**4. Potentiation of hormone secretion in pituitary gland**

Depolarization and Ca2+ influx stimulated by extracellular ATP has numerous functions also in endocrine cells. These involve stimulation of luteinizing hormone, prolactin, oxytocin and vasopressin hormone secretion by pituitary gland (Kapoor and Sladek, 2000; Stojilkovic, 2009; Stojilkovic et al., 2010b). The anterior pituitary is a heterogeneous gland with multiple cell types that secrete six major peptide hormones necessary for reproduction, lactation, growth, development, metabolic homeostasis, and the response to stress: FSH and LHproducing gonadotrophs, prolactin (PRL)-producing lactotrophs, GH-producing somatotrophs, TSH-producing thyrotrophs, and ACTH-producing corticotrophs. This lobe also contains the non-hormone-producing folliculostellate cells, which are glia-like cells, and endothelial cells that line the capillaries. Pituitary hormone secretion is under control of hypothalamic neurohormones and is also modulated by extracellular ATP (Chen et al., 1995) that is released by the anterior pituitary itself in a regulated manner, or coreleased with hypothalamic peptides (Tomic et al., 1996; Lazarowski et al., 2000; Stojilkovic and Koshimizu, 2001; He et al., 2005). The mRNA transcripts for several P2X subunits (P2X2R, P2X3R, P2X4R, and P2X7R) were identified in anterior pituitary cells from neonatal (Zemkova *et al.*, 2006) and adult rats (Koshimizu et al., 2000a; Koshimizu et al., 2000b; Stojilkovic and Koshimizu, 2001), including two spliced forms of the P2X2R subunit. Experiments with the plasma membrane-targeted luciferase expressed in HEK cells or ACN neuroblastoma cells indicated that endogenous extracellular ATP concentrations are in the range of 100-200 µM, which is more than sufficient to activate all types of P2X receptors (Pellegatti et al., 2005). *In vivo*, the ATP action on gonadotroph functions could be controlled by ectonucleotidases 1-3, which are expressed in pituitary cells (He et al., 2005) and provide an effective pathway for the control of extracellular ATP concentrations.

Detection and localization of mRNA transcripts for P2X2, P2X3 and P2X4 receptors in pituitary gland by in situ hybridization. *PP,* posterior pituitary, *AP,* anterior pituitary. For details see: (Stojilkovic et al., 2010b).

Most anterior pituitary cells express functional P2X receptors (Fig.4) (Carew et al., 1994; Villalobos et al., 1997; Koshimizu et al., 2000a). The P2X2Rs were identified in pituitary gonadotrophs (Tomic et al., 1996; Koshimizu et al., 2000b; Zemkova et al., 2006) and somatotrophs (Koshimizu et al., 2000a). Lactotrophs express functional P2X4Rs (Carew et al., 1994; He et al., 2003; Zemkova et al., 2010) as well as P2X7R subtypes (Chung et al., 2000; He et al., 2003). Corticotrophs seem not to respond to extracellular ATP directly by stimulation of any P2X receptor (Zhao et al., 2006), and identification of P2X receptors in remaining anterior pituitary cell type, thyrotrophs, has not yet been done. Functional studies showed that ATP application induces inward slowly desensitizig current and increases

Facilitation of Neurotransmitter and Hormone Release by P2X Purinergic Receptors 73

Fig. 5. Electrophysiological characterization of the P2X2R in pituitary gonadotrophs. A, Gonadotrophs can be identified in a mixed population of pituitary cells because of their specific GnRH-induced calcium oscillations monitored as outward calcium-activated potassium current (IK-Ca). B, Stimulation of electrical activity by application of ATP to identified gonadotrophs. C, The sensitivity of ATP-induced current to suramine, general P2XR blocker. D, Insensitivity of ATP-induced current to ivermectin indicates that gonadotrophs express the P2X2 receptor. For details see: (Zemkova et al., 2006).

To summarize, the cellular actions of extracellular ATP and P2XRs in the brain range from modulation of resting membrane potential and electrical activity to stimulation of neurotransmitter and hormone release, including release of anterior pituitary hormones. It is clear that the effects of ATP and its analogs are realized by membrane depolarization resulting in voltage-dependent Ca2+ entry and by Ca2+ entry through the pore of P2XR itself. Nonetheless, the molecular and neurochemical mechanisms underlying these effects are far from being fully understood and likely involve multiple receptor systems and various signaling pathways. Experimental evidences gathered so far suggest that whereas neurotransmitter release seems to be linked to the activation of presynaptic P2X1, P2X2, P2X3 and perhaps P2X4 and P2X7 receptors expressed in nerve terminals, neuropeptide and hormone secretion more likely involves P2X2 and P2X4 receptors on the surface of neuronal somata and pituitary cells. Thus, extracellular ATP together with P2XRs comprise a new excitatory system in the mammalian central nervous system and an increasing number of studies support also an indirect role of P2XRs in inhibitory system owing to its ability to facilitate GABA release. However, surprisingly few studies confirmed the role of ATP as neurotransmitter in the central nervous system, whereas its modulatory roles are well established. Further studies aimed at analyzing the cellular and molecular actions of ATP in various brain regions and under different physiological states would be required for a

**5. Conclusion** 

comprehensive understanding.

