Altered ischemic cerebral injury in mice lacking αIE subunit of the voltage—dependent Ca^2＋ channel

Abstract:Abstract: Objective ①To set up a stable and reproducible focal cerebral infarct model in mice;②To examine theinolvement of aIE subunit of voltage-dependent Ca~(2+ )channel in cerebral ischemic injury. Methods Male C57BL/6J Jclmice 8 -12w and F4-F6α_(IE )subunit of Ca~(2+) channel mutant mice were both used in this study. All animals were allowedto freely access to food and water before and after operation. Animals were anesthetized with pentobarbital sodium 60mg/kg,ip. Rectal temperature was continuously monitored before, during and after operation, and maintained at (36.6±

Altered ischemic cerebral injury in mice lacking αIE subunit of the voltage-dependent Ca2+ channel

Objective (1)To set up a stable and reproducible focal cerebral infarct model in mice; (2)To examine the involvement of aIEi subunit of voltage-dependent Ca2 + channel in cerebral ischemic injury. Methods Male C57BL/6J Jcl mice 8 - 12w and F4 - F6aIE subunit of Ca2+ channel mutant mice were both used in this study. All animals were allowed to freely access to food and water before and after operation. Animals were anesthetized with pentobarbital sodium 60mg/ kg,ip. Rectal temperature was continuously monitored before, during and after operation, and maintained at (36.6 + 0.1)C by a autoregulating pad. To produce pilot models, the middle cerebral artery (MCA) was occluded either by surgical ligation or electrical coagulation and in some models the common carotid artery (CCA) was surgically ligated in tandem . In our latter work the MCA was cut off soon after it was ligated or coagulated in order to make sure that the blood flow was occluded completely. The MCA was coagulated or ligated with a bipolar coagulator or microsurgery suture at the site just superior to the rhinal fissure. Twenty-four hours after the operation, the mice were anesthetized and decapitated, then their brains were dissected from the skull and put into cold artificial brain spinal fluid as soon as possible. 1mm thick coronal sections were cut by vibratome and stained with 2% 2, 3, 5-triphenyltetrazolium chloride (TTC) at 37C for 30min. Every section was photographed positively and the whole infarction volume was calculated by summing up the infarction volumes of all sections by NTH Image System. Infarction ratio ( % ) was also calculated by the following formula: (contralateral volume-ipsilateral undamaged volume)/contralateral volume X 100% to eliminate the influence of edema. In brief, the mutant mice were produced with gene targeting technique. F4 - F6 mice were used in this experiment. All offsprings were genotyped by the polymerase chain reaction (PCR) and the genotypes remained unknown to the researcher until the operation were finished and the infarction volumes were measured. Results In sham operation group there was no obvious injury to the brain. Compared with MCA occlusion only, MCA and CCA ligating and occlusion models have got the higher infarct ratio, however MCA coagulating plus CCA ligating and then cutting method could produce the stable infarct model with higher infarct ratio and higher successful rate. Among the different genotypes, the infarct ratio of wild type was the smallest, the heterozygote the medium and the homozygote were the largest. Conclusion (1)MCA coagulating combined with CCA ligating and cutting model is a relatively ideal acute local cerebral infart model in mice. (2)aIE subunit of voltage-dependent Ca2+ channel is involved in cerebral ischemic injury and may play a protective role in cerebral ischemia . (3)am subunit gene knocked out might influence the structure of function of other subunits.