Dysmaturation of Somatostatin Interneurons Following Umbilical Cord Occlusion in Preterm Fetal Sheep

Abstract

Introduction Cerebral white matter injury is the most common neuropathology observed in preterm infants. However, there is increasing evidence that gray matter development also contributes to neurodevelopmental abnormalities. Fetal cerebral ischemia can lead to both neuronal and non-neuronal structural-functional abnormalities, but less is known about the specific effects on interneurons. Objective In this study we used a well-established animal model of fetal asphyxia in preterm fetal sheep to study neuropathological outcome. We used comprehensive stereological methods to investigate the total number of oligodendrocytes, neurons and somatostatin (STT) positive interneurons as well as 3D morphological analysis of STT cells 14 days following umbilical cord occlusion (UCO) in fetal sheep. Materials and Methods Induction of asphyxia was performed by 25 min of complete UCO in five preterm fetal sheep (98–100 days gestational age). Seven, non-occluded twins served as controls. Quantification of the number of neurons (NeuN), STT interneurons and oligodendrocytes (Olig2, CNPase) was performed on fetal brain regions by applying optical fractionator method. A 3D morphological analysis of STT interneurons was performed using IMARIS software. Results The number of Olig2, NeuN, and STT positive cells were reduced in IGWM, caudate and putamen in UCO animals compared to controls. There were also fewer STT interneurons in the ventral part of the hippocampus, the subiculum and the entorhinal cortex in UCO group, while other parts of cortex were virtually unaffected ( p > 0.05). Morphologically, STT positive interneurons showed a markedly immature structure, with shorter dendritic length and fewer dendritic branches in cortex, caudate, putamen, and subiculum in the UCO group compared with control group ( p < 0.05). Conclusion The significant reduction in the total number of neurons and oligodendrocytes in several brain regions confirm previous studies showing susceptibility of both neuronal and non-neuronal cells following fetal asphyxia. However, in the cerebral cortex significant dysmaturation of STT positive neurons occurred in the absence of cell loss. This suggests an abnormal maturation pattern of GABAergic interneurons in the cerebral cortex, which might contribute to neurodevelopmental impairment in preterm infants and could implicate a novel target for neuroprotective therapies.

Connexin Hemichannel Mimetic Peptide Attenuates Cortical Interneuron Loss and Perineuronal Net Disruption Following Cerebral Ischemia in Near-Term Fetal Sheep

“…In the present study, cerebral ischemia in near-term fetal sheep was associated with a pattern of parasagittal cortical damage after 7 days of recovery, which included a marked reduction in the survival of cortical GAD + , parvalbumin + , calretinin + , and calbindin + interneuron populations. These findings are broadly consistent with previous studies showing loss and disrupted development of interneurons in the cerebral cortex and subcortical grey matter following HI in preterm and term fetal sheep [17,[32][33][34][35] and neonatal rodents [36][37][38][39][40][41][42][43] and following ventilatory support in preterm baboons [44]. Although there are no comparable term human data, limited pathology and imaging studies in preterm infants have shown reductions in the numbers and complexity of cortical interneurons, refs [25,45] cortical interneuron migration, [7] interneuron neurogenesis, [46] and cortical GABAergic signaling [4].…”

Regenerative Therapies to Restore Interneuron Disturbances in Experimental Models of Encephalopathy of Prematurity

“…Consistent with previous studies that showed susceptibility of interneurons to perinatal insults in animal models and post-mortem human tissue, we observed differences in interneuron density following induction of EoP in both our models [ 19 , 20 , 21 , 23 , 24 , 25 , 62 ]. However, the previous published studies generally use single-hit (fetal/postnatal inflammation or hypoxia) models, failing to reflect the multifactorial etiology of EoP [ 19 , 23 , 24 , 25 , 26 , 27 ]. Furthermore, it is good to note that different studies have used different markers of subpopulations of interneurons.…”

“…The net absence of GAD67+ abnormalities in our mouse model might be explained by the overshoot of VIP-expressing interneurons, masking the maturation arrest of PV+ cells. Additional evidence for interneuron maturational arrest of interneurons can be found in other preclinical studies [ 19 , 23 ]. In the first postnatal weeks, similar to excitatory neurons, a surplus of interneurons is eliminated via inactivity-dependent programmed cell death, leading to circuit refinement [ 16 ].…”

“…Recent findings in human post-mortem tissue have revealed cortical interneuron deficits after preterm birth [ 19 , 20 , 21 , 22 ]. Aberrations in interneuron development was confirmed in a few, very recent experimental studies, though a large variety of changes in cortical interneuron density and distribution after different preterm-birth related hits were observed [ 19 , 21 , 23 , 24 , 25 , 26 , 27 ]. Apart from the limited amount of preclinical evidence on the role of interneurons in EoP pathophysiology, studies exploring potential therapeutic interventions to restore interneuron deficits induced by EoP are currently lacking.…”

Cortical Gray Matter Injury in Encephalopathy of Prematurity: Link to Neurodevelopmental Disorders

“…In a study of partial uterine artery occlusion, modeling hypoxia-ischemia in the fetal sheep, immediate low level necrotic cell death was found throughout the deep and cortical GM, followed by extensive apoptosis in both the GM and WM at 3 h post-injury ( 131 ). Other studies of in utero hypoxia-ischemia in sheep have shown some increase in pyknotic cells and activated caspase-3 staining from 24 h up to 4 weeks in the caudate nucleus and subplate ( 132 , 133 ), reduced NeuN and somatostatin positive neurons in the caudate and putamen ( 134 ), specific loss of glutamate decarboxylase interneurons (a marker of GABAergic neurons) and their perineuronal nets in the cortex ( 135 ), along with reduced arborization complexity and spine density in both the caudate, hippocampus, and cortex ( 103 , 132 – 134 , 136 ).…”