Fig. 4. Expression of P2XR in pituitary gland.

frequency of action potentials (Fig.5) in pituitary gonadotrophs. These responses were sensitive to PPADS indicating that P2X2Rs could operate as depolarizing channels in the pituitary (Zemkova et al., 2006). ATP could play the role of modulator in the anterior pituitary lactotrophs (Stojilkovic and Koshimizu, 2001), P2X4R potentiator ivermectin per se augmented ATP-induced prolactin secretion and slightly potentiated the effect of TRH in lactrotrophs which express P2X4Rs (Zemkova et al., 2006). Thus in pituitary, ATP may serve to synchronize of spontaneous electrical activity, to initiate intercellular Ca2+ waves, as observed in other cell types (Guthrie et al., 1999) and modulate G-protein coupled receptorstimulated Ca2+ signaling by refilling of intracellular stores from calcium influx through P2XR pore (Zemkova et al., 2006). Such an action of extracellular ATP could provide a mechanism for the amplification of the effects of hypothalamic peptides on pituitary hormone secretion. Further experiments should clarify to what extent these capacities of P2X receptors are utilized *in vivo*.

Fig. 5. Electrophysiological characterization of the P2X2R in pituitary gonadotrophs. A, Gonadotrophs can be identified in a mixed population of pituitary cells because of their specific GnRH-induced calcium oscillations monitored as outward calcium-activated potassium current (IK-Ca). B, Stimulation of electrical activity by application of ATP to identified gonadotrophs. C, The sensitivity of ATP-induced current to suramine, general P2XR blocker. D, Insensitivity of ATP-induced current to ivermectin indicates that gonadotrophs express the P2X2 receptor. For details see: (Zemkova et al., 2006).

#### **5. Conclusion**

72 Neuroscience – Dealing with Frontiers

frequency of action potentials (Fig.5) in pituitary gonadotrophs. These responses were sensitive to PPADS indicating that P2X2Rs could operate as depolarizing channels in the pituitary (Zemkova et al., 2006). ATP could play the role of modulator in the anterior pituitary lactotrophs (Stojilkovic and Koshimizu, 2001), P2X4R potentiator ivermectin per se augmented ATP-induced prolactin secretion and slightly potentiated the effect of TRH in lactrotrophs which express P2X4Rs (Zemkova et al., 2006). Thus in pituitary, ATP may serve to synchronize of spontaneous electrical activity, to initiate intercellular Ca2+ waves, as observed in other cell types (Guthrie et al., 1999) and modulate G-protein coupled receptorstimulated Ca2+ signaling by refilling of intracellular stores from calcium influx through P2XR pore (Zemkova et al., 2006). Such an action of extracellular ATP could provide a mechanism for the amplification of the effects of hypothalamic peptides on pituitary hormone secretion. Further experiments should clarify to what extent these capacities of P2X

Fig. 4. Expression of P2XR in pituitary gland.

receptors are utilized *in vivo*.

To summarize, the cellular actions of extracellular ATP and P2XRs in the brain range from modulation of resting membrane potential and electrical activity to stimulation of neurotransmitter and hormone release, including release of anterior pituitary hormones. It is clear that the effects of ATP and its analogs are realized by membrane depolarization resulting in voltage-dependent Ca2+ entry and by Ca2+ entry through the pore of P2XR itself. Nonetheless, the molecular and neurochemical mechanisms underlying these effects are far from being fully understood and likely involve multiple receptor systems and various signaling pathways. Experimental evidences gathered so far suggest that whereas neurotransmitter release seems to be linked to the activation of presynaptic P2X1, P2X2, P2X3 and perhaps P2X4 and P2X7 receptors expressed in nerve terminals, neuropeptide and hormone secretion more likely involves P2X2 and P2X4 receptors on the surface of neuronal somata and pituitary cells. Thus, extracellular ATP together with P2XRs comprise a new excitatory system in the mammalian central nervous system and an increasing number of studies support also an indirect role of P2XRs in inhibitory system owing to its ability to facilitate GABA release. However, surprisingly few studies confirmed the role of ATP as neurotransmitter in the central nervous system, whereas its modulatory roles are well established. Further studies aimed at analyzing the cellular and molecular actions of ATP in various brain regions and under different physiological states would be required for a comprehensive understanding.

Facilitation of Neurotransmitter and Hormone Release by P2X Purinergic Receptors 75

Donato R, Rodrigues RJ, Takahashi M, Tsai MC, Soto D, Miyagi K, Villafuertes RG, Cunha

Duan S, Anderson CM, Keung EC, Chen Y, Chen Y, Swanson RA (2003) P2X7 receptormediated release of excitatory amino acids from astrocytes. J Neurosci 23:1320-1328. Duckwitz W, Hausmann R, Aschrafi A, Schmalzing G (2006) P2X5 subunit assembly

Dunn PM, Zhong Y, Burnstock G (2001) P2X receptors in peripheral neurons. Prog

Edwards FA, Gibb AJ, Colquhoun D (1992) ATP receptor-mediated synaptic currents in the

Egan TM, Khakh BS (2004) Contribution of calcium ions to P2X channel responses. J

Egan TM, Haines WR, Voigt MM (1998) A domain contributing to the ion channel of ATP-

Egan TM, Cox JA, Voigt MM (2004) Molecular structure of P2X receptors. Curr Top Med

Ennion SJ, Evans RJ (2002) Conserved cysteine residues in the extracellular loop of the

Evans RJ, Derkach V, Surprenant A (1992) ATP mediates fast synaptic transmission in

Ferrari D, Chiozzi P, Falzoni S, Hanau S, Di Virgilio F (1997a) Purinergic modulation of

Ferrari D, Chiozzi P, Falzoni S, Dal Susino M, Melchiorri L, Baricordi OR, Di Virgilio F

