The Roles Of CyclinD1-CDK4 In Synaptic Plasticity During The Postnatal Development And The Lead Induced Neuronal Death

In this paper, using immunofluorescence, western blot, immunohistochemical techniques and electrophysiology techniques of brain slices and cultured hippocampal neurons, we studied the roles of CDK4/cyclinD1 in synaptic pasticity during the postnatal development in mice hippocampus area CA1 and lead evoked cultured hippocampal neuronal death. Moreover, we investigated the possible upstream signal pathway involved.Techniques: field potential recording in brain slices, western blot and immunohistochemistryResults:1) The different roles of cyclinD1-CDK4 in STP and mGluR-LTD during thepostnatal development in mice hippocampus area CA1The expression and translocation of cyclinD1-CDK4 in post-mitotic neurons indicate that they may have supplementary functions in differentiated neurons that might be associated with neuronal plasticity. In the present study, our findings showed that the expression of CDK4 was localized mostly in nuclei and cytoplasm of pyramidal cells of CA1 at postnatal day 10 (P10); whereas at P28 staining of CDK4 could be detected predominantly in the cytoplasm but not nuclei. Basal synaptic transmission was normal in the presence of CDK4 inhibitor. Short-term synaptic plasticity (STP) was impaired in CDK4 inhibitor pre-treated slices both from neonatal (P8-15) and adolescent (P21-35) animals; however there was no significant change in paired-pulse facilitation (PPF) in slices pre-incubated with the CDK4 inhibitor from adolescent animals. By the treatment of CDK4 inhibitor, the induction or the maintenance of Long-term potentiation (LTP) in response to a strong tetanus and NMDA receptor-dependent long-term depression (LTD) were normal in hippocampus. However, long-term depression (LTD) induced either by group I metabotropic glutamate receptors (mGluRs) agonist or by paired-pulse low-frequency stimulation (PP-LFS) was impaired in CDK4 inhibitor pretreated slices both from neonatal and adolescent animals. But the effects of the CDK4 inhibitor at slices from adolescent animals were not as robust as at slices from neonatal animals. Our results indicated that the activation of cyclinD1-CDK4 is required for short-term synaptic plasticity and mGluR-dependent LTD, and suggested that this cyclin-dependent kinase may have different roles during the postnatal development in mice hippocampus area CA1.2) Involvement of cyclinD1/CDK4 and pRb mediated by PI3K/AKT pathway activation in Pb2+-induced neuronal death in cultured hippocampal neuronsThe present study sought to examine the obligate nature of cyclinD1CDK4, phosphorylation of its substrate retinoblastoma protein (pRb) and its select upstream signal phosphoinositide 3-kinase (PI3K)/AKT pathway in the death of primary cultured rat hippocampal neurons evoked by Pb²⁺. Our data showed that lead treatment of primary hippocampal cultures results in dose-dependent cell death. Inhibition of CDK4 prevented Pb2+-induced neuronal death significantly but incomplete. In addition, we demonstrated that the levels of cyclinD1 and pRb/p107 were increased during Pb2+ treatment. These elevated expression persisted up to 48h, returning to control levels after 72h. We also presented pharmacological and morphological evidences that cyclinD1/CDK4 and pRb/p107 were required for such kind of neuronal death. Addition of the PI3K inhibitor LY294002(30μM) or wortmannin (100nM) significantly rescued the cultured hippocampal neurons from death caused by Pb2+. And that Pb2+-elicited phospho-AKT (Ser473) participated in the induction of cyclinD1 and partial pRb/p107 expression. These results provide evidences that cell cycle elements play a required role in death of neurons evoked by Pb²⁺ and suggest that certain signaling elements upstream of cyclinD1/CDK4 are modified and/or required for this form of neuronal death.