Changes Of Sinoatrial Node And Ventricular Conexin43 Of Cerebral Ischemia Rats

Objective To investigate the morphological change of sinoatrial node (SAN) and the expression of connexion43 (Cx43) in left ventricle of cerebral ischemia rat from protein and RNA level. Provide reliable theory and data for recognition, prevention and treatment of cerebral-cardiac syndrome (CCS).Methods A total of 72 health male Sprague-Dawley rats, weight 250g～350g, were divided in to control group (n=12) and MCAO model group (MCAO, n=60), MCAO group were subdivided into 3 h,24 h,48 h,72 h, and 1 week group. According to the modified Zea Longa patching method, perform left middle cerebral artery occlusion. The success of rat model identified by the right limb movement disorder 2 hours after surgery. Control group were treated with the same procedure without the left middle cerebral artery occlusion.①Measurement of ECG.②TTC staining of damaged brain areas after MCAO.③HE staining and transmission electron microscope, TEM:SAN tissue were cut from sulcus terminals (front right section of the superior vena cava proximal wall) and performed HE staining and electron microscopy.④Immunofluorescent staining, Cx43:some frozen sections of left ventricle were stained and observe the changes of Cx43.⑤RT-PCR:detect the expression of Cx43 mRNA in each group.⑥Statistical analysis:immunofluorescent results were analyzed by Image J 1.44e, Statistical analysis was performed using analysis of variance and expressed as mean±SD. P<0.05 was considered statistically significant.Results1 ECG analysis Demonstrated that in control group, heart rate changes were significantly increased after surgery (P<0.05), but after 3 h heart rate gradually returned to the preoperative level. Experimental rats after middle cerebral artery occlusion showed significantly increased heart rate, heart rate accelerating was most obvious after 3 h and decreased after 24 h, but still hold significant changes than that before (P<0.05). After surgery the model groups T wave were elevated, and markedly inverted (upside down).2 HE staining All the SAN was located in the heart epicardium with no coverage of myocardium, cells staining plain. There are clear boundaries between sinus node and the surrounding myocardial in the control group, P cells were mainly located in the center with round, oval or irregular shape and smaller size, often gathered in 3 to 5 cells or scattered. T cell had the middle size between the P cells and atrial muscle cells and more peripheral located. Fibroblasts, collagen fibers, atrial muscle cells and varying amounts of adipose tissue can be seen around the sinus area. Blood vessels located in the central SAN. There no obvious abnormalities were found in Experimental group.3 Transmission electron microscope analysis The SAN in control group illustrated P cells were round or polygonal shaped, with smooth membrane, small or large size well-developed dense mitochondria, clear structure, electron-dense Golgi particles, and the complete sarcomeres surrounded nucleuses. Slender T cell is not smooth in membrane, and has more filaments in cytoplasm, clear Z line, oval nucleus. SAN cells of experimental rats showed rough nuclear membrane, unsharp structure, reduced glycogen granules, shrinking mitochondrial, swelling ridge, rough endoplasmic reticulum, expanse Golgi complex, vacuolus intracytoplasmic.4 Immunofluorescent staining:Cx43 The expression of Cx43 in rat ventricular after 3 h in operation group was statistically decreased than that in the control group (P<0.01). After 24 h,48 h,72 h and 1 week group, Cx43 expression in rat ventricular gradually recovered, but still less compared with the control group (P<0.05). After 48 h, the expression of Cx43 in rat ventricular trended to stabilize (P>0.05).5 RT-PCR:The expression of Cx43 mRNA in operation group were lower than that in control group (P<0.05).ConclusionAfter focal cerebral ischemia in rats found no structural cardiac pathological changes of SAN. Focal cerebral ischemia result in decreased ventricular myocardium junction protein Cx43 expression, and then induce cardiac conduction disturbance.