Pituitary adenylate cyclase‐activating polypeptide (PACAP) is an important regulator of the stress response in mammals, influencing both the hypothalamic–pituitary–adrenal (HPA) axis and the sympathetic nervous system (SNS). PACAP has been reported to influence energy homeostasis, including adaptive thermogenesis, an energy burning process in adipose tissue regulated by the SNS in response to cold stress and overfeeding. While research suggests PACAP acts centrally at the level of the hypothalamus, knowledge of PACAP's role within the sympathetic nerves innervating adipose tissues in response to metabolic stressors is limited. This work shows, for the first time, gene expression of PACAP receptors in stellate ganglia and highlights some differential expression with housing temperature. Additionally, we present our dissection protocol, analysis of tyrosine hydroxylase gene expression as a molecular biomarker for catecholamine producing tissue and recommend three stable reference genes for the normalization of quantitative real time‐polymerase chain reaction (qRT‐PCR) data when working with this tissue. This study adds to information about neuropeptide receptor expression in peripheral ganglia of the sympathetic nervous system innervating adipose tissue and provides insight into PACAP's role in the regulation of energy metabolism.

In obesity, energy intake exceeds energy expenditure and concomitantly increases risk of chronic diseases, including metabolic diseases such as diabetes. Therapeutics to correct dysregulations in energy balance are needed and one notable neuropeptide being studied is pituitary adenylate cyclase-activating polypeptide (PACAP), an overarching regulator of the stress response1,2. In the context of metabolism, PACAP has been shown to regulate adaptive thermogenesis, an energy burning process regulated by the sympathetic nervous system (SNS) in response to cold stress and overfeeding. While research suggests PACAP acts centrally at the level of the hypothalamus to regulate energy balance3-5, PACAP is also known to be expressed in the SNS. In preganglionic neurons innervating the adrenal medulla6, PACAP is co-localized with acetylcholine and regulates catecholamine synthesis and release. Recent neuronal tracing studies have shown that postganglionic fibres of the stellate ganglia innervate intrascapular brown adipose tissue (BAT), the main thermogenic tissue in mammals7. However, PACAP's role within the sympathetic nerves innervating and regulating energy metabolism in adipose tissues is not known. As such, our group is interested in characterizing PACAP and PACAP receptor expression in sympathetic nerves innervating and regulating energy metabolism in BAT (via the stellate ganglia) in response to metabolic stressors, such as cold stress. We hypothesize that PACAP is released from preganglionic nerves which binds PACAP receptors (PAC1, VPAC1, VPAC2) expressed in postganglionic nerves of the stellate ganglia. Here we report, for the first time, PAC1 and VPAC1 expression in the stellate ganglia with sex-specific differential gene expression based on housing temperature. Analysis of PAC1 splice variant expression identified at least two variants in tissues of the SNS (adrenal gland, superior cervical ganglia and the stellate ganglia). These results will support future functional studies characterizing PACAP regulation of catecholamine production in peripheral ganglia which will contribute to assessing G-protein coupled receptors (GPCRs) as potential therapeutic targets for obesity and metabolic disease. References 1. Gray SL, Cline DL. 2019. Fink G, Ed. Elsevier: London UK. p.279-91. 2. Mustafa T. 2013. Adv Pharmacol. San Diego, Calif. p.445-57. 3. Banki E, Pakai E, Gaszner B, et al. 2014. J Mol Neurosci 54(3): 543-54. 4. Contreras C, Gonzalez F, Fernø J et al. 2015. Ann Med. 47(2): 150-68. 5. Resch JM, Maunze B, Gerhardt AK, et al. 2013. Am J Physiol-Endoc M. 305(12): E1452-E1463. 6. Eiden LE, Emery AC, Zhang L, et al. 2018. Pflügers Arch. 470(1): 79-88. 7.François M, Torres H, Huesing C, et al. 2019. Ann N T Acad Sci. 1454(1): 3-13. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

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