Changes of free hydrogen in neurons after hydraulic impact injury and its influencing factors Zhang Xiangyu Liu Enzhong Liu Xiaoqian Su Junwu Pretty Dai Qinyu] i) changes to explore nimodipine (Nim), D2 aminovaleric acid (DAP5) and hypothermia on post-traumatic neurons The influence and mechanism of changes in [H]i. Methods BCECF AM was used as a fluorescent indicator of intracellular hydrogen ions. The changes of [H]i in rat neurons cultured by hydraulic impact injury were measured by laser confocal microscopy. Results The cerebral cortical neurons [ H ] i after hydraulic impact injury can reduce [ H ] i within 4 h after injury, while DAP5 is effective within 10 h after injury, and the application of hypothermia within 30 min can be reduced. The effect of [H] i in cells is better than that of Nim (P 0 .001). Conclusion Hydrocrine injury caused neuronal acidosis, Nim, DAP5 and mild hypothermia all decreased intracellular] i, but the effective time window of each effect was different, revealing that the post-traumatic neuronal acidosis should pay attention to comprehensive treatment, and select each The best time window for action.
Author: 150001 Department of Neurosurgery, the First Affiliated Hospital of Harbin Medical University (Zhang Xiangxuan, Department of Pathophysiology, Beijing Institute of Neurosurgery, 100050) Brain trauma is one of the important causes of death and disability in today's adolescents. In addition to the primary mechanical damage, the neurological damage after brain trauma is more conspicuous. These secondary pathological lesions are closely related to neurobiochemical reactions following brain trauma, including intracellular acidosis. Understanding the mechanism of acidosis in nerve cells after trauma is of great significance for the improvement of wound prognosis. The authors focused on the effects of nimodipine (Nim), D 2 aminovaleric acid (D AP 5) and mild hypothermia on [H] i in post-traumatic neuronal cells, and analyzed its possible mechanism of action.
Materials and methods 1. Cell culture: 104 cerebral cortex tissues of newborn Wistar rats were dissected into single cell suspension, and pre-washed with polylysine (American Sigma) in a 50 ml culture flask. On the sheet, the medium containing the volume fraction was cultured at 37 ° C, and the culture solution was RPMI 1640 containing fetal bovine serum with a volume fraction of 10. After 8 to 14 days of primary culture, the study was carried out. Neuron enolase (NSE) immunohistochemical staining showed 40 positive, and glial fibrillary acidic protein (GFAP) immunohistochemical staining showed 60 positive.
2. Hydraulic impact injury test: The hydraulic impactor device refers to the model of Scott et al., the impact pressure is 2.5 kPa, and the action time is 20-30 ms.
3. Experimental grouping: (1) Normal control group. (2) Trauma group: It was further divided into trauma treatment, and the effect on [H] i in cells was observed after 2 h. The number of rats in each of the above experimental groups was 8.
4. Determination of [H]i in neurons: Neurons cultured on coverslips were washed 3 times with phosphate buffered saline (PBS) and then placed on a chamber at 37 °C with Insight plus IQ laser confocal microscope 488 The nm argon ion laser was excited and the fluorescence of 530 640 nm emission was recorded to find the ratio of the two. The intracellular pH was determined from the standard curve.
The standard curve of intracellular pH was directly calibrated: the ratio of excitation fluorescence of BCECF in high potassium buffer, nm) cells was prepared, and the standard curve of the ratio and pH was determined.
5. Statistical analysis: The results were processed by SAS software system, expressed as x±s, and the comparison between groups was performed by two-sample heteroscedastic t test.
Results 1. Changes in [H]i in neurons after hydraulic impact injury: Compared with the normal control group, [H]i in the neurons at 0. 25 h after injury increased gradually, reaching a peak at 12 h after injury. , followed by [ H ] i gradually] i changes 2. Nim, D AP 5 and mild hypothermia on the changes of [ H ] i in neurons after hydraulic impact injury: Nim treatment group can be inhibited within 4 h after injury Intracellular [H above the application is ineffective (P 0 .05). The D AP 5 treatment group was significantly inhibited within 4 h after injury [more than H (P 0.05). The hypothermia treatment group was able to reduce [H cells within 0.5 h after injury [H see Figure 2.