Ferrari D, Pizzirani C, Adinolfi E, Lemoli RM, Curti A, Idzko M, Panther E, Di Virgilio F

Fields RD, Stevens B (2000) ATP: an extracellular signaling molecule between neurons and

Fields RD, Burnstock G (2006) Purinergic signalling in neuron-glia interactions. Nat Rev

Fountain SJ, Parkinson K, Young MT, Cao L, Thompson CR, North RA (2007) An

Gomes DA, Song Z, Stevens W, Sladek CD (2009) Sustained stimulation of vasopressin and

Gomez-Villafuertes R, Gualix J, Miras-Portugal MT (2001) Single GABAergic synaptic

trafficking to the cell surface. Mol Pharmacol 61:303-311.

receptor of human macrophages. J Immunol 159:1451-1458.

modulating Ca2+ influx. Cell Calcium 44:521-532.

aspartate. J Biol Chem 281:39561-39572.

central nervous system. Nature 359:144-147.

mammalian neurons. Nature 357:503-505.

Neurobiol 65:107-134.

Neurosci 24:3413-3420.

Neurosci 18:2350-2359.

J Exp Med 185:579-582.

glia. Trends Neurosci 23:625-633.

discoideum. Nature 448:200-203.

Comp Physiol 297:R940-949.

to induce GABA secretion. J Neurochem 77:84-93.

176:3877-3883.

Neurosci 7:423-436.

Chem 4:821-829.

RA, Edwards FA (2008) GABA release by basket cells onto Purkinje cells, in rat cerebellar slices, is directly controlled by presynaptic purinergic receptors,

requires scaffolding by the second transmembrane domain and a conserved

gated P2X2 receptors identified by the substituted cysteine accessibility method. J

human P2X(1) receptor form disulfide bonds and are involved in receptor

interleukin-1 beta release from microglial cells stimulated with bacterial endotoxin.

(1997b) Extracellular ATP triggers IL-1 beta release by activating the purinergic P2Z

(2006) The P2X7 receptor: a key player in IL-1 processing and release. J Immunol

intracellular P2X receptor required for osmoregulation in Dictyostelium

oxytocin release by ATP and phenylephrine requires recruitment of desensitization-resistant P2X purinergic receptors. Am J Physiol Regul Integr

terminals from rat midbrain exhibit functional P2X and dinucleotide receptors, able

#### **6. Acknowledgements**

This study was supported by the Internal Grant Agency of Academy of Sciences (Grant No. IAA500110910), the Grant Agency of the Czech Republic (305/07/0681, P304/12/P371, the Academy of Sciences of the Czech Republic (Research Project No. AVOZ 50110509) and the Centrum for Neuroscience (Research Project No. LC554).

#### **7. References**


This study was supported by the Internal Grant Agency of Academy of Sciences (Grant No. IAA500110910), the Grant Agency of the Czech Republic (305/07/0681, P304/12/P371, the Academy of Sciences of the Czech Republic (Research Project No. AVOZ 50110509) and the

Abbracchio MP, Burnstock G, Verkhratsky A, Zimmermann H (2009) Purinergic signalling

Adinolfi E, Cirillo M, Woltersdorf R, Falzoni S, Chiozzi P, Pellegatti P, Callegari MG,

Barrera NP, Ormond SJ, Henderson RM, Murrell-Lagnado RD, Edwardson JM (2005)

Buell G, Lewis C, Collo G, North RA, Surprenant A (1996) An antagonist-insensitive P2X

Burkhart CN (2000) Ivermectin: an assessment of its pharmacology, microbiology and

Burnstock G (2011) Introductory overview of purinergic signalling. Front Biosci (Elite Ed)

Calvert JA, Evans RJ (2004) Heterogeneity of P2X receptors in sympathetic neurons:

Carew MA, Wu ML, Law GJ, Tseng YZ, Mason WT (1994) Extracellular ATP activates

Clyne JD, Wang LF, Hume RI (2002) Mutational analysis of the conserved cysteines of the

Coddou C, Stojilkovic SS, Huidobro-Toro JP (2011) Allosteric modulation of ATP-gated P2X

Collo G, Neidhart S, Kawashima E, Kosco-Vilbois M, North RA, Buell G (1997) Tissue distribution of the P2X7 receptor. Neuropharmacology 36:1277-1283. Collo G, North RA, Kawashima E, Merlo-Pich E, Neidhart S, Surprenant A, Buell G (1996)

Di Virgilio F, Chiozzi P, Falzoni S, Ferrari D, Sanz JM, Venketaraman V, Baricordi OR (1998)

extended family of ATP-gated ion channels. J Neurosci 16:2495-2507. Day TA, Sibbald JR, Khanna S (1993) ATP mediates an excitatory noradrenergic neuron

input to supraoptic vasopressin cells. Brain Res 607:341-344.

