acids and newly synthesized derivatives of lithocholic acid on agonist-induced responses in HEK293T cells expressing rat P2X2, P2X4 and P2X7 receptors. Electrophysiology revealed that low micromolar concentrations of lithocholic acid and its structural analog 4-dafachronic acid strongly inhibit ATP-stimulated P2X2 but potentiate P2X4 responses, whereas primary bile acids and other secondary bile acids exhibit no or reduced effects only at higher concentrations. Agonist-stimulated P2X7 responses are significantly potentiated by lithocholic acid at moderate concentrations. Structural modifications of lithocholic acid at positions C-3, C-5 or C-17 abolish both inhibitory and potentiation effects to varying degrees, and the 3α-hydroxy group contributes to the ability of the molecule to switch between potentiation and inhibition. Lithocholic acid allosterically modulates P2X2 and P2X4 receptor sensitivity to ATP, reduces the rate of P2X4 receptor desensitization and antagonizes the effect of ivermectin on P2X4 receptor deactivation. Alanine-scanning mutagenesis of the upper halve of P2X4 transmembrane domain-1 revealed that residues Phe48, Val43 and Tyr42 are important for potentiating effect of lithocholic acid, indicating that modulatory sites for lithocholic acid and ivermectin partly overlap. Lithocholic acid also inhibits ATP-evoked currents in pituitary gonadotrophs expressing native P2X2, and potentiates ATP currents in nonidentified pituitary cells expressing P2X4 receptors. These results indicate that lithocholic acid is a bioactive steroid that may help to further unveil the importance of the P2X2, and P2X4 receptors in many physiological processes.

• Klíčová slova
• ATP, Allosteric modulation, Bile acids, Ivermectin, Lithocholic acid, Purinergic P2X receptors,
• MeSH
• adenohypofýza cytologie   účinky léků   fyziologie   MeSH
• agonisté purinergního receptoru P2X farmakologie   MeSH
• antagonisté purinergních receptorů P2X farmakologie   MeSH
• gating iontového kanálu účinky léků   MeSH
• HEK293 buňky MeSH
• hypothalamus cytologie   účinky léků   fyziologie   MeSH
• kyselina lithocholová analogy a deriváty   farmakologie   MeSH
• lidé MeSH
• neurony účinky léků   fyziologie   MeSH
• potkani Wistar MeSH
• purinergní receptory P2X2 fyziologie   MeSH
• purinergní receptory P2X4 fyziologie   MeSH
• purinergní receptory P2X7 fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• agonisté purinergního receptoru P2X MeSH
• antagonisté purinergních receptorů P2X MeSH
• kyselina lithocholová MeSH
• purinergní receptory P2X2 MeSH
• purinergní receptory P2X4 MeSH
• purinergní receptory P2X7 MeSH

The functional relevance of aromatic residues in the upper part of the transmembrane domain-1 of purinergic P2X receptors (P2XRs) was examined. Replacement of the conserved Tyr residue with Ala had a receptor-specific effect: the P2X1R was non-functional, the P2X2R, P2X4R, and P2X3R exhibited enhanced sensitivity to ATP and alphabeta-meATP accompanied by prolonged decay of current after washout of agonists, and the P2X7R sensitivity for agonists was not affected, though decay of current was delayed. The replacement of the P2X4R-Tyr42 with other amino acids revealed the relevance of an aromatic residue at this position. Mutation of the neighboring Phe and ipsilateral Tyr/Trp residues, but not the contralateral Phe residue, also affected the P2X2R, P2X3R, and P2X4R function. Double mutation of ipsilateral Tyr42 and Trp46 P2X4R residues restored receptor function, whereas the corresponding P2X2R double mutant was not functional. In contrast, mutation of the contralateral Phe48 residue in the P2X4R-Y42A mutant had no effect. These results indicate that aromatic residues in the upper part of TM1 play important roles in the three-dimensional structure of the P2XRs and that they are required not only for ion conductivity but also for specificity of agonist binding and/or channel gating.

