Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Mexico.
Departamento de Farmacología, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Mexico.
Neurobiol Dis. 2015 Feb;74:336-46. doi: 10.1016/j.nbd.2014.12.008. Epub 2014 Dec 14.
In striatonigral projections activation of dopamine D3 receptors (D3Rs) potentiates the stimulation of GABA release and cAMP production caused by activation of dopamine D1 receptors (D1Rs). Cytoplasmic [Ca(2+)] in the terminals controls this response by modulating CaMKII, an enzyme that depresses D3R action. To examine the effects of dopamine deprivation on D3R signaling we investigated their function in striatonigral terminals of hemiparkinsonian rats. Denervation switched the signaling cascade initiated by D3R activation. In the non-lesioned side activation of D3R potentiated the stimulatory effects of D1R activation on cAMP production and K(+)-depolarization induced [(3)H] GABA release. In contrast, in the denervated side the stimulatory effects of both D1R activation and forskolin administration were blocked by D3R activation. In non-lesioned slices, D3R responses were inhibited by the activation of CaMKII produced by K(+)-depolarization (via increased Ca(2+) entry). The CaMKII-induced inhibition was blocked by the selective inhibitor KN-62. In denervated tissues the response to D3R stimulation was not modified either by K(+) depolarization or by blocking CaMKII with KN-62. Immunoblotting studies showed that depolarization-induced CaMKII binding to the D3 receptor and CaMKII phosphorylation were suppressed in denervated tissues. We also determined calmodulin expression with PCR and immunoblot techniques. Both techniques showed that calmodulin expression was depressed in the lesioned side. In sum, our studies show that dopaminergic denervation switches the D3R signaling cascade and depresses CaMKII signaling through a process that appears to involve reduced calmodulin levels. Since calmodulin is a major cytoplasmic Ca(2+) buffer our findings suggest that abnormal Ca(2+) buffering may be an important component of the abnormalities observed during dopaminergic denervation.
在纹状体苍白球投射中，多巴胺 D3 受体 (D3R) 的激活增强了多巴胺 D1 受体 (D1R) 激活引起的 GABA 释放和 cAMP 产生的刺激。末梢细胞质[Ca(2+)]通过调节 CaMKII 来控制这种反应，CaMKII 是一种抑制 D3R 作用的酶。为了研究多巴胺剥夺对 D3R 信号的影响，我们研究了它们在半帕金森大鼠纹状体苍白球末梢中的功能。去神经作用改变了 D3R 激活引发的信号级联。在未受损侧，D3R 的激活增强了 D1R 激活对 cAMP 产生和 K(+)-去极化诱导的[(3)H]GABA 释放的刺激作用。相比之下，在去神经侧，D1R 激活和 forskolin 给药的刺激作用都被 D3R 激活所阻断。在非损伤切片中，D3R 反应被 K(+)-去极化（通过增加 Ca(2+)内流）产生的 CaMKII 的激活所抑制。选择性抑制剂 KN-62 阻断了 CaMKII 诱导的抑制。在去神经组织中，D3R 刺激的反应既不受 K(+)去极化的影响，也不受 KN-62 阻断 CaMKII 的影响。免疫印迹研究表明，去神经组织中，去极化诱导的 CaMKII 与 D3 受体的结合以及 CaMKII 的磷酸化均受到抑制。我们还通过 PCR 和免疫印迹技术测定了钙调蛋白的表达。这两种技术都表明，损伤侧的钙调蛋白表达受到抑制。总之，我们的研究表明，多巴胺能去神经作用改变了 D3R 信号级联，并通过一种似乎涉及降低钙调蛋白水平的过程抑制了 CaMKII 信号。由于钙调蛋白是主要的细胞质 Ca(2+)缓冲剂，我们的发现表明，异常的 Ca(2+)缓冲可能是多巴胺能去神经作用期间观察到的异常的一个重要组成部分。
