The Effect And Mechanism Of Prophylactic Use Of Nimodipine In The Protection Of Facial Nerve

Background and purposes of the research:Great advances in neuroimaging, intraoperative cranial nerve monitoring, and microsurgical technique have shifted the focus of vestibular schwannomas surgery from prolonging life to preserving cranial nerve function in patients. However, the function impairment of nerves after surgery remained the most difficult problem of the treatment of vestibular schwannomas. Facial paralysis is a major complication of vestibular schwannomas surgery.It is the key how not only to resect the tumor but also to preserve the function of facial nerve for the treatment of vestibular schwannomas.To explore methods of protection of the facial nerve, to retain the pursuit of the anatomy of facial nerve function preservation even has been a hotspot research of vestibular schwannomas.It was reported that nimodipine can accelerate the regeneration of rat facial nerve axons, increase the expression of glial fibrillary acidic protein (GFAP), promote the growth of motor neurons.The results were significant for a better outcome after vestibular schwannoma surgery for both hearing and facial nerve preservation in the group of patients who received a prophylactic vasoactive prophylaxis consisting of nimodipine and hydroxyethylstarch in Clinical studies. The effect of Nimodipine alone for the role of acoustic neuroma patients has not been described, the vasoactive effect of nimodipine may improve the intraneural microcirculation of the facial and cochlear nerves, but it was not conducted in-depth discussion.The protective mechanism of nimodipine on the brain and neuronal cells has the following aspects:①blocking voltage-dependent Na+ or/and Ca+ channel;②activateγ-aminobutyric acid (GABA)-induced Cl- influx;③blockα-amino-3-carboxy -5-methyl-Isoxazole-4-propionic acid (AMPA) and kainic acid (KA) AMPA /KA-type glutamate receptors;④inhibiting the activity of carbonic anhydrase;⑤reduce mitochondrial damage, but its exact mechanisms are not yet clear.Calcium overload is an important mechanism of ischemic injury, neuronal apoptosis and it is considered to be the common pathway of cell death after ischemic injury. As the dihydropyridine calcium antagonist and the L-type voltage-sensitive receptor calcium channels, its receptor of Nimodipine, mainly located in the nerve cell bodies, a combination of both to enable to the reduction of intracellular Ca2+; and nimodipine priority combined with local ischemic area. Nimodipine used extensively in clinic, proved to have neurotrophic effects, and can pass through the blood-brain barrier. With regard to the application of time, delivery time, long-term effects, nerve protection mechanisms of Nimodipine, these still need further study.Facial nerve injury is mainly mechanical stretch and electrical coagulation of the thermal damage.The pathological and physiological changes of facial nerve injury has been not observed, if the experiment can copy such a model of thermal damage and the facial nerve stretch injuries can be help clinicians understand and judge the degree of facial nerve injury and patho-physiological changes.There are no injuries reported in the literature which includes both the stretch and damage electrocoagulation applied to facial nerve injury of animal model studies. This topic aims at establishing a rat facial nerve combined injury model. To make a similar facial nerve injury model with acoustic neuroma. To observe the pathology, physiological changes of facial nerve injury, while observing the effect of Nimodipine in facial nerve injury model.Glial cell line-derived neurotrophic factor and brain-derived neurotrophic factor are the members of neurotrophin family, they are important sensory and motor neuron trophic factor. The role of GDNF and BDNF mainly include:①regulate normal growth of nerve cells;②resist the injury of stimulus,to promote regeneration of neuron, protect the ischemic and hypoxic neurons, and inhibit neuronal apoptosis, induce axon regeneration.The possible protective mechanism of BDNF and GDNF against the injury of neuron as follows:antagonism of glutamate cytotoxicity and intracellular calcium overload. BDNF, GDNF can increase the expression of superoxide dismutase and glutathione peroxidase, reduce the production of free radicals, thereby reduce the damage of free radicals on neuronal. As a calcium antagonist, the effect of nimodipine on the expression of GDNF and BDNF and the neurotrophic effects remain unclear.In clinical study, nimodipine were used prophylacticly in patients with acoustic neuroma, We evaluate facial nerve function according to House-Brackmann score and to explore new ways to protect the function of facial nerve.The subject of innovation are:1,make a facial nerve complex damage model; 2, observe facial nerve pathology and analysis the mechanisms of prophylactic use of nimodipine; 3 observe the protective effect of nimodipine in clinical preventive application; 4 study the expression of GDNF and BDNF prophylactic by use of nimodipine on the facial nerve protection.Materials and methods:Chapter One:The application and assessment of rats model with facial nerve injury. 