Expression of the non-neuronal cholinergic system components in Malpighian tubules of Mythimna separata and evidence for non-neuronal acetylcholine synthesis

IF 3.2 2区农林科学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Insect Biochemistry and Molecular Biology Pub Date : 2023-12-09 DOI:10.1016/j.ibmb.2023.104047

Baolei Zhan, Ting Zhou, Di Tong, Zile Xu, Xiaoyu Liang, Anqi Ma, Shumin Lü

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Abstract

The non-neuronal cholinergic system, widely distributed in nature, is an ancient system that has not been well studied in insects. This study aims to investigate the key components of the cholinergic system and to identify the non-neuronal acetylcholine (ACh)-producing cells and the acting sites of ACh in the Malpighian tubules (MTs) of Mythimna separata. We found that non-neuronal ACh in MTs is synthesized by carnitine acetyltransferase (CarAT), rather than choline acetyltransferase (ChAT), as confirmed by using enzyme inhibitors and high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS). Fluorescence in situ hybridization revealed the presence of CarAT mRNA within MTs, specifically localized in the principal cells. Immunohistochemistry showed strong staining for A-mAChR, a muscarinic acetylcholine receptor, in the principal cells. Pharmacological analysis further demonstrated that ACh acts through A-mAChR in the principal cells to increase the intracellular Ca²⁺ concentration. These findings provide compelling evidence for the existence of a non-neuronal cholinergic system in the MTs of M. separata, and the principal cells play a crucial role in ACh synthesis via CarAT.

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Mythimna separata Malpighian 小管中非神经元胆碱能系统成分的表达以及非神经元乙酰胆碱合成的证据

非神经元胆碱能系统广泛分布于自然界中，是一个古老的系统，但在昆虫中却没有得到很好的研究。本研究旨在探究胆碱能系统的关键组成部分，并确定Mythimna separata麦氏管（MTs）中的非神经元乙酰胆碱（ACh）产生细胞和ACh的作用位点。通过使用酶抑制剂和高效液相色谱-质谱联用技术（HPLC-MS/MS），我们发现MTs中的非神经元乙酰胆碱是由肉碱乙酰转移酶（CarAT）而不是胆碱乙酰转移酶（ChAT）合成的。荧光原位杂交显示，MTs 中存在 CarAT mRNA，特别定位于主细胞。免疫组化显示，主细胞中的毒蕈碱乙酰胆碱受体 A-mAChR 染色较强。药理分析进一步证明，ACh 通过主细胞中的 A-mAChR 起到增加细胞内 Ca2+ 浓度的作用。这些发现为M. separata的MT中存在非神经元胆碱能系统提供了令人信服的证据，主细胞通过CarAT在ACh合成过程中发挥了关键作用。

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来源期刊

Insect Biochemistry and Molecular Biology 生物-昆虫学

CiteScore

7.40

自引率

5.30%

发文量

105

审稿时长

40 days

期刊介绍： This international journal publishes original contributions and mini-reviews in the fields of insect biochemistry and insect molecular biology. Main areas of interest are neurochemistry, hormone and pheromone biochemistry, enzymes and metabolism, hormone action and gene regulation, gene characterization and structure, pharmacology, immunology and cell and tissue culture. Papers on the biochemistry and molecular biology of other groups of arthropods are published if of general interest to the readership. Technique papers will be considered for publication if they significantly advance the field of insect biochemistry and molecular biology in the opinion of the Editors and Editorial Board.