Synaptic Plasticity Of Rat Retina After Acute High Intraocular Pressue And The Possible Modulatory Factors

PARTⅠExpression of marker proteins of synapse in rat retina after acute high intraocular pressureObjective To investigate the synaptic plasticity of rat retina after acute high intraocular pressure by observing the expression changes of synaptophsin(SYN),Vesicular glutamate transporter 1(VGLUT1)and Protein kinase Cα(PKCα).Methods Adult SD rats were divided into groups according to survival time.The intraocular pressure of the left eyes was elevated gradually until b wave of flash electroretinogram(fERG)disappeared and maintained for 60 minutes.The right eyes served as normal controls. After survival for 1,3,7 or 14 days,the serial frozen sectioned retinae were carried out with SYN immunohistochemistry and SYN mRNA in situ hybridization.Double labeling of SYN and VGLUT1,SYN and PKCαby immunofluorescence was also carried out.Results The synaptic plasticity was observed in rat retina after acute high intraocular pressure.It was mainly viewed as:The expression of SYN was up-regulated and the distribution of the protein was widened(It was also found in the inner part of the outer nuclear layer,where there was no distribution of SYN in normal condition).The expression up-regulation and widened distribution reached the peak at the 7^thday after acute high intraocular pressure,while reverted to normal pattern at the 14^thday.VGLUT1 had the same expression pattem as SYN and showed perfect co-localization with SYN in normal condition and at all time points after injury.PKCαshowed little changes in distribution after high intraocular pressure.Conclusion There was synaptic plasticity in rat retina after acute high intraocular pressure.It presented as expression up-regulation and widened distribution of pre-synaptic functional proteins in the outer plexiform layer and outer nuclear layer.While this kind of synaptic plasticity seemed be restricted in the pre-synaptic element. PARTⅡThe morphologic changes of synapses in rat retina after acute high intraocular pressureObjective To provide direct morphologic evidences of synaptic plasticity of rat retina after high intraocular pressure by observing the morphologic changes of ribbon synapses in the outer plexiform layer and the structures in the outer nuclear layer with electron microscopy.Methods Adult SD rats were divided into groups according to survival time.The intraocular pressure of the left eyes was elevated gradually until b wave of fERG disappeared and maintained for 60 minutes.The right eyes served as normal controls.After survival for 1,3, 7 or 14 days,animals were sacrificed and the inferior temporal quadrant of retinae were selected and fixed.After dehydration and embedding, ultrathin sections were cut with ultramicrotome and observed under electron microscope.Results Plenty of ribbon synapses were found in the outer nuclear layer in normal retina with standard characters:The electronic density ribbon was seen near and vertically standing to the pre-synaptic membrane.Synaptic vesicles around the ribbon were packed and well arranged.The synaptic cleft was seen between the thickened pre-and post-synaptic membrane.After acute high intraocular pressure,the number and the basic structures of ribbon synapse in the outer plexiform layer remained normal.While at the 3^rdand 7^thday after acute high intraocular pressure,bouton-like structures containing with vesicles and mitochondria were found in the outer nuclear layer,where normally be packed with nucleus of photoreceptors and absent of boutons and synapses.There wasn't standard synapse observed in the outer nuclear layer after injury.Conclusion These results indicated that the ectopic distribution of bouton-like structures in the outer nuclear layer after acute high intraocular pressure may be the morphologic basis of the widened distribution of the pre-synaptic functional proteins,such as SYN and VGLUT1. PARTⅢExploring of the possible factors modulating synaptic plasticity in rat retina after acute high intraocular pressureObjective To investigate the possible influence of BDNF and cholesterol on the synaptic plasticity in rat retina following acute high intraocular pressure.Methods Adult SD rats were divided into groups according to different pretreatment(the left eyes received vehicle,BDNF or BDNF antibody pretreated)and different survival time.2 days after pretreatment, the intraocular pressure of all left eyes was increased gradually until b wave of fERG disappeared and maintained for 60 minutes.The right eyes served as normal controls.1,3,7or 14 days later,serial frozen cross retinal sections of different groups were processed for SYN,glial fibrillary acidicprotein(GFAP)immunohistochemistry and FilipinⅢstaining.Results BDNF pretreatment resulted in delay of the widening of SYN distribution.The area of immuno-positive products of SYN in the outer plexiform and nuclear layer markedly increased at the 14^thday after acute high intraocular pressure.Contrarily,BDNF antibody pretreatment brought the time point ahead of schedule to the 3^rdday.Cholesterol content was increased after injury shown by FilipinⅢstaining and reverted to some extend at the 14^thday,and GFAP expression was also up-regulated after acute high intraocular pressure,indicating glia activation.Conclusion BDNF may affect synaptic plasticity of retina after acute high intraocular pressure by delay the regenerative changes through retinal ganglion cells protection and visual signal pathway stabilization. The activated retinal glia may participate the regulation of retinal synaptic plasticity through cholesterol release after acute high intraocular pressure.