Cytolytic P2X purinoceptors. Cell Death Differ 5:191-199.

contribution of neuronal P2X1 receptors revealed using knockout mice. Mol

calcium entry and mobilization via P2U-purinoceptors in rat lactotrophs. Cell

Cloning OF P2X5 and P2X6 receptors and the distribution and properties of an

Sandona D, Markwardt F, Schmalzing G, Di Virgilio F (2010) Trophic activity of a naturally occurring truncated isoform of the P2X7 receptor. Faseb J 24:3393-3404. Ballerini P, Rathbone MP, Di Iorio P, Renzetti A, Giuliani P, D'Alimonte I, Trubiani O,

Caciagli F, Ciccarelli R (1996) Rat astroglial P2Z (P2X7) receptors regulate

Atomic force microscopy imaging demonstrates that P2X2 receptors are trimers but that P2X6 receptor subunits do not oligomerize. J Biol Chem 280:10759-10765. Bavan S, Straub VA, Blaxter ML, Ennion SJ (2009) A P2X receptor from the tardigrade

species Hypsibius dujardini with fast kinetics and sensitivity to zinc and copper.

in the nervous system: an overview. Trends Neurosci 32:19-29.

intracellular calcium and purine release. Neuroreport 7:2533-2537.

receptor expressed in epithelia and brain. Embo J 15:55-62.

Burnstock G (2004) Cotransmission. Curr Opin Pharmacol 4:47-52.

rat P2X2 purinoceptor. J Neurosci 22:3873-3880.

receptor channels. Rev Neurosci 22:335-354.

**6. Acknowledgements** 

BMC Evol Biol 9:17.

Pharmacol 65:139-148.

Calcium 16:227-235.

3:896-900.

safety. Vet Hum Toxicol 42:30-35.

**7. References** 

Centrum for Neuroscience (Research Project No. LC554).


Facilitation of Neurotransmitter and Hormone Release by P2X Purinergic Receptors 77

Ireland MF, Noakes PG, Bellingham MC (2004) P2X7-like receptor subunits enhance

Jacobson KA, Jarvis MF, Williams M (2002) Purine and pyrimidine (P2) receptors as drug

Jelinkova I, Yan Z, Liang Z, Moonat S, Teisinger J, Stojilkovic SS, Zemkova H (2006)

channel gating and interaction with ivermectin. Pflugers Arch 456:939-950. Jiang LH, Rassendren F, Spelta V, Surprenant A, North RA (2001) Amino acid residues

Jiang LH, Kim M, Spelta V, Bo X, Surprenant A, North RA (2003) Subunit arrangement in

Jin YH, Bailey TW, Li BY, Schild JH, Andresen MC (2004) Purinergic and vanilloid receptor

Jindrichova M, Vavra V, Obsil T, Stojilkovic SS, Zemkova H (2009) Functional relevance of

Jindrichova M, Khafizov K, Skorinkin A, Fayuk D, Bart G, Zemkova H, Giniatullin R (2011)

Jo YH, Role LW (2002) Coordinate release of ATP and GABA at in vitro synapses of lateral

Jo YH, Donier E, Martinez A, Garret M, Toulme E, Boue-Grabot E (2011) Crosstalk between

Kapoor JR, Sladek CD (2000) Purinergic and adrenergic agonists synergize in stimulating

Kato F, Shigetomi E (2001) Distinct modulation of evoked and spontaneous EPSCs by purinoceptors in the nucleus tractus solitarii of the rat. J Physiol 530:469-486. Kawate T, Michel JC, Birdsong WT, Gouaux E (2009) Crystal structure of the ATP-gated

Kawate T, Robertson JL, Li M, Silberberg SD, Swartz KJ (2011) Ion access pathway to the transmembrane pore in P2X receptor channels. J Gen Physiol 137:579-590. Khakh BS, Henderson G (1998) ATP receptor-mediated enhancement of fast excitatory

Khakh BS, Egan TM (2005) Contribution of transmembrane regions to ATP-gated P2X2

Khakh BS, North RA (2006) P2X receptors as cell-surface ATP sensors in health and disease.

Khakh BS, Gittermann D, Cockayne DA, Jones A (2003) ATP modulation of excitatory

Permeability of ATP-Gated P2X3 Receptor. J Neurochem 119:676-685. Jo YH, Schlichter R (1999) Synaptic corelease of ATP and GABA in cultured spinal neurons.

Neuroscience 128:269-280.

targets. J Med Chem 45:4057-4093.

P2X(2) receptor. J Biol Chem 276:14902-14908.

P2X receptors. J Neurosci 23:8903-8910.

tractus solitarius. J Neurosci 24:4709-4717.

hypothalamic neurons. J Neurosci 22:4794-4804.

vasopressin and oxytocin release. J Neurosci 20:8868-8875.

P2X(4) ion channel in the closed state. Nature 460:592-598.

neurotransmitter release in the brain. Mol Pharmacol 54:372-378.

channel permeability dynamics. J Biol Chem 280:6118-6129.

synapses onto interneurons. J Neurosci 23:7426-7437.

Neurochem 109:923-934.

Nat Neurosci 2:241-245.

Biol Chem 256:19993-20004.

Nature 442:527-532.

excitatory synaptic transmission at central synapses by presynaptic mechanisms.

Identification of P2X(4) receptor-specific residues contributing to the ivermectin effects on channel deactivation. Biochem Biophys Res Commun 349:619-625. Jelinkova I, Vavra V, Jindrichova M, Obsil T, Zemkova HW, Zemkova H, Stojilkovic SS

(2008) Identification of P2X(4) receptor transmembrane residues contributing to

involved in gating identified in the first membrane-spanning domain of the rat

activation releases glutamate from separate cranial afferent terminals in nucleus

aromatic residues in the first transmembrane domain of P2X receptors. J

Highly Conserved Tyrosine 37 Stabilizes Desensitized States and Restricts Calcium

P2X4 and GABA-A receptors determines synaptic efficacy at central synapses. J


Gordon GR, Iremonger KJ, Kantevari S, Ellis-Davies GC, MacVicar BA, Bains JS (2009)

