Reinebrant Hanna E, Wixey Julie A, Buller Kathryn M
UQ Centre for Clinical Research, Faculty of Medicine, The University of Queensland, Herston, Queensland, Australia.
Neural Regen Res. 2020 Mar;15(3):457-463. doi: 10.4103/1673-5374.266067.
Neonatal hypoxia-ischemia (HI) results in losses of serotonergic neurons in specific dorsal raphé nuclei. However, not all serotonergic raphé neurons are lost and it is therefore important to assess the function of remaining neurons in order to understand their potential to contribute to neurological disorders in the HI-affected neonate. The main objective of this study was to determine how serotonergic neurons, remaining in the dorsal raphé nuclei after neonatal HI, respond to an external stimulus (restraint stress). On postnatal day 3 (P3), male rat pups were randomly allocated to one of the following groups: (i) control + no restraint (n = 5), (ii) control + restraint (n = 6), (iii) P3 HI + no restraint (n = 5) or (iv) P3 HI + restraint (n = 7). In the two HI groups, rat pups underwent surgery to ligate the common carotid artery and were then exposed to 6% O for 30 minutes. Six weeks after P3 HI, on P45, rats were subjected to restraint stress for 30 minutes. Using dual immunolabeling for Fos protein, a marker for neuronal activity, and serotonin (5-hydroxytrypamine; 5-HT), numbers of Fos-positive 5-HT neurons were determined in five dorsal raphé nuclei. We found that restraint stress alone increased numbers of Fos-positive 5-HT neurons in all five dorsal raphé nuclei compared to control animals. However, following P3 HI, the number of stress-induced Fos-positive 5-HT neurons was decreased significantly in the dorsal raphé ventrolateral, interfascicular and ventral nuclei compared with control animals exposed to restraint stress. In contrast, numbers of stress-induced Fos-positive 5-HT neurons in the dorsal raphé dorsal and caudal nuclei were not affected by P3 HI. These data indicate that not only are dorsal raphé serotonergic neurons lost after neonatal HI, but also remaining dorsal raphé serotonergic neurons have reduced differential functional viability in response to an external stimulus. Procedures were approved by the University of Queensland Animal Ethics Committee (UQCCR958/08/NHMRC) on February 27, 2009.
新生儿缺氧缺血（HI）会导致特定背侧中缝核中的5-羟色胺能神经元缺失。然而，并非所有的中缝5-羟色胺能神经元都会丢失，因此评估剩余神经元的功能对于理解它们在受HI影响的新生儿中导致神经障碍的可能性非常重要。本研究的主要目的是确定新生儿HI后仍存在于背侧中缝核中的5-羟色胺能神经元如何对外界刺激（束缚应激）作出反应。在出生后第3天（P3），将雄性幼鼠随机分为以下几组：（i）对照+无束缚（n = 5），（ii）对照+束缚（n = 6），（iii）P3 HI+无束缚（n = 5）或（iv）P3 HI+束缚（n = 7）。在两个HI组中，幼鼠接受手术结扎颈总动脉，然后暴露于6%的氧气中30分钟。P3 HI后六周，即P45时，对大鼠施加30分钟的束缚应激。使用针对Fos蛋白（一种神经元活动标记物）和5-羟色胺（5-羟色胺；5-HT）的双重免疫标记，在五个背侧中缝核中确定Fos阳性5-HT神经元的数量。我们发现，与对照动物相比，单独的束缚应激会增加所有五个背侧中缝核中Fos阳性5-HT神经元的数量。然而，在P3 HI后，与接受束缚应激的对照动物相比，背侧中缝腹外侧核、束间核和腹核中应激诱导的Fos阳性5-HT神经元数量显著减少。相比之下，背侧中缝背核和尾核中应激诱导的Fos阳性5-HT神经元数量不受P3 HI的影响。这些数据表明，新生儿HI后不仅背侧中缝5-羟色胺能神经元会丢失，而且剩余的背侧中缝5-羟色胺能神经元对外界刺激的反应性差异功能活力也会降低。本实验程序于2009年2月27日获得昆士兰大学动物伦理委员会（UQCCR958/08/NHMRC）批准。
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