Maraula Giovanna, Lana Daniele, Coppi Elisabetta, Gentile Francesca, Mello Tommaso, Melani Alessia, Galli Andrea, Giovannini Maria Grazia, Pedata Felicita, Pugliese Anna Maria
Dept. of Neuroscience, Psychology, Drug Research and Child Health, NEUROFARBA, Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.
Dept. of Health Sciences, Clinical Pharmacology and Oncology Unit, University of Florence, Florence, Italy.
PLoS One. 2014 Dec 19;9(12):e115273. doi: 10.1371/journal.pone.0115273. eCollection 2014.
Purinergic P2X and P2Y receptors are broadly expressed on both neurons and glial cells in the central nervous system (CNS), including dentate gyrus (DG). The aim of this research was to determine the synaptic and proliferative response of the DG to severe oxygen and glucose deprivation (OGD) in acute rat hippocampal slices and to investigate the contribution of P2X7 and P2Y1 receptor antagonism to recovery of synaptic activity after OGD. Extracellular field excitatory post-synaptic potentials (fEPSPs) in granule cells of the DG were recorded from rat hippocampal slices. Nine-min OGD elicited an irreversible loss of fEPSP and was invariably followed by the appearance of anoxic depolarization (AD). Application of MRS2179 (selective antagonist of P2Y1 receptor) and BBG (selective antagonist of P2X7 receptor), before and during OGD, prevented AD appearance and allowed a significant recovery of neurotransmission after 9-min OGD. The effects of 9-min OGD on proliferation and maturation of cells localized in the subgranular zone (SGZ) of slices prepared from rats treated with 5-Bromo-2'-deoxyuridine (BrdU) were investigated. Slices were further incubated with an immature neuron marker, doublecortin (DCX). The number of BrdU+ cells in the SGZ was significantly decreased 6 hours after OGD. This effect was antagonized by BBG, but not by MRS2179. Twenty-four hours after 9-min OGD, the number of BrdU+ cells returned to control values and a significant increase of DCX immunofluorescence was observed. This phenomenon was still evident when BBG, but not MRS2179, was applied during OGD. Furthermore, the P2Y1 antagonist reduced the number of BrdU+ cells at this time. The data demonstrate that P2X7 and P2Y1 activation contributes to early damage induced by OGD in the DG. At later stages after the insult, P2Y1 receptors might play an additional and different role in promoting cell proliferation and maturation in the DG.
嘌呤能P2X和P2Y受体广泛表达于中枢神经系统（CNS）的神经元和神经胶质细胞上，包括齿状回（DG）。本研究的目的是确定急性大鼠海马切片中DG对严重氧糖剥夺（OGD）的突触和增殖反应，并研究P2X7和P2Y1受体拮抗剂对OGD后突触活动恢复的作用。从大鼠海马切片记录DG颗粒细胞的细胞外场兴奋性突触后电位（fEPSP）。9分钟的OGD导致fEPSP不可逆转地丧失，并总是伴随着缺氧去极化（AD）的出现。在OGD之前和期间应用MRS2179（P2Y1受体的选择性拮抗剂）和BBG（P2X7受体的选择性拮抗剂），可防止AD出现，并使9分钟OGD后神经传递显著恢复。研究了9分钟OGD对用5-溴-2'-脱氧尿苷（BrdU）处理的大鼠制备的切片颗粒下区（SGZ）中细胞增殖和成熟的影响。切片进一步与未成熟神经元标记物双皮质素（DCX）一起孵育。OGD 6小时后，SGZ中BrdU+细胞的数量显著减少。这种作用被BBG拮抗，但未被MRS2179拮抗。9分钟OGD后24小时，BrdU+细胞的数量恢复到对照值，并观察到DCX免疫荧光显著增加。当在OGD期间应用BBG而非MRS2179时，这种现象仍然明显。此外，此时P2Y1拮抗剂减少了BrdU+细胞的数量。数据表明，P2X7和P2Y1的激活促成了OGD在DG中诱导的早期损伤。在损伤后的后期阶段，P2Y1受体可能在促进DG中的细胞增殖和成熟方面发挥额外且不同的作用。