The influence of i discusses acidosis is an important aspect of the process of secondary pathological damage in traumatic brain injury. Therefore, direct detection of intracellular [H]i is one of the important means to further explore the neurochemical changes in brain trauma. Previous studies have mainly used biochemical analysis of plasma lactic acid content or microdialysis technology to monitor lactic acid content in brain tissue extracellular fluid and other methods to indirectly reflect brain acidosis. The authors used the intracellular H fluorescent probe BCECF AM to directly detect the changes of [H]i levels in the cytoplasm of individual neurons at different time after brain hydraulic impact injury, and found that there was acidosis in neurons after brain trauma. The intracellular [H]i increased at 0.25 h after injury and reached a peak at 12 h, then gradually decreased, and remained at a high level for 48 h. The reason for the increase of [H]i in the neuronal cell body after brain trauma is not completely clear, and may be related to the following two aspects: (1) a large outflow of K in the neuron after brain trauma, which leads to depolarization of the cell membrane. Depolarization in turn induces the release of excitatory amino acids (EAA), which further promotes K efflux, while Ca influx, thus forming a vicious circle between ion transmembrane abnormalities and EAA release. In order to maintain normal ion balance inside and outside the cell, this abnormal transmembrane ion barrier activates an energy-dependent ion pump and stimulates the hydrolysis of ATP, eventually enhancing glycolysis, resulting in accumulation of lactic acid and formation of intracellular acidosis. (2) The release of gamma aminobutyric acid (GABA) after brain trauma activates the Cl channel, which causes the HCO outflow and simultaneous glutamate release to activate the N-methyl D-naproline (NMDA) receptor, thereby influx H, thus forming The extracellular fluid is briefly alkaline. This transient extracellular fluidization, intracellular H elevation, and membrane depolarization activate the Na exchange pump. In addition, protein kinase C (PKC) coupled to the EAA receptor after brain trauma can also be activated, thereby phosphorylating the Na exchange pump and activating the exchange pump. The indirect dependence of the Na pump is directly dependent on the large consumption of Na ATP. On the other hand, it stimulates the anaerobic glycolysis to enhance, eventually producing lactic acid accumulation and intracellular acidosis. It can be seen that the activation of EAA receptors and the activation of Na pumps and Na pumps are an important source of lactic acid accumulation.
After brain trauma, neuronal cell membrane depolarization, voltage-dependent calcium channel is open, and nimodipine specifically blocks the opening of the channel, thereby reducing the abnormal distribution of transmembrane Ca, thereby reducing the consumption of ATP by Ca pump, and finally producing H. Reduced and reduced glycolysis. This] i competes for intracellular protein binding sites, and the influx decreases to reduce [H]i. The results of this experiment showed that nimodipine was effective within 4 h after trauma and was ineffective for more than 10 h. This suggests that early Ca influx is likely to be dominated by voltage-dependent channels after brain trauma, while the number of NMDA receptors that may be activated by receptor-dependent channels in the late stage is reduced, thereby reducing Ca influx and K outflow, and restoring transmembrane ion anomalies. The disorder reduces glycolysis, which in turn reduces the increase in [H]i. The authors used the NMDA receptor competitive antagonist D AP 5, which was best used within 4 hours after trauma, and the application was not effective within 4-10 hours, but it was ineffective for more than 22 hours. This indicates that the release of EAA is mainly within 10 h after injury, while the release of EAA is reduced on the one hand over 10 h, and the formation of organic acids is increased due to other biochemical cascades (such as arachidonic acid), and intracellular Ca overload The interaction of free radical formation and acidosis eventually leads to the formation of acidosis in the late stage of trauma, which is not limited to the ion disturbance mediated by EAA receptors. The protective effect of mild hypothermia is mainly to reduce the release of EAA (2) by reducing the ATP consumption [8], thereby reducing H production on the one hand, and reducing the stimulation of glycolysis on the other hand (3) inhibiting the activity of PKC [ 9], thereby inhibiting the phosphorylation of the G protein receptor by PKC, thereby controlling the Na influx, alleviating the transmembrane ion disorder, and finally weakening the glycolysis. The author observed that the application of the hypothermia treatment group was effective within 0.5 hours after injury, but the application was not effective for more than 1 hour. The reason may be that more than 1 h after injury, EAA release increased, and PKC has been activated, ATP consumption increased, and eventually the application of sub-hypothermia is not effective for more than 1 h.
The results of this experiment show that Nim, D AP 5 and mild hypothermia protect post-traumatic neurons from different mechanisms, and the effective treatment time window is different. It is suggested that the clinical treatment of post-traumatic neuronal acidosis should pay attention to comprehensive treatment, and select the best time window for each effect. This experiment is based on a traumatic model at the cellular level. As for the overall level of effect, further research is needed.
(Editor: Jiang Wei)? notice?
Baiyou Absorbable Internal Fixture General Agent Relocation Notice Beijing Putai Technology Developers Agency moved to the new office on November 8, 2000. Beijing Baiyou Putai Medical Products Co., Ltd. is officially operating at the new site. If you want to contact Finnish Baiyou absorbable fracture internal fixation and German aap groove interlocking intramedullary nail, please contact us as follows. Address: 17F (100029), Mingzhu Building, Shenfang Building, 23rd Floor, 4th Floor, Anzhen Xili District, Chaoyang District, Beijing. Tel: 010 Finnish Baiyou absorbable fracture internal fixation German aap groove interlocking intramedullary nail China general agent Beijing Baiyou Putai Medical Products Co., Ltd. Beijing Putai Technology Developers Association
LED Step Light
The Step Light install in wall,step,corner. has the characteristics of small size, low power consumption, long service life, sturdy and durable. Low power consumption, long life, easy installation, chic and elegant shape, anti-leakage, waterproof;
1. The LED light source has a long life, no accidents and almost no need to change the bulb, one time construction, several years of use.
2. Low power consumption, no need to pay high electricity bills for lighting and beautification.
3. Waterproof, dustproof, pressure resistant and corrosion resistant
Our other products range:LED Underground Light, LED Underwater Light, LED Wall Washer Light, LED Linear Light , LED Outdoor Flood Light, LED Garden Light , LED landscape light , LED Strip Light , LED Step Light etc.
LED stair Lamp,LED Foot Lamp,Waterproof Step Lamp,Stair Step Lamp,LED Corridors Light,Corner light
SHENGYA LIGHTING TECHNOLOGY CO., LTD. , https://www.syalighting.com