DESCRIPTION (provided by applicant): Sex differences in cognitive outcome after early childhood anesthesia have been poorly studied and no mechanism for such differences is known. The long-term goal of this laboratory is to improve the safety of anesthesia delivered to children by eliminating lasting cognitive effects from anesthetic-induced changes in the developing brain. The objective of this proposal is to identify specific sex related factors that lead to divergent cognitive outcomes in male and female rodents. The central hypothesis is that sex-specific cognitive outcomes after early anesthesia are due to testosterone-mediated delay of KCC2 expression which slows the transition from excitatory to inhibitory GABAergic signaling in the hippocampus of male rats leading to loss of neuronal spines and eventual cognitive dysfunction. This hypothesis will be tested in two specific aims: 1) Establish the role of testosterone in dendritic spinogenesis and cognitive outcome after neonatal anesthesia in rats. 2) Establish the role of testosterone in KCC2 expression and determine whether excitatory GABAergic neurotransmission is necessary for anesthetic-mediated spine loss and cognitive deficits. The first aim will compare dendritic spine densities of hippocampal pyramidal neurons and cognitive function following anesthesia exposure in male and female rats after gonadectomy and/or treatment with exogenous testosterone. Under the second aim, section 2.1, will define the role of testosterone in sex-specific temporal expression of KCC2 in control, male and female rats after gonadectomy or treatment with exogenous testosterone. And section 2.2, will use in utero electroporation, to genetically manipulate hippocampal excitatory signaling by early expression of KCC2 or the inward rectifying Kir2.1 (two mechanisms that inhibit GABAergic depolarization) then assess dendritic spine morphology and density as well as cognitive function after early anesthesia exposure. The approach is innovative because cellular level mechanistic studies are guided by and interpreted in light of cognitive function testing, the most relevant outcome that can be measured in animals. The expected contribution from these studies is a detailed understanding of the sex-specific factors leading to deficits of recognition memory in males' vs females after early anesthesia exposure. The proposed research is significant because 1)~ T of children anesthetized in the first three years of life are male, and 2)few other studies have investigated the role of sex in cognitive outcome after early anesthesia, and 3)there is no known mechanism for sex-specific cognitive outcomes after early childhood anesthesia exposure. The results obtained from the proposed research will lead to eventual trials to improve risk stratification and guide decision making around the youngest patients that must be exposed to anesthesia.

PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because more than a million children under age five are anesthetized in the United States every year, but the potential risk factors for cognitive dysfunction after anesthesia are not well understood. This project is also relevant to the NIH mission. The primary goal of the NIH Strategic Plan for Women's Health Research published in 2010 is to "increase sex differences research in basic science studies". The proposed experiments will examine possible mechanisms that lead to sex difference in cognitive outcome after early anesthesia exposure.