1. Animal model:8 male Sprague-Dawley rats were random distribute, these animals were own control on both sides of the face, divided into two groups, each group has 8 sides:the right to contralateral control; the left facial nerve buccal branch for the production of facial injury model.2. In the experiment model, we separate 10mm length the buccal branch of facial nerve. Using microvascular clips, we can make the left facial nerve crushed, it spends 30 seconds. Coaxial bipolar electrode (using one side as a unipolar stimulation) was used on the branch of facial nerve, the power of injury was 12.5watt.3. After 2 weeks, we examined compound action potential amplitude, latency and nerve conduction velocity of the left orbicularis muscle by nerve electrophysiological monitoring.4. Specimen obtained and HE staining were observed.5. Statistics software SPSS 13.0 was used, the facial nerve action potential latency, amplitude and nerve conduction velocity with paired t test, P<0.05 indicated significant difference.ChapterⅡ:The prophylactic use of nimodipine in the rat facial nerve injury1.96 rats, animals were randomly divided into two randomized block design into four groups:sham operation group, injury group, nimodipine pretreatment group, nimodipine after treatment group, n= 24. rats were killed at 1 month,3 months, 6 months points in batches.2. Examine action potential amplitude, latency and nerve conduction velocity by neural electrophysiological examination at 1 month,3 month,6 month, animals were sacrificed, tissue specimens were examined by HE staining and Biels-chowsky staining of nerve fibers after neural electrophysiological examination. 3. The expression of GDNF and BDNF in tissue of facial nerve was examined by Western Blot at 1 month,3 months,6 months.4. Statistics software SPSS 13.0 was used, the facial nerve nerve conduction velocity, action potential amplitude and latency with a mean±standard deviation ((?)±s), the facial nerve nerve conduction velocity, action potential amplitude and latency compare between the two groups by the analysis of variance (one -way ANOVA), homogeneity of variance with LSD, missing variance with Tamhane's T2, P<0.05 indicated significant difference.ChapterⅢ:The preliminary observations of nimodipine prophylaxis in vestibular schwannoma1. There were 32 cases with acoustic neuroma surgery from December 2008 to December 2009, all these cases were diagnosed with acoustic neuroma by pathology examination and MRI,the results showed that the facial nerve had been anatomic preservation.2. Treatment:There were 5 patients in nimodipine pretreatment group, they were given nimodipine tablets 3 days before surgery. Given intravenously nimodipine injection after operation. There were 6 patients in nimodipine aftertreatment group, they were given nimodipine tablets 7 days after 2 days of surgery. There were 21 patients in untreated group, they were treated with conventional methods without Nimodipine.3. Physiological monitoring Methods:Cascade IOM nerve monitoring system made by American Cadwell corporation was used.Stimulus current flow:0.1～10mA, wave width 0.2ms, frequency 1Hz, smoothing range:10-3000Hz, spontaneous and evoked electromyogram were recorded.4. Postoperative assessment:①All cases were examined by CT or MRI;②facial nerve function were assessed by House-Brackmann scores;③All of the cases regular follow-up postoperation, three months to one year follow up.Results:ChapterⅠ:1. When dissecting the facial nerve, we found the facial nerve close to the surface of masseter muscle is divided into temporal masseter branch, zygomatic branch, buccal branch, marginal mandibular and cervical branches of the upward, which distributed to the nasal part, upper/lower lips and facial muscles.2.8 All rats survived after surgery, all rats on the left side and the towering bearded-lip sip activities significantly decreased, the left caudal whiskers extendedto the tail straightly.2 weeks, mild atrophy can be seen on the left facial muscles, left beard can be seen a slight jitter. The function of control side was perserved well compared with the side of sham effect.3. Electrophysiological examination:after 2 weeks, nerve conduction velocity action, potential amplitude and latency compared with the injury group were significant differences (P<0.05).4. The tissue HE staining showed the fibers of normal facial nerve were lined up with parallel, while those of the injured side group discontinued.ChapterⅡ:1. The whiskers blow of injured side improved significantly over the previous time, slightly worse than the sham side. The whiskers blow of nimodipine pretreatment group improved significantly better than other groups.2. The pathological results of the group of nimodipine pretreatment performed better than the other groups(nimodipine after treatment group, injury group sham operation group).3. Nerve conduction velocity was significantly different (P= 0.000<0.01) between 1-month group and 3-month group; in 6-month group, nerve conduction velocity was significantly different (P= 0.000<0.01) between nimodipine pretreatment group and nimodipine aftertreatment