• MeSH
• adenosintrifosfát analogy a deriváty   farmakologie   MeSH
• aminokyseliny aromatické genetika   metabolismus   MeSH
• biofyzika MeSH
• elektrická stimulace MeSH
• konformace proteinů MeSH
• lidé MeSH
• membránové potenciály účinky léků   genetika   MeSH
• metoda terčíkového zámku metody   MeSH
• mutageneze genetika   MeSH
• purinergní receptory P2 klasifikace   genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• signální transdukce účinky léků   fyziologie   MeSH
• terciární struktura proteinů genetika   fyziologie   MeSH
• transfekce metody   MeSH
• transformované buněčné linie MeSH
• vazba proteinů genetika   fyziologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zelené fluorescenční proteiny genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• adenosintrifosfát MeSH
• alpha,beta-methyleneadenosine 5'-triphosphate MeSH Prohlížeč
• aminokyseliny aromatické MeSH
• enhanced green fluorescent protein MeSH Prohlížeč
• purinergní receptory P2 MeSH
• zelené fluorescenční proteiny MeSH

The purpose of this review is to present current knowledge regarding purinergic receptors and their subtypes. The main endogenous ligans for these receptors are adenosine and ATP which are released from cells and neurons under various pathophysiological conditions, and adenine nucleotides which are released as contransmiters together with noradrenaline, acetylcholine and substance P. Purinergic receptors which are present in various tissues and mediate numerous physiological effects can be divided into two main groups, P1 (for adenosine) and P2 (for adenine nucleotides) receptors. Both these types are further divided into subtypes. P1 receptors are better characterized than P2 receptors whose classification must be regarded as tentative rather than definitive. P1 receptors are named according to their natural ligand adenosine as A1, A2a, A2b, A3 and possibly A4 receptors. Their characteristics are summarized in tables which also present the main selective agonists and antagonists of their receptors. Since the classification of P2 receptors is still tenative, their nomenclatue does not follow the exact rules which are provided by IUPHAR. P2 receptors are subdivided into these subtypes: P2X, P2Y, P2U, P2T, P2Z and P2D. The article also lists the main agonists and antagonists for these receptors. The introduction of new selective agonists and antagonists not only helps to classify various receptors subtypes of purinoceptors but it also has a big therapeutic potential for various diseases.

• MeSH
• lidé MeSH
• purinergní receptory * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• purinergní receptory * MeSH

Ivermectin (IVM), a large macrocyclic lactone, specifically enhances P2X(4) receptor-channel function by interacting with residues of transmembrane (TM) helices in the open conformation state. In this paper, we used cysteine-scanning mutagenesis of rat P2X(4)-TMs to identify and map residues of potential importance for channel gating and interaction with IVM. The receptor function was unchanged by mutations in 29 different residues, and among them, the IVM effects were altered in Gln(36), Leu(40), Val(43), Val(47), Trp(50), Asn(338), Gly(342), Leu(346), Ala(349), and Ile(356) mutants. The substitution-sensitive Arg(33) and Cys(353) mutants could also be considered as IVM-sensitive hits. The pattern of these 12 residues was consistent with helical topology of both TMs, with every third or fourth amino acid affected by substitution. These predominantly hydrophobic-nonpolar residues are also present in the IVM-sensitive Schistosoma mansoni P2X subunit. They lie on the same side of their helices and could face lipids in the open conformation state and provide the binding pocket for IVM. In contrast, the IVM-independent hits Met(31), Tyr(42), Gly(45), Val(49), Gly(340), Leu(343), Ala(344), Gly(347), Thr(350), Asp(354), and Val(357) map on the opposite side of their helices, probably facing the pore of receptor or protein and playing important roles in gating.

• MeSH
• antiparazitární látky metabolismus   MeSH
• buněčné linie MeSH
• gating iontového kanálu * MeSH
• ivermektin metabolismus   MeSH
• konformace proteinů * MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• metoda terčíkového zámku MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• mutace MeSH
• purinergní receptory P2 chemie   genetika   metabolismus   MeSH
• purinergní receptory P2X4 MeSH
• sekvence aminokyselin MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• antiparazitární látky MeSH
• ivermektin MeSH
• P2RX4 protein, human MeSH Prohlížeč
• P2rx4 protein, rat MeSH Prohlížeč
• purinergní receptory P2 MeSH
• purinergní receptory P2X4 MeSH