Gourine AV, Melenchuk EV, Poputnikov DM, Gourine VN, Spyer KM (2002) Involvement

Gu JG, MacDermott AB (1997) Activation of ATP P2X receptors elicits glutamate release

Guo W, Sun J, Xu X, Bunstock G, He C, Xiang Z (2009) P2X receptors are differentially

paraventricular nuclei of rat hypothalamus. Histochem Cell Biol 131:29-41. Guthrie PB, Knappenberger J, Segal M, Bennett MV, Charles AC, Kater SB (1999) ATP released from astrocytes mediates glial calcium waves. J Neurosci 19:520-528. Haines WR, Voigt MM, Migita K, Torres GE, Egan TM (2001a) On the contribution of the

Haines WR, Migita K, Cox JA, Egan TM, Voigt MM (2001b) The first transmembrane

He ML, Gonzalez-Iglesias AE, Stojilkovic SS (2003) Role of nucleotide P2 receptors in

He ML, A.E. GI, M. T, Stojilkovic SS (2005) Release and extracellular metabolism of ATP by

Heine C, Heimrich B, Vogt J, Wegner A, Illes P, Franke H (2006) P2 receptor-stimulation

Hervas C, Perez-Sen R, Miras-Portugal MT (2003) Coexpression of functional P2X and P2Y nucleotide receptors in single cerebellar granule cells. J Neurosci Res 73:384-399. Hiruma H, Bourque CW (1995) P2 purinoceptor-mediated depolarization of rat supraoptic

Hugel S, Schlichter R (2000) Presynaptic P2X receptors facilitate inhibitory GABAergic

Chen ZP, Levy A, Lightman SL (1994) Activation of specific ATP receptors induces a rapid

Chen ZP, Kratzmeier M, Levy A, McArdle CA, Poch A, Day A, Mukhopadhyay AK,

Chung HS, Park KS, Cha SK, Kong ID, Lee JW (2000) ATP-induced [Ca(2+)](i) changes and

Illes P, Ribeiro JA (2004) Neuronal P2 receptors of the central nervous system. Curr Top

Inoue K, Koizumi S, Tsuda M (2007) The role of nucleotides in the neuron--glia communication responsible for the brain functions. J Neurochem 102:1447-1458.

of pituitary function. Proc Natl Acad Sci U S A 92:5219-5223.

depolarization in GH3 cells. Br J Pharmacol 130:1843-1852.

Neuron 64:391-403.

278:46270-46277.

138:303-311.

20:2121-2130.

641:249-256.

Med Chem 4:831-838.

Signalling 1:135-144.

rats. Br J Pharmacol 135:2047-2055.

from sensory neuron synapses. Nature 389:749-753.

ionotropic P2X receptor. J Neurosci 21:5885-5892.

neurosecretory cells in vitro. J Physiol 489:805-811.

the channel. J Biol Chem 276:32793-32798.

Astrocyte-mediated distributed plasticity at hypothalamic glutamate synapses.

of purinergic signalling in central mechanisms of body temperature regulation in

expressed on vasopressin- and oxytocin-containing neurons in the supraoptic and

first transmembrane domain to whole-cell current through an ATP-gated

domain of the P2X receptor subunit participates in the agonist-induced gating of

calcium signaling and prolactin release in pituitary lactotrophs. J Biol Chem

ecto-nucleotidase eNTPDase 1-3 in hypothalamic and pituitary cells. Purinergic

influences axonal outgrowth in the developing hippocampus in vitro. Neuroscience

transmission between cultured rat spinal cord dorsal horn neurons. J Neurosci

increase in intracellular calcium ions in rat hypothalamic neurons. Brain Res

Lightman SL (1995) Evidence for a role of pituitary ATP receptors in the regulation


Facilitation of Neurotransmitter and Hormone Release by P2X Purinergic Receptors 79

Mackenzie AB, Young MT, Adinolfi E, Surprenant A (2005) Pseudoapoptosis induced by brief activation of ATP-gated P2X7 receptors. J Biol Chem 280:33968-33976. Matute C, Torre I, Perez-Cerda F, Perez-Samartin A, Alberdi E, Etxebarria E, Arranz AM,

experimental autoimmune encephalomyelitis. J Neurosci 27:9525-9533. Mehta VB, Hart J, Wewers MD (2001) ATP-stimulated release of interleukin (IL)-1beta and

Migita K, Haines WR, Voigt MM, Egan TM (2001) Polar residues of the second

Michel AD, Chambers LJ, Clay WC, Condreay JP, Walter DS, Chessell IP (2007) Direct

Mio K, Kubo Y, Ogura T, Yamamoto T, Sato C (2005) Visualization of the trimeric P2X2

Nakatsuka T, Gu JG (2001) ATP P2X receptor-mediated enhancement of glutamate release and evoked EPSCs in dorsal horn neurons of the rat spinal cord. J Neurosci 21:6522-6531. Nakatsuka T, Tsuzuki K, Ling JX, Sonobe H, Gu JG (2003) Distinct roles of P2X receptors in

Nelson DW, Gregg RJ, Kort ME, Perez-Medrano A, Voight EA, Wang Y, Grayson G, Namovic

Nicke A, Kerschensteiner D, Soto F (2005) Biochemical and functional evidence for heteromeric assembly of P2X1 and P2X4 subunits. J Neurochem 92:925-933. Nicke A, Baumert HG, Rettinger J, Eichele A, Lambrecht G, Mutschler E, Schmalzing G

North R (1996) P2X receptors: a third major class of ligand-gated ion channels. Ciba Found

