Protective Effects Of Hydrogen Sulifde On Neuronal Injury Induced By Ischemia In Rats

Objective⑴To investigate whether exogenous hydrogen sulfide （H₂S） attenuates theneuronal injury in the hippocampal CA1region induced by chronic brain ischemia.⑵To address the neuroprotective effect of H₂S on aberrant glial cells inglioblastoma （GBM）.⑶To explore the mechanism of the neuroprotective effect of H₂S on ischemicneurons in hippocampus as well as glial cell in GBM.MethodsIn the present study, experiments were divided into3parts. In the first andsecond parts, a global model of cerebral ischemia was established by two vesselocclusion （2VO）, and the experimental rats were divided into4groups: sham group,H₂S group （NaHS injected,5.6mg/kg/day, i.p.）,2VO group and2VO-H₂S group（NaHS injected post2VO,5.6mg/kg/day, i.p.）. For the first part experiment, theperformances of learning and memory were examined by the Morris water maze（MWM）. As an H₂S donor, NaHS was administered intraperitoneally （5.6mg/kg/day,i.p.）. The morphology of neurons in hippocampus was determined by hematoxylinand eosin （HE） staining, and H₂S content in hippocampus was evaluated. In thesecond part of the present experiments, LTP from hippocampus Schaffer collaterals toCA1region was performed. Before the LTP induction, the local field potentials （LFPs）were recorded. Growth-associated protein-43（GAP-43） expression in thehippocampus of each rat was analyzed by immunohistochemical staining. In addition,a novel general partial directed coherence （gPDC） algorithm was employed todetermine the coupling interaction between CA3and CA1in hippocampus in twofrequency bands （theta rhythm and gamma rhythm）. In the third part of the presentstudy, in vivo GBM model was conducted using adult rats with intracerebral injectionof rat C6glioma cell line, and another4groups were involved: sham group, H₂Sgroup, GBM group and GBM-H₂S group. To observe the H₂S enhance effect onangiogenesis in GBM, an intraperitoneal injection of NaHS was administrated. HE staining, angiogenesis examination and immunohistochemical analysis of thehypoxia-inducible factor-1alpha （HIF-1α）, matrix metalloproteinase-2（MMP-2） andCD34expressions were performed, respectively. Each section stained withCD34-related antigen was also to highlight the blood vessels for evaluating the tumormicrovessel density （MVD-CD34）.Results⑴Exogenous H₂S significantly improved spatial learning and memory deficitsinduced by brain ischemia （P＜0.01）. Exogenous H₂S inhibited the edema aroundpyramidal neurons and the nuclear shrink induced by ischemia. Intraperitonealinjection with a certain concentration of NaHS （5.6mg/kg/day） could increase thecontent of H₂S in hippocampus of2VO animals. A subnormal level of H₂S content inhippocampus was existed in2VO rats. The H₂S level in hippocampus in2VO-H₂Sgroup was elevated after treated with NaHS, while it was lower than that in shamgroup. It can be seen that the H₂S level in hippocampus was associated with the dataobtained from behavior and morphology tests.⑵Exogenous H₂S enhanced the LTP in the hippocampus of2VO rats, andpromoted the expression of GAP-43in the CA1region of hippocampus post ischemia.The phase locked values （PLV） were significantly increased in theta and gammarhythms after H₂S treatment in2VO rats. The unidirectional influence from CA3toCA1reduced significantly at theta and gamma rhythms in2VO rats, and enhancedafter H₂S treatment, which were associated with the LTP alterations.⑶After NaHS injection, the clinical symptoms of tumor-bearing rats becamemore serious, and the dramatic distress and obvious weight loss were found. HEstaining showed more evidence of tumor in GBM-H₂S animal brains than that inGBM animal brains. The mean tumor volume became much larger in GBM-H₂S ratsthan that in GBM animals （P＜0.001）. Immunohistochemical analysis exhibited thatthe HIF-1α, MMP-2and CD34expressions were obviously increased after NaHSintraperitoneal injection in GBM-H₂S rats. And the MVD-CD34was markedly higherin GBM-H₂S group.Conclusions⑴the H₂S content in hippocampus could be increased by treating with a certain concentration of exogenous H₂S. H₂S could improve impairment of learning andmemory in2VO rats. The underlying mechanism was associated with improvementof the edema around pyramidal neurons and the nuclear shrink induced by ischemia.⑵Exogenous H₂S enhanced synaptic plasticity in the hippocampus of2VO rats,The underlying mechanism was associated with H₂S promoted the expression ofGAP-43in the CA1region of post-ischemic hippocampus. The changes of oscillatorypattern in hippocampal neurons induced by2VO got retrieved in a degree by H₂Streatment, which might represent a novel recognition to the mechanism of H₂Simproving cognitive impairments.⑶H₂S serves as a stimulator in the angiogenesis of GBM via multiplemechanisms including the increase of HIF-1α and MMP-2.⑷Exogenous H₂S has neuroprotective effects on both pyramidal neurons in theCA1region of hippocampus and glial cell in GBM when ischemic injury existed inthese two kinds of neuron.