Deletion or mutations of SHANK3 lead to Phelan–McDermid syndrome and monogenic forms of autism spectrum disorder (ASD). SHANK3 encodes its eponymous scaffolding protein at excitatory glutamatergic synapses. Altered morphology of dendrites and spines in the hippocampus, cerebellum, and striatum have been associated with behavioral impairments in Shank3-deficient animal models. Given the attentional deficit in these animals, our study explored whether deficiency of Shank3 in a rat model alters neuron morphology and synaptic ultrastructure in the medial prefrontal cortex (mPFC). We assessed dendrite and spine morphology and spine density in mPFC layer III neurons in Shank3-homozygous knockout (Shank3-KO), heterozygous (Shank3-Het), and wild-type (WT) rats. We used electron microscopy to determine the density of asymmetric synapses in mPFC layer III excitatory neurons in these rats. We measured postsynaptic density (PSD) length, PSD area, and head diameter (HD) of spines at these synapses. Basal dendritic morphology was similar among the three genotypes. Spine density and morphology were comparable, but more thin and mushroom spines had larger head volumes in Shank3-Het compared to WT and Shank3-KO. All three groups had comparable synapse density and PSD length. Spine HD of total and non-perforated synapses in Shank3-Het rats, but not Shank3-KO rats, was significantly larger than in WT rats. The total and non-perforated PSD area was significantly larger in Shank3-Het rats compared to Shank3-KO rats. These findings represent preliminary evidence for synaptic ultrastructural alterations in the mPFC of rats that lack one copy of Shank3 and mimic the heterozygous loss of SHANK3 in Phelan–McDermid syndrome. The Shank3 deletion in the rat model we used does not affect all isoforms of the protein and would only model the effect of mutations resulting in loss of the N-terminus of the protein. Given the higher prevalence of ASD in males, the ultrastructural study focused only on synaptic structure in male Shank3-deficient rats. We observed increased HD and PSD area in Shank3-Het rats. These observations suggest the occurrence of altered synaptic ultrastructure in this animal model, further pointing to a key role of defective expression of the Shank3 protein in ASD and Phelan–McDermid syndrome.

中文翻译：

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Shank3缺陷大鼠前额叶皮层突触超微结构改变

SHANK3 的缺失或突变导致 Phelan-McDermid 综合征和单基因形式的自闭症谱系障碍 (ASD)。SHANK3 在兴奋性谷氨酸能突触处编码其同名支架蛋白。在缺乏 Shank3 的动物模型中，海马、小脑和纹状体中树突和棘的形态改变与行为障碍有关。鉴于这些动物的注意力缺陷，我们的研究探讨了大鼠模型中 Shank3 的缺乏是否会改变内侧前额叶皮层 (mPFC) 的神经元形态和突触超微结构。我们评估了 Shank3 纯合敲除 (Shank3-KO)、杂合 (Shank3-Het) 和野生型 (WT) 大鼠中 mPFC III 层神经元的树突和脊柱形态以及脊柱密度。我们使用电子显微镜来确定这些大鼠 mPFC 第三层兴奋性神经元中不对称突触的密度。我们测量了这些突触处棘突的突触后密度 (PSD) 长度、PSD 面积和头部直径 (HD)。三种基因型的基底树突形态相似。与 WT 和 Shank3-KO 相比，Shank3-Het 中的脊椎密度和形态相似，但更薄的蘑菇刺具有更大的头部体积。所有三组都具有可比的突触密度和 PSD 长度。Shank3-Het 大鼠（但不是 Shank3-KO 大鼠）的总和非穿孔突触的脊柱 HD 显着大于 WT 大鼠。与 Shank3-KO 大鼠相比，Shank3-Het 大鼠的总和非穿孔 PSD 面积显着更大。这些发现代表了缺乏一个 Shank3 拷贝并模拟 Phelan-McDermid 综合征中 SHANK3 杂合缺失的大鼠 mPFC 突触超微结构改变的初步证据。我们使用的大鼠模型中的 Shank3 缺失不会影响蛋白质的所有同种型，只会模拟导致蛋白质 N 端丢失的突变的影响。鉴于雄性 ASD 的患病率较高，超微结构研究仅关注雄性 Shank3 缺陷大鼠的突触结构。我们观察到 Shank3-Het 大鼠的 HD 和 PSD 面积增加。这些观察结果表明该动物模型中突触超微结构发生了改变，进一步指出了 Shank3 蛋白表达缺陷在 ASD 和 Phelan-McDermid 综合征中的关键作用。