Role of Kalirin 7 in Estrogen-Mediated Spine Plasticity in the Hippocampus

描述（由申请人提供）：雌激素在体内和体外均可引起海马神经元棘和兴奋性突触的形成，但其潜在机制尚不完全清楚。Kalirin (Kal) 7仅定位于海马神经元兴奋性突触的突触后侧。在体外实验中，外源性Kal7的表达增加了海马神经元的脊柱密度，而内源性Kal7的表达减少了海马神经元的脊柱密度。Kal7基因敲除（Kal7KO）小鼠CA1神经元的突触结构和功能需要Kal7。我的初步数据表明，体内和体外海马神经元的Kal7免疫反应性受雌激素的调节；Western blot证实了这一结果。当内源性Kal7表达降低时，雌激素不再能够增加突触的形成。这些数据导致了Kal7在雌激素调节突触可塑性的机制中起关键作用的假设。Aim1。确定雌激素如何增加海马中Kal7的表达。雌激素可能直接作用于锥体神经元和/或通过中间神经元增加CA1锥体神经元的棘/突触密度。目前尚不清楚雌激素是在转录水平（mRNA）还是转录后水平调控Kal7的表达。目的1是确定：(1)雌激素影响Kal7 mRNA水平的时间过程；(2)在发情周期中Kal7表达是否发生变化；(3)内源性雌激素是否影响Kal7的表达；(4)哪个雌激素受体（1或2）在这一过程中起关键作用。目标2。确定Kal7是否对雌激素介导的脊柱形成和海马神经元突触功能至关重要。了解雌激素调控Kal7表达的机制将有助于调控雌激素介导的突触可塑性。如果Kal7对于雌激素介导的脊柱形成和突触功能至关重要，那么与Ovx野生型对照相比，卵巢切除（Ovx） Kal7KO小鼠中雌激素替代介导的脊柱形成和突触功能可能减弱或消失。目的2是确定：(1)Kal7是否在体外雌激素介导的突触功能中发挥重要作用；(2)体内雌激素介导的脊柱形成是否需要Kal7；(3) Kal7是否对雌激素介导的LTP诱导至关重要；(4)雌激素调控Kal7表达的途径。

DESCRIPTION (provided by applicant): Estrogen causes the formation of spines and excitatory synapses in hippocampal neurons in vitro and in vivo, but the underlying mechanisms are not fully understood. Kalirin (Kal) 7 is exclusively localized to the postsynaptic side of excitatory synapses in hippocampal neurons. Expression of exogenous Kal7 increases spine density whereas reduced endogenous Kal7 decreases spine density in hippocampal neurons in vitro. Kal7 is required for synaptic structure and function in CA1 neurons in Kal7 knockout (Kal7KO) mice. My preliminary data show that Kal7 immunoreactivity in hippocampal neurons is regulated by estrogen in vivo and in vitro; this result was confirmed by Western blot. Estrogen was no longer able to increase synapse formation when endogenous Kal7 expression was reduced. These data led to the hypothesis that Kal7 plays a key role in the mechanisms by which estrogen regulates synaptic plasticity. Aim1. To determine how estrogen increases Kal7 expression in the hippocampus. Estrogen may act directly on pyramidal neurons and/or through interneurons to increase spine/synaptic density in CA1 pyramidal neurons. It is not yet clear whether estrogen regulates Kal7 expression at the transcriptional (mRNA) or post-transcriptional level. Aim1 is to determine: (1) the time course over which estrogen affects Kal7 mRNA levels; (2) whether Kal7 expression changes during the estrous cycle; (3) whether endogenous estrogen affects Kal7 expression; (4) which estrogen receptor (1 or 2) plays a key role in this process. Aim 2. To determine whether Kal7 is essential for estrogen-mediated spine formation and synaptic functions in hippocampal neurons. Understanding the mechanisms through which estrogen regulates Kal7 expression will facilitate manipulation of estrogen-mediated synaptic plasticity. If Kal7 is essential for estrogen-mediated spine formation and synaptic functions, estrogen replacement-mediated spine formation and synaptic functions in ovariectomized (Ovx) Kal7KO mice are likely attenuated or abolished compared to OVX wildtype controls. Aim 2 is to determine: (1) whether Kal7 plays an essential role in estrogen-mediated synaptic function in vitro; (2) whether Kal7 is essential for estrogen-mediated spine formation in vivo; (3) whether Kal7 is essential for estrogen-mediated LTP induction; (4) the pathway through which estrogen regulates Kal7 expression. PUBLIC HEALTH RELEVANCE: Estrogen causes the formation of dendritic spines and excitatory synapses in hippocampal neurons in vitro and in vivo, but the underlying mechanisms are not fully understood. The aim of this research is to determine the effects of Kal7 on estrogen-mediated spine plasticity in hippocampal neurons.