Foong Jaime Pei Pei, Hirst Caroline S, Hao Marlene M, McKeown Sonja J, Boesmans Werend, Young Heather M, Bornstein Joel C, Vanden Berghe Pieter
Departments of Physiology and.
Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia and.
J Neurosci. 2015 May 6;35(18):7106-15. doi: 10.1523/JNEUROSCI.4175-14.2015.
Acetylcholine-activating pentameric nicotinic receptors (nAChRs) are an essential mode of neurotransmission in the enteric nervous system (ENS). In this study, we examined the functional development of specific nAChR subtypes in myenteric neurons using Wnt1-Cre;R26R-GCaMP3 mice, where all enteric neurons and glia express the genetically encoded calcium indicator, GCaMP3. Transcripts encoding α3, α4, α7, β2, and β4 nAChR subunits were already expressed at low levels in the E11.5 gut and by E14.5 and, thereafter, α3 and β4 transcripts were the most abundant. The effect of specific nAChR subtype antagonists on evoked calcium activity in enteric neurons was investigated at different ages. Blockade of the α3β4 receptors reduced electrically and chemically evoked calcium responses at E12.5, E14.5, and P0. In addition to the α3β4 antagonist, antagonists to α3β2 and α4β2 also significantly reduced responses by P10-11 and in adult preparations. Therefore, there is an increase in the diversity of functional nAChRs during postnatal development. However, an α7 nAChR antagonist had no effect at any age. Furthermore, at E12.5 we found evidence for unconventional receptors that were responsive to the nAChR agonists 1-dimethyl-4-phenylpiperazinium and nicotine, but were insensitive to the general nicotinic blocker, hexamethonium. Migration, differentiation, and neuritogenesis assays did not reveal a role for nAChRs in these processes during embryonic development. In conclusion, there are significant changes in the contribution of different nAChR subunits to synaptic transmission during ENS development, even after birth. This is the first study to investigate the development of cholinergic transmission in the ENS.
乙酰胆碱激活的五聚体烟碱型受体（nAChRs）是肠神经系统（ENS）中神经传递的一种重要方式。在本研究中，我们使用Wnt1-Cre;R26R-GCaMP3小鼠来研究肌间神经元中特定nAChR亚型的功能发育，在这些小鼠中，所有肠神经元和神经胶质细胞都表达基因编码的钙指示剂GCaMP3。编码α3、α4、α7、β2和β4 nAChR亚基的转录本在E11.5肠道中已低水平表达，到E14.5时以及之后，α3和β4转录本最为丰富。研究了特定nAChR亚型拮抗剂在不同年龄对肠神经元诱发钙活性的影响。α3β4受体的阻断在E12.5、E14.5和P0时降低了电和化学诱发的钙反应。除了α3β4拮抗剂外，α3β2和α4β2的拮抗剂在P10 - 11时以及成年制剂中也显著降低了反应。因此，在出生后发育过程中，功能性nAChRs的多样性增加。然而，α7 nAChR拮抗剂在任何年龄都没有作用。此外，在E12.5时，我们发现了非常规受体的证据，这些受体对nAChR激动剂1 - 二甲基 - 4 - 苯基哌嗪和尼古丁有反应，但对一般的烟碱型阻滞剂六甲铵不敏感。迁移、分化和神经突发生测定未揭示nAChRs在胚胎发育过程中的这些过程中起作用。总之，即使在出生后，ENS发育过程中不同nAChR亚基对突触传递的贡献也有显著变化。这是第一项研究ENS中胆碱能传递发育的研究。
