Dilger, R. N. | bioRxiv Science

bioRxiv2021

Epigenetic MRI: Noninvasive Imaging of DNA Methylation in the Brain

Both neuronal and genetic mechanisms regulate brain function. While there are excellent methods to study neuronal activity in vivo, there are no nondestructive methods to measure global gene expression in living brains. Here we present a method, epigenetic magnetic resonance imaging (eMRI), that overcomes this limitation via direct imaging of DNA methylation, a major gene expression regulator. eMRI exploits the methionine metabolic pathways for DNA methylation to label genomic DNA through 13C-enriched diets. A novel 13C magnetic resonance spectroscopic imaging method then maps the spatial distribution of labeled DNA. We validated eMRI using pigs, whose brains have stronger similarity to humans in volume and anatomy than rodents, and confirmed efficient 13C labeling of brain DNA. We also discovered strong regional differences in global DNA methylation. Just as MRI measurements of regional neuronal activity have had a transformational effect on neuroscience, we expect that the eMRI signal as a surrogate for regional gene expression will enable many new investigations into the roles of gene expression in human brain function, behavior, and disease.

bioengineering↗

bioRxiv2021

Pigs as a new behavioral model for studying Pavlovian eyeblink conditioning

IntroPigs have been an increasingly popular preclinical model in nutritional neuroscience, as their anatomy, physiology, and nutrition requirements are highly comparable to those of humans. Eyeblink conditioning is one of the most well-validated behavioral paradigms in neuroscience to study underlying mechanisms of learning and memory formation in the cerebellum. Eyeblink conditioning has been performed in many species but has never been done on young pigs. Therefore, our aim here was to develop and validate an eyeblink conditioning paradigm in young pigs. MethodEighteen intact male pigs were artificially reared from postnatal day 2 to 30. The eyeblink conditioning setup consisted of a sound-damping box with a hammock that pigs were placed in, which allowed the pig to remain comfortable yet maintain a typical range of head motion. In a delay conditioning paradigm, the conditional stimulus (CS) was a 550 ms blue light-emitting diode (LED), the unconditional stimulus (US) was a 50 ms eye air-puff, the CS-US interval was 500 ms. Starting at postnatal day 14, pigs were habituated for five days to the eyeblink conditioning setup, followed by 5 daily sessions of acquisition training (40 paired CS-US trials each day). ResultsThe group-averaged amplitude of eyelid responses gradually increased over the course of the five days of training, indicating that pigs learned to make the association between the LED light CS and the air-puff US. A similar increase was found for the conditioned response (CR) probability: the group-averaged CR probability on session 1 was about 12% and reached a CR probability of 55% on day 5. The latency to CR peak time lacked a temporal preference in the first session, but clearly showed preference from the moment that animals started to show more CRs in session 2 and onwards whereby the eyelid was maximally closed exactly at the moment that the US would be delivered. ConclusionWe concluded that 4-week-old pigs have the capability of performing in a cerebellar classical association learning task, demonstrating for the first time that eyeblink conditioning in young pigs has the potential to be a valuable behavioral tool to measure neurodevelopment.

neuroscience↗