Vitamin D₃ (VD₃), an essential nutrient for animals, has been demonstrated to stimulate the uptake of certain amino acids. However, the role of VD₃ in the intestine, the primary site for digestion and absorption of nutrients, remains poorly characterized. Here, the grass carp (Ctenopharyngodon idella) was studied to assess the influence of different doses of VD₃ (15.2, 364.3, 782.5, 1,167.9, 1,573.8, and 1,980.1 IU/kg) on growth performance, intestinal morphology, digestive absorption, amino acid transport, and potential signaling molecule levels in a feeding experiment. As a result, dietary VD₃ improved growth performance, intestinal structure, and digestive and brush border enzyme activities. Additionally, most intestinal free amino acids and their transporters were upregulated after VD₃ intake, except for Ala, Lys, Asp, Leu, solute carrier (SLC) 7A7, SLC1A5, and SLC1A3 mRNA in different segments, Leu and SLC6A14 mRNA in the proximal intestine, and SLC7A5 mRNA in the mid and distal intestine. In the crucial target of rapamycin (TOR) signal pathway of amino acid transport, the gene and protein expression of TOR, S6 kinase 1, and activating transcription factor 4 were elevated, whereas 4E-binding protein 1 was decreased, further suggesting an advanced amino acid absorption capacity in the fish due to VD₃ supplementation. Based on percentage weight gain, feed efficiency, and trypsin activity, the VD₃ requirements of on-growing grass carp were estimated to be 968.33, 1,005.00, and 1,166.67 IU/kg, respectively. Our findings provide novel recommendations for VD₃ supplementation to promote digestion and absorption capacities of fish, contributing to the overall productivity of aquaculture.

维生素 D ₃ (VD ₃ ) 是动物必需的营养素，已被证明可以刺激某些氨基酸的吸收。然而，VD ₃在肠道（消化和吸收营养物质的主要场所）中的作用仍然知之甚少。本研究以草鱼 ( Ctenopharyngodon idella ) 为研究对象，评估不同剂量的 VD ₃ (15.2、364.3、782.5、1,167.9、1,573.8 和 1,980.1 IU/kg) 对生长性能、肠道形态、消化吸收、氨基酸的影响运输和喂养实验中潜在的信号分子水平。结果，膳食 VD ₃改善了生长性能、肠道结构以及消化酶和刷状缘酶活性。此外，摄入VD ₃后，大多数肠道游离氨基酸及其转运蛋白均上调，不同片段中的Ala、Lys、Asp、Leu、溶质载体（ SLC ） 7A7 、 SLC1A5和SLC1A3 mRNA，以及不同片段中的Leu和SLC6A1 4 mRNA除外。近端肠，以及中段和远端肠中的SLC7A5 mRNA。在氨基酸转运的雷帕霉素（TOR）信号通路的关键靶标中，TOR、S6激酶1和激活转录因子4的基因和蛋白表达升高，而4E结合蛋白1降低，进一步提示高级氨基酸由于补充了 VD ₃ ，鱼的酸吸收能力增强。根据增重百分比、饲料效率和胰蛋白酶活性，生长草鱼的 VD ₃需求量估计分别为 968.33、1,005.00 和 1,166.67 IU/kg。 我们的研究结果为补充 VD ₃以促进鱼类的消化和吸收能力提供了新的建议，从而有助于提高水产养殖的整体生产力。