Neurochemical and behaviour changes in animal models for studing different aspects of Alzheimer's Disease: effects of chronic treatment of standardized extract of Ginkgo biloba (EGb)

Abstract

Alzheimer's disease (AD) is the most frequent cause of dementia associated with aging, occurring with higher prevalence in women. Among the factors associated with the disease are the alteration in the function of acetylcholine vesicular transporter (VAChT) in the prefrontal cortex (CPF) and in the hippocampal formation (FH) of patients with AD. Another risk factor associated with AD is impaired insulin signaling in these same brain areas. These changes result in changes in expression CREB-1, NF-kB and BDNF, cytokines such as GSK3-²; PI3K / Akt, IL-1², IL-10 and ERK1 / 2 and Cholinergic Receptors (N7 and M1); 5HT1A and GABAA in neurons and glia. To better understand these issues, this study proposes to investigate the effects of reduced expression of VAChT (Knockdown, KD) in mice or of insulin resistance induced by intracerebroventricular (icv) injection of streptozotocin (STZ) in memory formation, short and long term, and if chronic treatment with standardized Ginkgo biloba extract (EGb) can reduce cognitive impairment of AD and modulate cellular mechanisms. Additionally, we will evaluate the same parameters in aging. In experiment I, female C57BL / 6 (N3) mice wild type (WT) or 45% (VAChT KDHet) or 65% (VAChT KDHom) strains will be used. The animals will be divided into 2 groups [adults (3- 6 months) and elderly (24 months)] Each group will be redistributed into 18 subgroups: i-iii) naive animals (KDHet, KDHom and WT; n = 8 / group); iv-vi) vehicle groups (0.9% saline) (KDHet, KDHom and WT; n = 8 / group); vii) donepezil hydrochloride groups 5 mg / kg (DPZ) (KDHet, KDHom and WT; n = 8 / group); x-xviii) EGb groups at doses 250, 500 or 1000 mg / kg (KDHet, KDHom and WT; n = 8 / group). All animals except naives will be treated by gavage (v.o) with EGb, DPZ or vehicle for 31 days. After the 24th treatment will be subjected to acquisition and retention test aversive memory evaluated in discriminative avoidance in high cross maze (PM-DAT) where will be evaluated behaviors related to anxiety, spontaneous motricity and short-term memory and long term. On day 26 you will be exposed to the object recognition task to evaluate item memory (MRO) and object location (MLO). In the 31st will be anesthetized and perfused with 4% formaldehyde or beheaded for removal of FH and CPF, which will be kept in -80º freezer for analysis of differential expression of proteins highlighted above by western blot and / or immunohistochemistry. In experiment II, we aimed to assess whether brain insulin resistance induces molecular and behavioral changes proposed above and whether these differ with animal age and / or are modulated by EGb. Adult Wistar rats 3 to 6 months and 12 months will be used (CEMIB, UNICAMP). Except for the naive group, all groups will undergo stereotactic surgery for icv injection of STZ (3 mg / kg) or vehicle (citrate buffer, CT). 72 hours after surgery the animals will be treated by EGb, 0.9% saline or DPZ for 14 days. The animals will be divided into groups, namely: controls [TC icv + vehicle, EGb (250, 500 or 1,000 mg / kg) or DPZ 5 mg / kg] or submitted to STZ infusion (SZT icv + vehicle, EGb (250 500 or 1,000 mg / kg) or DPZ] and will be evaluated in the same tests described in experiment I, MRO-MLO (7 to 11 days) and PM-DAT (13-14 days) of treatment. and / or beheaded for analysis of the differential expression of proteins described in Experiment I. For data analysis we will use the One-way or Two-way ANOVA test, according to the evaluated parameter followed, when necessary, by the post hoc test. (AU)