| Background: Many severe neurological and psychiatric diseases such as cerebrovascular disease, brain traum, brain tumor and the following intracerebral syndrome, anxiety, depresser, epilepsy, dementia may vary in pathogenies, however, they are confirmded to be caused at least by glutamate-induced excitotoxicity. The N-methyl-D-aspartate receptor (NMDAR) plays a pivotal role in the process of glutamate-induced excitotoxicity. In the past decades, a large variety of NMDAR antagonists have been evaluated for their potential neuroprotective effects in clinic. However, clinical trials with NMDA antagonists have failed for the reasons being side effects of these drugs that occur because of high binding affinity towards NMDA receptors, differential actions on neurons from various regions of brain and interaction with neurotransmitter receptors other than NMDA receptors. Among many neuroprotective drugs, extracts of Ginkgo biloba (GBE) as a neuroprotective drug has attracted much attention. Initially it was conceived as a novel therapeutic principle with a combination of preventive and curative effects on diverse pathologies associated with vascular occlusions, and other circulatory disturbances. Since then a plenty of evidences show that GBE benefits in a variety of pathological conditions of the brain associated with excitotoxicity. Numerous experiments have already demonstrated that GBE has many clinical therapeutic applications not only for cerebral (or cerebrovascular) insufficiency, coronary artery disease, hypertension, angina, arteriosclerosis, but also for cognitive speed, dementia, and aging damages. GBE has been well prescribed as a neuroprotective drug in 1980s. However, the common GBE is microsome, which has difficulty in crossing the brain blood barrier because of big molecule diameter. Furthermore, GBE is a complex mixture containing flavonoid glycosides, terpene lactones (non-flavone fraction) and various other constituents,Which only dissolve in organic compounds. All of these limit the application of this herb drug in clinic. Therefore it is important to explore a way to enhance the neurprotective effects of GBE. Our group adopts Supercritical Anti-Solvent (SAS) upon CO2-supercritical fluid extraction (CO2-SPF) to prepare Nano-GBE, Which molecule diameters are smaller than 100nm in electron microscope. Our aim is to explore the neuroprotective effect of GBE by means of patch clamp technique, to further study the pharmacological mechanisms of GBE. Objectives and Methods: In this study, our group adopts Supercritical Anti-Solvent (SAS) upon CO2-supercritical fluid extraction (CO2-SPF) to prepare Nano-GBE, Which molecule diameters are smaller than 100nm in electron microscope. To further explore the neuroprotective effects of Nano-GBE, whole–cell patch clamp recording was carried out to evaluated the modulatory effects of Micro-GBE (dissolved in DMSO)/Nano-GBE (dissolved in the standard extracellular solution) on NMDAR-activated currents in acutely isolated rat hippocampal neurons, the difference between Nano-GBE and Micro-GBE in respect of protective potency and to determine the possible modes of actions. Results: 1. The majority of the neurons examined (81.8%, 90/110) weresensitive to NMDA (1mM). NMDA activated an inward current, which manifested minimum apparent desensitization and could be blocked by MK-801, the specific antagonist of NMDAR. 2. After the neurons were treated with Micro (dissolved in DMSO) /Nano GBE (dissolved in the standard extracellular solution) (0.1mg/ml) prior to the application of NMDA (1mM) and Gly (10μM) for 30s, NMDA-activated currents was obviously inhibited (n=8). The modulatory effect of Nano-GBE (dissolved in the stander extracellular solution) on NMDA-activated current is significantly different from that of Micro-GBE (dissolved in DMSO) (P < 0.05). 3. The NMDA-activated current could be blocked by MK-801 (50μM), this inhibitory effects is a little higher than that of Nano-GBE (dissolved in the standard extracellular solution), But the difference between them h...
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