Single cell atlas of domestic pig brain illuminates the conservation and divergence of cell types at spatial and species levels

Abstract

Domestic pig (Sus scrofa domesticus) has drawn much attention from researchers worldwide due to its implications in evolutionary biology, regenerative medicine and agriculture. The brain atlas of Homo sapiens (primate), Mus musculus (rodent), Danio rerio (fish) and Drosophila melanogaster (insect) have been constructed at single cell resolution, however, the cellular compositions of pig brain remain largely unexplored. In this study, we investigated the single-cell transcriptomic profiles of five distinct regions of domestic pig brain, from which we identified 21 clusters corresponding to six major cell types, characterized by unique spectrum of gene expression. By spatial comparison, we identified cell types enriched or depleted in certain brain regions. Inter-species comparison revealed cell-type similarities and divergences in hypothalamus between mouse and pig, providing invaluable resources for the evolutionary exploration of brain functions at single cell level. Besides, our study revealed cell types and molecular pathways closely associated with several diseases (obesity, anorexia, bulimia, epilepsy, intellectual disability, and autism spectrum disorder), bridging the gap between gene mutations and pathological phenotypes, which might be of great use to the development precise therapies against neural system disorders. Taken together, we reported, so far as we know, the first single cell brain atlas of Sus scrofa domesticus, followed by comprehensive comparisons across brain region and species, which could throw light upon future evo-devo, regenerative medicine, and agricultural studies. Introduction:The domestic pig (Sus scrofa domesticus), one of the most important livestocks, shares close interaction relationships with humans during evolution history [1][2][3]. It belongs to the eutherian mammal from cetartiodactyla order, a clade distinct from primates and rodents [4]. Domestic pig has been widely studied because of its significances in evolutionary biology and regenerative medicine [5]. During the past decades, pig is increasingly used in neuroscience researches, due to the much higher correspondence in its brain to human brain in anatomy, physiology, and development than that of commonly used small laboratory animals such as mouse and rat [6,7]. Similar to human brain, pig brain is gyrencephalic and easy to recognize anatomically. For example, the cerebral cortex is clearly differentiated into four lobes including temporal lobe (TL), frontal lobe (FL), parital lobe (PL), and occipital lobe (OL) [8]. Recently, a transgenetic pig Huntington's disease (HD) model was established by a mutant huntingtin knockin, who shows classical HD phenotypes such as behavioral abnormalities, consistent movement and early death [9]. Accumulating genetic manipulation tools have facilitated the applications of pig as an animal model for researches into brain function, particularly studies concerning brain malfunctioning mechanism. Besides, for ethical and economical reasons, pig is a more acceptable lab animal than primat...

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“…The collected tissues were rinsed by 1X PBS, then quick-frozen and stored in liquid nitrogen. The single cell nucleus of each tissue was separated by mechanical extraction method26 . Briefly, the tissues were first thawed, infiltrated by 1X homogenization buffer (containing 30mM CaCl 2 , 18mM Mg(Ac) 2 , 60mM Tris-HCl (pH 7.8), 320mM sucrose, 0.1% NP-40, 0.1mM EDTA and 0.2U/µl RNase inhibitor), then cut into smaller pieces, and the single nucleus was isolated by 2ml Dounce homogenizer set.…”

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