RESUMO
Communication between neuronal and glial cells is important for many brain functions. Astrocytes can modulate synaptic strength via Ca(2+)-stimulated release of various gliotransmitters, including glutamate and ATP. A physiological role of ATP release from astrocytes was suggested by its contribution to glial Ca(2+)-waves and purinergic modulation of neuronal activity and sleep homeostasis. The mechanisms underlying release of gliotransmitters remain uncertain, and exocytosis is the most intriguing and debated pathway. We investigated release of ATP from acutely dissociated cortical astrocytes using "sniff-cell" approach and demonstrated that release is vesicular in nature and can be triggered by elevation of intracellular Ca(2+) via metabotropic and ionotropic receptors or direct UV-uncaging. The exocytosis of ATP from neocortical astrocytes occurred in the millisecond time scale contrasting with much slower nonvesicular release of gliotransmitters via Best1 and TREK-1 channels, reported recently in hippocampus. Furthermore, we discovered that elevation of cytosolic Ca(2+) in cortical astrocytes triggered the release of ATP that directly activated quantal purinergic currents in the pyramidal neurons. The glia-driven burst of purinergic currents in neurons was followed by significant attenuation of both synaptic and tonic inhibition. The Ca(2+)-entry through the neuronal P2X purinoreceptors led to phosphorylation-dependent down-regulation of GABAA receptors. The negative purinergic modulation of postsynaptic GABA receptors was accompanied by small presynaptic enhancement of GABA release. Glia-driven purinergic modulation of inhibitory transmission was not observed in neurons when astrocytes expressed dn-SNARE to impair exocytosis. The astrocyte-driven purinergic currents and glia-driven modulation of GABA receptors were significantly reduced in the P2X4 KO mice. Our data provide a key evidence to support the physiological importance of exocytosis of ATP from astrocytes in the neocortex.
Assuntos
Trifosfato de Adenosina/metabolismo , Astrócitos/metabolismo , Neocórtex/metabolismo , Inibição Neural/fisiologia , Neurônios/metabolismo , Animais , Astrócitos/citologia , Bestrofinas , Cálcio/metabolismo , Comunicação Celular , Exocitose , Proteínas do Olho/genética , Proteínas do Olho/metabolismo , Regulação da Expressão Gênica , Ácido Glutâmico/metabolismo , Canais Iônicos/genética , Canais Iônicos/metabolismo , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Transgênicos , Neocórtex/citologia , Neurônios/citologia , Técnicas de Patch-Clamp , Canais de Potássio de Domínios Poros em Tandem/genética , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Receptores de GABA-A/genética , Receptores de GABA-A/metabolismo , Receptores Purinérgicos P2X4/deficiência , Receptores Purinérgicos P2X4/genética , Proteínas SNARE/genética , Proteínas SNARE/metabolismo , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/metabolismoRESUMO
The importance of communication between neuronal and glial cells for brain function is recognized by a modern concept of 'tripartite synapse'. Astrocytes enwrap synapses and can modulate their activity by releasing gliotransmitters such as ATP, glutamate and D-serine. One of the regulatory pathways in the tripartite synapse is mediated by P2X purinoreceptors. Release of ATP from synaptic terminals and astrocytes activates Ca(2+) influx via P2X purinoreceptors which co-localize with NMDA (N-methyl-D-aspartate) and GABA (gamma-aminobutyric acid) receptors and can modulate their activity via intracellular cascades which involve phosphatase II and PKA (protein kinase A).