Ormond SJ, Barrera NP, Qureshi OS, Henderson RM, Edwardson JM, Murrell-Lagnado

Pankratov Y, Castro E, Miras-Portugal MT, Krishtal O (1998) A purinergic component of the

North RA (2002) Molecular physiology of P2X receptors. Physiol Rev 82:1013-1067. North RA (2004) P2X3 receptors and peripheral pain mechanisms. J Physiol 554:301-308. Novakovic SD, Kassotakis LC, Oglesby IB, Smith JA, Eglen RM, Ford AP, Hunter JC (1999)

naive rats and following neuropathic injury. Pain 80:273-282.

benzyl-5-phenyltetrazole P2X7 antagonists. J Med Chem 49:3659-3666. Newman EA (2003) Glial cell inhibition of neurons by release of ATP. J Neurosci 23:1659-

cleavage. J Biol Chem 276:3820-3826.

receptor. J Biol Chem 276:30934-30941.

Commun 337:998-1005.

1666.

Symp 198:91-105.

69:1692-1700.

cord. J Neurophysiol 89:3243-3252.

cooperativity of binding. Br J Pharmacol 151:103-114.

ligand-gated ion channels. Embo J 17:3016-3028.

the rat hippocampus. Eur J Neurosci 10:3898-3902.

Ravid R, Rodriguez-Antiguedad A, Sanchez-Gomez M, Domercq M (2007) P2X(7) receptor blockade prevents ATP excitotoxicity in oligodendrocytes and ameliorates

IL-18 requires priming by lipopolysaccharide and is independent of caspase-1

transmembrane domain influence cation permeability of the ATP-gated P2X(2)

labelling of the human P2X7 receptor and identification of positive and negative

receptor with a crown-capped extracellular domain. Biochem Biophys Res

modulating glutamate release at different primary sensory synapses in rat spinal

MT, Donnelly-Roberts DL, Niforatos W, Honore P, Jarvis MF, Faltynek CR, Carroll WA (2006) Structure-activity relationship studies on a series of novel, substituted 1-

(1998) P2X1 and P2X3 receptors form stable trimers: a novel structural motif of

Immunocytochemical localization of P2X3 purinoceptors in sensory neurons in

RD (2006) An uncharged region within the N terminus of the P2X6 receptor inhibits its assembly and exit from the endoplasmic reticulum. Mol Pharmacol

excitatory postsynaptic current mediated by P2X receptors in the CA1 neurons of


Khakh BS, Proctor WR, Dunwiddie TV, Labarca C, Lester HA (1999) Allosteric control of gating and kinetics at P2X(4) receptor channels. J Neurosci 19:7289-7299. King BF, Townsend-Nicholson A, Wildman SS, Thomas T, Spyer KM, Burnstock G (2000)

Knott TK, Velazquez-Marrero C, Lemos JR (2005) ATP elicits inward currents in isolated

Kodama N, Funahashi M, Mitoh Y, Minagi S, Matsuo R (2007) Purinergic modulation of area postrema neuronal excitability in rat brain slices. Brain Res 1165:50-59. Koshimizu TA, Tomic M, Wong AO, Zivadinovic D, Stojilkovic SS (2000a) Characterization

Koshimizu TA, Van Goor F, Tomic M, Wong AO, Tanoue A, Tsujimoto G, Stojilkovic SS

Kracun S, Chaptal V, Abramson J, Khakh BS (2010) Gated access to the pore of a P2X

Labasi JM, Petrushova N, Donovan C, McCurdy S, Lira P, Payette MM, Brissette W, Wicks

Lazarowski ER, Boucher RC, Harden TK (2000) Constitutive release of ATP and evidence for

Le Feuvre RA, Brough D, Iwakura Y, Takeda K, Rothwell NJ (2002) Priming of macrophages

Le KT, Babinski K, Seguela P (1998) Central P2X4 and P2X6 channel subunits coassemble

Lewis C, Neidhart S, Holy C, North RA, Buell G, Surprenant A (1995) Coexpression of P2X2

Li M, Kawate T, Silberberg SD, Swartz KJ (2011) Pore-opening mechanism in trimeric P2X

Li P, Calejesan AA, Zhuo M (1998) ATP P2x receptors and sensory synaptic transmission

Li Z, Migita K, Samways DS, Voigt MM, Egan TM (2004) Gain and loss of channel function

Loesch A, Burnstock G (2001) Immunoreactivity to P2X(6) receptors in the rat hypothalamo-

into a novel heteromeric ATP receptor. J Neurosci 18:7152-7159.

expressed in excitable cells. Mol Pharmacol 58:936-945.

20:4871-4877.

10121.

31068.

277:3210-3218.

Pflugers Arch 450:381-389.

Endocrinology 141:4091-4099.

Neurosci Lett 421:158-162.

neurons. Nature 377:432-435.

Neurophysiol 80:3356-3360.

receptor. J Neurosci 24:7378-7386.

gold-silver labelling. Neuroscience 106:621-631.

receptor channels. Nat Commun 1:44.

Coexpression of rat P2X2 and P2X6 subunits in Xenopus oocytes. J Neurosci

vasopressinergic neurohypophysial terminals via P2X2 and P2X3 receptors.

of purinergic receptors and receptor-channels expressed in anterior pituitary cells.

