Plasticity of the axon initial segment (AIS) has aroused great interest in recent years because it regulates action potential initiation and neuronal excitability. AIS plasticity manifests as modulation of ion channels or variation in AIS structure. However, the mechanisms underlying structural plasticity of the AIS are not well understood. Here, we combined immunofluorescence, patch-clamp recordings, and pharmacological methods in cultured hippocampal neurons to investigate the factors participating in AIS structural plasticity during development. With lowered neuronal density, the distance between the AIS and the soma increased, while neuronal excitability decreased, as shown by the increased action potential threshold and current threshold for firing an action potential. This variation in the location of the AIS was associated with cellular secretory substances, including brain-derived neurotrophic factor (BDNF) and neurotrophin 3 (NT3). Indeed, blocking BDNF and NT3 with TrkB-Fc eliminated the effect of conditioned medium collected from high-density cultures on AIS relocation. Elevating the extracellular concentration of BDNF or NT3 promoted movement of the AIS proximally to the soma and increased neuronal excitability. Furthermore, knockdown of neurotrophin receptors TrkB and TrkC caused distal movement of the AIS. Our results demonstrate that BDNF and NT3 regulate AIS location and neuronal excitability. These regulatory functions of neurotrophic factors provide insight into the molecular mechanisms underlying AIS biology.
A decline in proteasome function is causally connected to neuronal aging and aging-associated neuropathologies. By using hippocampal neurons in culture and in vivo, we show that aging triggers a reduction and a cytoplasm-to-nucleus redistribution of the E3 ubiquitin ligase mahogunin (MGRN1). Proteasome impairment induces MGRN1 monoubiquitination, the key post-translational modification for its nuclear entry. One potential mechanism for MGRN1 monoubiquitination is via progressive deubiquitination at the proteasome of polyubiquitinated MGRN1. Once in the nucleus, MGRN1 potentiates the transcriptional cellular response to proteotoxic stress. Inhibition of MGRN1 impairs ATF3-mediated neuronal responsiveness to proteosomal stress and increases neuronal stress, while increasing MGRN1 ameliorates signs of neuronal aging, including cognitive performance in old animals. Our results imply that, among others, the strength of neuronal survival in a proteasomal deterioration background, like during aging, depends on the fine-tuning of ubiquitination-deubiquitination.
Contributions from estrogen receptor (ER) subtypes (ERα and ERβ) to postpartum anxiogenic and depressive responses remain unresolved in rats. Using the elevated-plus maze (EPM) and forced swim (FS) tests, we confirmed that primiparous rats exhibited anxiogenic and depressive responses 3 weeks postpartum, improved 5 weeks postpartum (EPM), and recovered at 5 (FS) or 10 weeks postpartum (EPM) compared with diestrus nulliparous females. Immunohistochemistry suggested that these behavioral changes were temporally associated with decreased ERα but not ERβ expression in the medial amygdala (MEA). Additionally, ERα expression in the medial preoptic area (MPOA) significantly increased 10 weeks postpartum. Brain-derived neurotrophic factor (BDNF) expression was significantly elevated in the MEA 3 weeks postpartum. BDNF receptor tropomyosin-related kinase expression was significantly elevated in the MEA at 3 and 10 weeks but not at 5 weeks postpartum. The phosphorylation of ERK (pERK)-2 in the MEA, MPOA, and hippocampal CA1 region was significantly elevated 3 and 5 weeks postpartum. The effects of single daily sc injections of the ERα-selective agonist, propyl pyrazoletriol (PPT); ERβ-selective agonist, diarylpropionitrile; 17β-estradiol (E₂); and vehicle for 6 days in primiparous rats were assessed. PPT and E₂ significantly produced anxiolytic and antidepressant actions in the EPM and FS tests but PPT to a lesser degree than E₂ in the EPM test. Diarylpropionitrile affected the EPM test but was not significantly different from vehicle. BDNF expression was significantly increased 3 weeks postpartum by all treatments in the MPOA but not the CA1 and MEA. E₂ and PPT treatment significantly increased tropomyosin-related kinase and pERK1/2 expression in the MEA and MPOA and increased pERK1/2 expression in the CA1. The onset of anxiety- and depression-like behaviors in postpartum rats may be partly caused by a complex estrogen-mediated mechanism; nevertheless, changes in the ERα-related system, likely in the MEA, are predominantly involved.