Programming of Embryonic Blood Brain Barrier and Neurovascular Transcriptome by an Anticipatory Acoustic Signal of Heat in the Zebra Finch

Abstract Organisms have evolved mechanisms to adjust to rapid environmental change. A dramatic example is the Australian zebra finch, where incubating parents produce an acoustic signal ("heat call") during extreme heat, triggering adaptive phenotypic plasticity in their offspring growth, thermoregulation, and reproductive success. To elucidate for the first time the molecular mechanisms underlying heat call-induced programming, we hypothesized a prenatal shift in hypothalamic gene expression, given the hypothalamus’s central role in neuroendocrine signaling controlling metabolism and growth. In addition, we tested whether prenatal heat-call exposure induces local changes in the brain, to protect this highly heat-sensitive organ from upcoming heat challenges. We exposed zebra finch embryos to chronic playback of parental heat calls or control calls, then isolated the hypothalamus for RNA sequencing to identify differentially expressed genes and gene regulatory networks. Heat-call exposure elicited modest neuroendocrine gene expression changes, but robust downregulation of genes tied to muscle contraction and cytoskeletal dynamics, with evidence of isoform usage shifts. These changes were prominently localized to hypothalamic neurovascular endothelial, mural, and ependymal cell populations, forming the blood–brain barrier (BBB). Because embryos experienced heat-associated sound, but not heat itself, and changes matched the loosening of the BBB to avoid breakages, these transcriptomic shifts likely represent an anticipatory response to enhance subsequent brain resilience to heat. Our study provides the first genome-wide characterization of embryonic hypothalamic gene expression in a songbird and reveals that prenatal acoustic cues can developmentally program neurovascular systems, expanding current understanding of developmental plasticity under climate change.