| Dipfluzine (Dip), a new diphenylpiperazine compoud, was first developed by School of Pharmacy, Hebei Medical University. Its effects of selective cerebral vasodilation, increasing cerebral blood flow, improving energy metablism, decreasing concentration of extracellular excitatory amino acids, and blocking calcium channels are confirmed. From all these we can see that Dip maybe is a good protector to cerebrum. In order to go on studying the protective effects of Dip, we take Aβ25-35 and AlCl3 as models and observe its effects on cultured hippocampal neuron. Our purpose is to observe whether Dip possesses the protective effects against the injury in cultured hippocampal neuron of foetal rats induced by Aβ25-35 and AlCl3. If it has the protective effects, we continue to explore whether it is related to its effects on intracellular calcium overload. The projects and results were as follow:1 The culture of hippocampal neuronObjective: To establish the method of the culture of hippocampal neuron and choose the best grown status.Methods: We used foetal rats of 18 days. Detached its hippocampal tissue, got cell suspension by digesting, flushing, suspending. Cultured in culture broad of 24 holes. The maximal diametes, minimal diametes and soma area in different periods (1st, 3rd, 7th, 14th, 21th) were observed by Laser Scanning Confocal Microscopy (LSCM) and the best grown status was chose. Results: The hippocampal neurons grew quickly in the first three days. The maximal diametes (15.1±2.5), minimal diametes (8.8±1.1) and soma area (119±25) in the third day were all increased markedly (p<0.05) compared with the maximal diametes (12.7±2.4),minimal diametes (7.8±1.1) and soma area (87±17) in the first day. From the third day to the fourteenth day the hippocampal neurons grew steadily. After 21 days, the hippocampal neurons began to shrink. The maximal diametes, minimal diametes and soma area began to decrease. The enations began to light and dissociation.Conclusion: The hippocampal neurons grew quickly in the first three days and got a steady grown status in two weeks. It shows us that it took about one week the hippocampal neurons got its best grown status. The adminstration should be in this periods.2 Protective effects of Dipfluzine on the injury in cultured hippocampal neuron of foetal rats induced by Aβ25-35Objective: To study the effects of Aβ25-35 on hippocampal neuron of foetal rats and the protective effects of Dip .Methods: Added four different concentrations of Aβ25-35 (0.01μmol/L, 0.1μmol/L, 1μmol/L, 10μmol/L) to the hippocampal neurons which cultured for 7 days and fostered together. After 24 hours, assayed the lactate dehydrogenase release( LDH %) by the lactate dehydrogenase Kit. Choose the concentration which could lead to injury of the hippocampal neurons as the concentration of model group. We use Nimodipine as the contrast. Added three different concentrations of Dip (0.1μmol/L, 1μmol/L, 10μmol/L) to the hippocampal neurons which cultured for 7 days and fostered together for 24 hour. Assayed LDH % by the lactate dehydrogenase Kit. Assayed the level of intracellular free calcium fluorescence intensity by LSCM.Results: Aβ25-35 could cause injury to the hippocampal neurons. The LDH% of 0.1μmol/L Aβ25-35, 1μmol/L Aβ25-35, 10μmol/L Aβ25-35 (43±3, 49±4, 39±3) were all increased significantly, compared with the blank group (31±3, p<0.05). We choose the 1μmol/L Aβ25-35 as the model group, which could cause the injury markedly. The LDH% decreased after added Dip. The model group was 48±4,far higher than the blank group (31±5). The contrast group Nimodipine (37±4, p<0.05), 1μmol/L Dip ( 37±5, p<0.01), 10μmol/L Dip (34±5, p<0.01) were all decreased compared with the model group. Assayed the level of intracellular free calcium fluorescence intensity by LSCM, The model group was 122±21,far higher than the blank group (72±14),The contrast group Nimodipine (92±16, p<0.01),1μmol/L Dip (90±13, p<0.01), 10μmol/L Dip (84±15, p<0.01) wer...
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