P2X7 receptors, a subtype of ATP-gated cation channel, have gained attention due to their involvement in inflammatory and neurodegenerative diseases, chronic pain, and cancer. However, despite extensive medicinal chemistry efforts, no P2X7 antagonists have reached clinical approval due to suboptimal pharmacokinetic properties, poor selectivity, and insufficient efficacy in comparison to placebo controls. To address these challenges, we employed a virtual screening workflow integrating ligand-based and structure-based approaches to identify novel P2X7 allosteric antagonists. A 3D pharmacophore model derived from three known P2X7 antagonists (A740003, A804598, and JNJ47965567) was used to filter four libraries of commercially available compounds (approximately 10,000,000 total). These compounds were docked into a human P2X7 homology model and ranked by four distinct scoring functions. Eleven compounds were selected based on drug-like properties and key interactions with residues lining the target pocket. Among those, six compounds inhibited P2X7 activation in a YO-PRO 1 dye uptake assay (30 μM), while just two of those (2 and 9) were also active in a Membrane Potential Red assay (10 μM). Further screening of 10 analogues of 2 and 9 led to the identification of 2g, which displayed comparable potency (IC50 = 1.31 μM) to 2 (IC50 = 1.88 μM) in the YO-PRO 1 dye uptake assay. Docking studies of 2g within the negative allosteric pocket provided insights into its binding mode and key interacting residues. These findings offer a promising starting point for the development of optimized P2X7 antagonists.

• MeSH
• antagonisté purinergních receptorů P2X * chemie   farmakologie   metabolismus   MeSH
• lidé MeSH
• ligandy MeSH
• preklinické hodnocení léčiv metody   MeSH
• purinergní receptory P2X7 * metabolismus   chemie   MeSH
• simulace molekulového dockingu MeSH
• uživatelské rozhraní počítače * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antagonisté purinergních receptorů P2X * MeSH
• ligandy MeSH
• purinergní receptory P2X7 * MeSH

P2X receptors (P2XRs) are ATP-gated cationic channels that are allosterically modulated by numerous compounds, including steroids and neurosteroids. These compounds may both inhibit and potentiate the activity of P2XRs, but sex steroids such as 17β-estradiol or progesterone are reported to be inactive. Here, we tested a hypothesis that testosterone, another sex hormone, modulates activity of P2XRs. We examined actions of native testosterone and a series of testosterone derivatives on the gating of recombinant P2X2R, P2X4R and P2X7R and native channels expressed in pituitary cells and hypothalamic neurons. The 17β-ester derivatives of testosterone rapidly and positively modulate the 1 µM ATP-evoked currents in P2X2R- and P2X4R-expressing cells, but not agonist-evoked currents in P2X7R-expressing cells. In general, most of the tested testosterone derivatives are more potent modulators than endogenous testosterone. The comparison of chemical structures and whole-cell recordings revealed that their interactions with P2XRs depend on the lipophilicity and length of the alkyl chain at position C-17. Pre-treatment with testosterone butyrate or valerate increases the sensitivity of P2X2R and P2X4R to ATP by several fold, reduces the rate of P2X4R desensitization, accelerates resensitization, and enhances ethidium uptake by P2X4R. Native channels are also potentiated by testosterone derivatives, while endogenously expressed GABA receptors type A are inhibited. The effect of ivermectin, a P2X4R-specific allosteric modulator, on deactivation is antagonized by testosterone derivatives in a concentration-dependent manner. Together, our results provide evidence for potentiation of particular subtypes of P2XRs by testosterone derivatives and suggest a potential role of ivermectin binding site for steroid-induced modulation. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

• Klíčová slova
• ATP, P2XR, allosteric modulation, ivermectin, purinergic receptors, testosterone derivatives,
• MeSH
• gating iontového kanálu účinky léků   fyziologie   MeSH
• HEK293 buňky MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• potkani Wistar MeSH
• purinergní receptory P2X2 metabolismus   MeSH
• purinergní receptory P2X4 metabolismus   MeSH
• testosteron farmakologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• purinergní receptory P2X2 MeSH
• purinergní receptory P2X4 MeSH
• testosteron MeSH