Functional Enlarged Pituitary Stalk Plays A Role In Central Diabetes Insipidus After Pituitary Stalk Lesion In Rats

Hypothalamo-neurohypophyseal system(HNS)is the most important functional structure for the regulation of body water and electrolyte balance in human and mammals.Hypothalamo-neurohypophyseal system consists of hypothalamo-neurohypophyseal tract(HNT)and hypophyseal portal system.Hypothalamo-neurohypophyseal tract is formed by the axons of the magnocellular neurons in hypothalamic suparoptic and paraventricular nuclei.These axons pass through the median eminence(ME)and the pituitary stalk and finally project into the posterior pituitary,where the neurohypophysial hormones(oxytocin and vaspressin)are released into the blood through hypophyseal portal system.AVP and OT can regulate the reabsorption of water in renal distal convoluted tubule and the metabolism of water and electrolytes.Injury of hypothalamo-neurohypophyseal system by trauma,tumor and operation in neurosurgery could make the insufficient of AVP synthesis or releasing,which can cause central diabetes insipidus(CDI).It has been proved that CDI can occur after hypothalamo-neurohypophyseal system injury in rats.There are several research methods to induce CDI in animal experiments,such as hypophysectormy,damage of the nuclei and axotomy or compression of pituitary stalk,and each method has advantages and disadvantages.In our previous study,we have demonstrated that different CDI characteristics and prognosis are shown in rats that underwent pituitary stalk electric lesion(PEL),and an enlarged stalk was formed at the lesion site in CDI recovery rats after PEL by histological research.Based on the former studies and our research foundations,we built a stable and reproducible stereotactic pituitary stalk lesion model in rats,and described the CDI pattern to explore the role of the enlarged pituitary stalk in CDI recovery after pituitary stalk lesion in rats and pathophysiological mechanism.Chapter 1.The pituitary stalk lesion rat model and the CDI characteristicsMethods:The lesion knife was designed using AutoCAD software,and PEL was performed by parietal approach.Then the biological parameters of CDI were monitored daily for 28 days post-operation.Brains magnetic resonance imaging(MRI)of PEL rats were captured,and rat brains were obtained via the retaining sphenoid and dura technique.Finally,the brain sections were used for immunohistochemistry and immunofluorescence studies.Results:A triphasic CDI pattern was observed in the PEL rats,and the PEL method for CDI model is efficient and stable and repeatable.Compared to the sham-operated rats,the MRI signals of pituitary stalk was interrupted in PEL rats.Through the retaining sphenoid and dura technique,the sagittal paraffin tissue sections of hypothalamo-neurohypophyseal tract can be obtained and applied for AVP immunohistochemical staining.Moreover,the AVP immunohistochemistry studies demonstrated that no AVP-immunoreactivities were detected in the atrophic posterior pituitary and abundant AVP staining were detected in the enlarged proximal end of pituitary stalk on 28 days after PEL operation.Furthermore,compared to the sham-operated rats,AVP Immunofluorescent staining of hypothalamic region showed that the number of AVP positive magnocellular neurons were reduced in both SON and PVN in PEL rats.Chapter 2.Functional enlarged pituitary stalk plays a role in central diabetes insipidus after pituitary stalk lesion in rats and the physiopathological mechanismMethods:PEL was performed by parietal approach and then the biological parameters of CDI were monitored daily.On 28th day post-operation,the blood samples of rats were collected from the retro-orbital sinus and the brains were obtained via the retaining sphenoid and dura technique.The serum AVP levels were quantified using an ELISA kit and brain tissues were used for immunohistochemistry studies or Western Blot.Results:Two distinct CDI biological characteristics appeared in rats subjected to PEL,and PEL rats were classified as the recovery group and the no-recovery group.Compared to the sham-operated rats,the serum AVP concentrations in the recovery group increased significantly,but the difference in serum AVP content between the sham and no-recovery group was not statistically significant.Significant reduction was observed in the number of AVP neurons in both the recovery group and the no-recovery group compared with the sham-operated group,and abundant AVP staining were detected in the enlarged pituitary stalk of recovery group compared to both the sham-operated and no-recovery groups.Moreover,a significant difference in CD34-positive cells was observed between the median eminence-pituitary stalk of recovery and no-recovery groups,and a darker GFAP staining was observed in the no-recovery group than in the sham and recovery groups in median eminence and pituitary stalk.Furthermore,GAP-43 expression was detected by western blot and Immunostaining at 4 weeks after surgery and was significantly increased in the recovery group compared to both the sham-operated and no-recovery groups.Finally,p-PI3K and p-AKT expression was significantly increased in the recovery group but not in the no-recovery and the sham-operated groups,and no significant difference was observed between the no-recovery and sham-operated groups.In conclusion,the results of this study indicated that an enlarged structure was formed as a functional ectopic neural lobe after pituitary stalk lesion.Its regeneration might help the recovery from CDI and might be due to an increase in GAP-43 expression through the PI3K/AKT signaling pathway.