(2000b) Characterization of calcium signaling by purinergic receptor-channels

receptor: Structural implications for closed-open transitions. J Biol Chem 285:10110-

JR, Audoly L, Gabel CA (2002) Absence of the P2X7 receptor alters leukocyte function and attenuates an inflammatory response. J Immunol 168:6436-6445. Lalo U, Verkhratsky A, Pankratov Y (2007) Ivermectin potentiates ATP-induced ion currents

in cortical neurones: evidence for functional expression of P2X4 receptors?

major contribution of ecto-nucleotide pyrophosphatase and nucleoside diphosphokinase to extracellular nucleotide concentrations. J Biol Chem 275:31061-

with lipopolysaccharide potentiates P2X7-mediated cell death via a caspase-1 dependent mechanism, independently of cytokine production. J Biol Chem

and P2X3 receptor subunits can account for ATP-gated currents in sensory

between primary afferent fibers and spinal dorsal horn neurons in rats. J

by alanine substitutions in the transmembrane segments of the rat ATP-gated P2X2

neurohypophysial system: an ultrastructural study with extravidin and colloidal


Facilitation of Neurotransmitter and Hormone Release by P2X Purinergic Receptors 81

Solle M, Labasi J, Perregaux DG, Stam E, Petrushova N, Koller BH, Griffiths RJ, Gabel CA

Song Z, Vijayaraghavan S, Sladek CD (2007) ATP increases intracellular calcium in

Sperlagh B, Heinrich A, Csolle C (2007) P2 receptor-mediated modulation of

Sperlagh B, Kofalvi A, Deuchars J, Atkinson L, Milligan CJ, Buckley NJ, Vizi ES (2002)

Stojilkovic SS (2009) Purinergic regulation of hypothalamopituitary functions. Trends

Stojilkovic SS, Koshimizu T (2001) Signaling by extracellular nucleotides in anterior pituitary

Stojilkovic SS, Yan Z, Obsil T, Zemkova H (2010a) Structural Insights into the Function of

Stojilkovic SS, He ML, Koshimizu TA, Balik A, Zemkova H (2010b) Signaling by purinergic receptors and channels in the pituitary gland. Mol Cell Endocrinol 314:184-191. Stoop R, Thomas S, Rassendren F, Kawashima E, Buell G, Surprenant A, North RA (1999)

based on methanethiosulfonate block at T336C. Mol Pharmacol 56:973-981. Surprenant A, North RA (2009) Signaling at purinergic P2X receptors. Annu Rev Physiol

Surprenant A, Rassendren F, Kawashima E, North RA, Buell G (1996) The cytolytic P2Z

Terasawa E, Keen KL, Grendell RL, Golos TG (2005) Possible role of 5'-adenosine

hormone-releasing hormone neurons. Mol Endocrinol 19:2736-2747. Tomic M, Jobin RM, Vergara LA, Stojilkovic SS (1996) Expression of purinergic receptor

reticulum-derived calcium oscillations. J Biol Chem 271:21200-21208. Torres GE, Egan TM, Voigt MM (1999) Identification of a domain involved in ATP-gated ionotropic receptor subunit assembly. J Biol Chem 274:22359-22365. Troadec JD, Thirion S, Nicaise G, Lemos JR, Dayanithi G (1998) ATP-evoked increases in

neurotransmitter release-an update. Purinergic Signal 3:269-284.

Physiol Regul Integr Comp Physiol 292:R423-431.

rat hippocampus. J Neurochem 81:1196-1211.

cells. Trends Endocrinol Metab 12:218-225.

P2X2 purinoceptor. J Physiol 511:89-103.

Endocrinol Metab 20:460-468.

276:125-132.

30:1251-1258.

71:333-359.

371:516-519.

Neuroscience 188:1-12.

738.

(2001) Altered cytokine production in mice lacking P2X(7) receptors. J Biol Chem

supraoptic neurons by activation of both P2X and P2Y purinergic receptors. Am J

Involvement of P2X7 receptors in the regulation of neurotransmitter release in the

P2X4: An ATP-Gated Cation Channel of Neuroendocrine Cells. Cell Mol Neurobiol

Contribution of individual subunits to the multimeric P2X(2) receptor: estimates

receptor for extracellular ATP identified as a P2X receptor (P2X7). Science 272:735-

triphosphate in synchronization of Ca2+ oscillations in primate luteinizing

channels and their role in calcium signaling and hormone release in pituitary gonadotrophs. Integration of P2 channels in plasma membrane- and endoplasmic

[Ca2+]i and peptide release from rat isolated neurohypophysial terminals via a

of ligand-gated ion channel defined by P2x receptor for extracellular ATP. Nature

P2X receptor activation in supraoptic neurons from freshly isolated rat brain slices.

Valera S, Hussy N, Evans RJ, Adami N, North RA, Surprenant A, Buell G (1994) A new class

Vavra V, Bhattacharya A, Zemkova H (2011) Facilitation of glutamate and GABA release by

Villalobos C, Alonso-Torre SR, Nunez L, Garcia-Sancho J (1997) Functional ATP receptors in

rat anterior pituitary cells. Am J Physiol 273:C1963-1971.