group, nimodipine aftertreatment group and the injury groups was not statistically significant (P= 0.058> 0.05). Action potential amplitude was significantly different (P= 0.000<0.01) between 1-month group and 3-month group; in 6-month group, action potential amplitude was significantly different (P= 0.000<0.01) between nimodipine pretreatment group and nimodipine aftertreatment group, it was not statistically significant (P = 0.141> 0.05) between nimodipine after treatment group and damage groups.Nerve action potential latency was significantly different (P= 0.000 <0.01) between 1-month group and 3-month group; in 6-month group, nerve action potential latency was significantly different (P= 0.012<0.01) between nimodipine pretreatment group and nimodipine aftertreatment group, it was not statistically significant (P= 0.340> 0.05) between nimodipine after treatment group and damage groups.4. Statistics software SPSS 13.0 was used, Image processing software Image Tool 3.0 was used to measure electrophoresis band density values.we can see protein bands of GDNF in experiment group, nimodipine pretreatment group was higher than post-treatment group and the injury group, the difference was statistically significant (p<0.01), the expression of GDNF in group of nimodipine aftertreatment was higher than the injury group in 1-month and in 3 month, the difference was statistically significant (p<0.05), there was no significant difference between groups (p= 0.095> 0.05) in 6 month; the expression were statistically significant (p<0.05) in pretreatment and aftertreatment group in each period. The expression of BDNF protein bands can be seen in nimodipine treatment groups and injury group, nimodipine pretreatment group was significantly higher than aftertreatment group and the injury group, the difference was statistically significant (p<0.01), the expression of BDNF in the treated group was higher than injury group in 1-month group, the difference was statistically significant (p<0.05), there was no significant difference in 3-month group (p= 0.087> 0.05) and 6-month group (p= 0.180> 0.05); the expression were statistically significant (p<0.05) in pretreatment and aftertreatment group in each period.ChapterⅢ1. There were 32 cases of vestibular schwannoma, facial nerve was kept anatomic intact in 32 cases of the patients(100%). the mortality rate was 0%.2. In cerebellopontine angle, the facial nerve was found on the anterior superior third in 6(18.8%),the anterior middle portion of the tumor in 11 cases (34.4%),the anterior inferior third in 14 (43.7%),the posterior location was rare and was found in only 1 cases (3.1%). On the medial side, the facial nerve is located on top of the glossopharyngeal nerve and in the anterior-superior of the flocculus and the choroid plexus.3.3 patients had extradural hematoma or hematoma in operative areas which required reoperation. Others complications such as:intracranial infection about 3 cases (9.4%), pulmonary infection about 3 cases (9.4%), hoarseness about 2 cases (6.4%), dysphagia about 2 cases (6.4%) and cerebrospinal fluid leakage about 1 cases (3.2%). Intracranial infections and pulmonary infections were cured by the way of standard antibiotic treatment, injecting antibiotics into intrathecal or lumbar drainage. Subcutaneous fluid was gradually absorbed through the pressure bandage. Cerebrospinal fluid leakage could be resolved by continuous lumbar drainage.4. The function of facial nerve in pretreatment of nimodipine after 1 week:1 case of gradeⅠ,1 cases of gradeⅡ,3 cases of gradeⅢ, the rate of good recovery was 40%; the function of facial nerve in pretreatment of nimodipine in group of 3-month:one case of gradeⅠ,3 cases of gradeⅡ,the rate of good recovery was 80%. The function of facial nerve in aftertreatment of nimodipine after 1 week: one case of gradeⅡ, the rate of good recovery of 16.7%; in 3-month group:1 case of gradeⅠ,2 cases of gradeⅡ, the rate of good recovery of 50%. Untreated group after 1 week:1 case of gradeⅡ, good facial nerve function recovery rate 4.8%, in 3-month group:in 3 cases of gradeⅠ,6 cases of gradeⅡ, the rate of good recovery was 43%.Conclusion:1. The established rats facial nerve injury model is stable and uniform, conveniently manufactured and easily repeatitive.With own control, the error made by factitiousness was enormously reduced. The evaluation method is effective and feasible, which provide a proper animal model for revealing facial protection mechanism of facial paralysis.2. Nimodipine prophylaxis of the rat facial nerve injury is better than nimodipine aftertreatment group and control group. 3. Nimodipine prophylaxis can improve facial nerve conduction velocity, action potential amplitude of the facial nerve, reducing latency, accelerating the repair of facial nerve functions.3. Nimodipine prophylaxis of facial nerve can enhance the expression of GDNF and BDNF.4. Prophylactic nimodipine protective effect of facial nerve may achieve by improving the expression of GDNF and BDNF.5. Preliminary clinical observation showed prophylactic use of nimodipine in the treatment of facial nerve injury increased rate of postoperative facial nerve recovered well.