Department of Physiology and Pharmacology, Oregon Health and Science University, 3181 SW Sam Jackson Park Road, Portland, OR 97239, USA.
Neuroscience. 2012 Jan 10;201:34-45. doi: 10.1016/j.neuroscience.2011.11.021. Epub 2011 Nov 17.
Cerebellar adaptive plasticity regulates posture and movement in response to changing conditions of sensory stimulation. Study of adaptive plasticity of cerebellar circuitry in vitro confines experimental interest to mechanisms with a time scale of minutes. However, cerebellar plasticity, measured behaviorally or electrophysiologically in vivo, occurs over a time scale of tens of minutes and hours. Here we investigate how optokinetically-evoked increases in climbing fiber activity influence expression of key subcellular signaling proteins that regulate the accumulation of GABA(A) receptors (GABA(A)Rs) in the cytoplasm of Purkinje cells and their insertion into the plasma membrane. We used long-term horizontal optokinetic stimulation (HOKS) to activate climbing fibers that project to the flocculus of mice. Although long-term increases in climbing fiber activity in vivo do not alter the expression of any of the subunits of GABA(A)Rs expressed by Purkinje cells, they do influence other subcellular events such as transcription and interaction of signaling proteins. Specifically, increased climbing fiber activity evoked decreased expression of 14-3-3-θ, reduced serine phosphorylation of GABA(A)g(2), and reduced the interaction of 14-3-3-θ with protein kinase C-γ (PKC-γ). Knockdown of 14-3-3-θ in vivo reduced the serine phosphorylation of GABA(A)γ(2). Conversely, treatment of cerebellar lysates with phorbol 12-myristate-13-acetate (PMA), a PKC activator, increased serine phosphorylation of GABA(A)γ(2). Knockdown of 14-3-3-θ or PKC-γ in N2a cells in vitro reduced serine phosphorylation of GABA(A)γ(2) and reduced its cell-surface expression. We interpret these data to mean that a prolonged increase in climbing fiber activity decreases the cell-surface expression of GABA(A)Rs in Purkinje cells and thereby reduces their sensitivity to GABAergic inhibition. This provides a homeostatic mechanism by which Purkinje cells become less sensitive to stellate cell inhibition also evoked by climbing fiber activity.
小脑的适应性可塑性调节姿势和运动，以响应感觉刺激变化的条件。体外小脑电路的适应性可塑性研究将实验兴趣局限于具有分钟时间尺度的机制。然而，小脑可塑性，无论是在体内通过行为还是电生理学测量，都发生在几十分钟到几个小时的时间尺度上。在这里，我们研究了光运动诱发的 climbing 纤维活动增加如何影响调节 Purkinje 细胞细胞质中 GABA(A)受体 (GABA(A)R) 积累及其插入质膜的关键亚细胞信号蛋白的表达。我们使用长期水平光运动刺激 (HOKS) 来激活投射到小脑绒球的 climbing 纤维。尽管体内 climbing 纤维活性的长期增加不会改变由 Purkinje 细胞表达的任何 GABA(A)R 亚基的表达，但它们确实会影响其他亚细胞事件，如转录和信号蛋白的相互作用。具体而言，增加的 climbing 纤维活性诱发 14-3-3-θ 的表达减少，GABA(A)g(2)的丝氨酸磷酸化减少，并且 14-3-3-θ 与蛋白激酶 C-γ (PKC-γ) 的相互作用减少。体内 14-3-3-θ 的敲低减少了 GABA(A)γ(2)的丝氨酸磷酸化。相反，用佛波醇 12-肉豆蔻酸-13-醋酸盐 (PMA) 处理小脑裂解物，一种 PKC 激活剂，增加了 GABA(A)γ(2)的丝氨酸磷酸化。体外 N2a 细胞中 14-3-3-θ 或 PKC-γ 的敲低减少了 GABA(A)γ(2)的丝氨酸磷酸化，并减少了其细胞表面表达。我们将这些数据解释为意味着 climbing 纤维活性的长期增加降低了 Purkinje 细胞中 GABA(A)R 的细胞表面表达，从而降低了它们对 GABA 抑制的敏感性。这提供了一种平衡机制，通过该机制，Purkinje 细胞对也由 climbing 纤维活性诱发的星状细胞抑制变得不那么敏感。