Papp L, Balazsa T, Kofalvi A, Erdelyi F, Szabo G, Vizi ES, Sperlagh B (2004) P2X receptor

Patti L, Raiteri L, Grilli M, Parodi M, Raiteri M, Marchi M (2006) P2X(7) receptors exert a

Pellegatti P, Falzoni S, Pinton P, Rizzuto R, Di Virgilio F (2005) A novel recombinant plasma

Raivich G (2005) Like cops on the beat: the active role of resting microglia. Trends Neurosci

Ralevic V, Burnstock G (1998) Receptors for purines and pyrimidines. Pharmacol Rev

Rassendren F, Buell G, Newbolt A, North RA, Surprenant A (1997) Identification of amino acid residues contributing to the pore of a P2X receptor. Embo J 16:3446-3454. Robertson SJ, Ennion SJ, Evans RJ, Edwards FA (2001) Synaptic P2X receptors. Curr Opin

Rodrigues RJ, Almeida T, Richardson PJ, Oliveira CR, Cunha RA (2005) Dual presynaptic

Rokic M, Tvrdonova V, Vavra V, Jindrichova M, Obsil T, Stojilkovic SS, Zemkova H (2010)

Samways DS, Egan TM (2007) Acidic amino acids impart enhanced Ca2+ permeability and

Samways DS, Migita K, Li Z, Egan TM (2008) On the role of the first transmembrane domain

Seguela P, Haghighi A, Soghomonian JJ, Cooper E (1996) A novel neuronal P2x ATP receptor ion channel with widespread distribution in the brain. J Neurosci 16:448-455. Shibuya I, Tanaka K, Hattori Y, Uezono Y, Harayama N, Noguchi J, Ueta Y, Izumi F,

Shigetomi E, Kato F (2004) Action potential-independent release of glutamate by Ca2+ entry

Silberberg SD, Chang TH, Swartz KJ (2005) Secondary Structure and Gating Rearrangements

Sim JA, Chaumont S, Jo J, Ulmann L, Young MT, Cho K, Buell G, North RA, Rassendren F

Sokolova E, Nistri A, Giniatullin R (2001) Negative cross talk between anionic GABAA and

Agonist Binding and Channel Gating. Physiol Res 59 59:927-935.

expressed in rat supraoptic neurones. J Physiol 514:351-367.

autonomic network. J Neurosci 24:3125-3135.

control by ATP of glutamate release via facilitatory P2X1, P2X2/3, and P2X3 and inhibitory P2Y1, P2Y2, and/or P2Y4 receptors in the rat hippocampus. J Neurosci

Roles of Conserved Ectodomain Cysteines of the Rat P2X4 Purinoreceptor in

flux in two members of the ATP-gated P2X receptor family. J Gen Physiol 129:245-

in cation permeability and flux of the ATP-gated P2X2 receptor. J Biol Chem

Yamashita H (1999) Evidence that multiple P2X purinoceptors are functionally

through presynaptic P2X receptors elicits postsynaptic firing in the brainstem

of Transmembrane Segments in Rat P2X4 Receptor Channels. J Gen Physiol

(2006) Altered hippocampal synaptic potentiation in P2X4 knock-out mice. J

cationic P2X ionotropic receptors of rat dorsal root ganglion neurons. J Neurosci

slices. J Pharmacol Exp Ther 310:973-980.

Neuropharmacology 50:705-713.

Cell 16:3659-3665.

Neurobiol 11:378-386.

28:571-573.

50:413-492.

25:6286-6295.

283:5110-5117.

125:347-359.

21:4958-4968.

Neurosci 26:9006-9009.

256.

activation elicits transporter-mediated noradrenaline release from rat hippocampal

permissive role on the activation of release-enhancing presynaptic alpha7 nicotinic receptors co-existing on rat neocortex glutamatergic terminals.

membrane-targeted luciferase reveals a new pathway for ATP secretion. Mol Biol


**4** 

Saša Branković *Clinic for Psychiatry,* 

*Serbia* 

**Assessment of Brain Monoaminergic Signaling** 

The majority of mental disorders (schizophrenia and other psychotic disorders, mood and anxiety disorders) have become treatable for the last several decades mostly due to development of the agents which modulate function of the brain dopaminergic, noradrenergic, and serotonergic system. The art of pharmacological healing in psychiatry consists actually in a great part in choice of the monoaminergic agent (i.e. antipsychotic or antidepressant) or their combinations which will modify the brainstem monoaminergic systems in an appropriate way. In other words, a daily problem in psychiatric practice could be realized as the task of estimation of the actual "neurochemical status" of the patient mostly in respect to the three brainstem monoamines. In clinical setting the task is still accomplished relying on the clinical picture and identifying the dominant symptoms that

The search for biological correlates (biomarkers) of neurochemical processes associated with mental disorders has not yet resulted in convincing and applicable findings (Bartova et al., 2010; Mössner et al., 2007). A reason that may account for the fail could be that attempts that have been made had been focused on a single neurotransmitter activity (Mössner et al., 2007). On the other hand, "Models which postulate too little or too much of a single neurotransmitter are not consistent with the complex regulation of neurotransmitter systems… There are major functional interactions among different neurotransmitter and neuropeptide systems which make single neurotransmitter theories simplisti A task for future investigations… is to develop clinically applicable biological tests that can assess the functional interactions among different neurotransmitter and neuropeptide systems and specific brain structures. Successful development of such paradigms could result in improved diagnostic classification and

In our interpretation of the task set by Charney & colleagues "functional interactions among different neurotransmitter and neuropeptide systems" are viewed as neurobiological signals in control system theory meaning of the word 'signal'. With this interpretation the task appear to be "to develop clinically applicable biological tests" which will allow insight into

the patient manifests (e.g. Nutt, 2008; Nutt et al., 2007).

prediction of treatment response…" (Charney et al., 1995).

**1. Introduction** 

**Through Mathematical Modeling of Skin** 

**Conductance Response** 

 *Clinical Center of Serbia, Belgrade* 

