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The Department of Biological Sciences at Minnesota State University, Mankato has recently implemented a first year undergraduate research experience (called the Research Immersive Scholastic Experience in Biology program; RISEbio) designed to improve student success and engagement in biology. In this program, first year students exchange introductory biology labs for the RISEbio curriculum, where they learn basic laboratory, analytical and scientific reasoning skills before beginning authentic mentored research projects in their second and third semesters. Students in one of three research tracks examine the neural control of reproductive behavior by examining gene expression in the brain of the seasonally breeding green anole lizard (Anolis carolinensis). Working in groups, students gain experience with bioinformatics by examining preliminary RNA-seq data and selecting a gene of interest. Then, students design and test primers to amplify their gene of interest, followed by isolating RNA from the hypothalamus of breeding and non-breeding lizard brains. Lastly, students utilize their isolated RNA samples and validated primers in quantitative RT-PCR studies to determine if their gene of interest is differentially expressed in the anole brain. Preliminary work has identified melatonin receptor 1A (MTNR1A) as more highly expressed in the breeding compared to non-breeding anole hypothalamus, while corticotropin releasing hormone binding protein (CRHBP) expression does not differ seasonally. Together with other aspects of the RISEbio program, these early research experiences have led to increased student outcomes, including increased academic success and enhanced scientific motivation. 

It has been well supported among mammalian species that the hypothalamicpituitary- gonad (HPG) axis is regulated positively by kisspeptin and negatively by gonadotropin inhibitory hormone (GnIH). Studies with seasonal breeding models have generally shown higher levels of kisspeptin in areas of the hypothalamus associated with reproduction, such as the preoptic area (POA) and arcuate nucleus, during the breeding season. Conversely, when examining models during the non-breeding season, studies have indicated GnIH to be higher in hypothalamic nuclei, such as the POA. However, kisspeptin’s role in regulating reproduction may not be consistent among all vertebrate groups. While kisspeptin has been shown to upregulate reproduction in mammals, this peptide has not been detected in avian species and recent work in 􀃕sh has suggested that kisspeptin may not play a regulatory role in reproduction. Relatively little is known about these peptides in reptiles and the seasonal regulation of kisspeptin and GnIH has not been investigated in this group. Green anole lizards (Anolis carolinensis) have a distinct breeding and nonbreeding season, and during the breeding season, steroid hormone levels in the plasma are elevated, males are more territorial, and display reproductive behaviors at a higher frequency. Previous work in this species has demonstrated that, in non-breeding anoles, kisspeptin-positive neurons were present in the POA and the dorsomedial hypothalamus. It is currently unknown whether kisspeptin expression is altered in breeding lizards and there is no data available on GnIH expression in this species. We hypothesize that there is a seasonal effect on kisspeptins and GnIH in green anole lizards, with kisspeptins more highly expressed in the breeding season, while GnIH is more highly expressed in the non-breeding season. Preliminary data using quantitative PCR has revealed no signi􀃕cant di􀃠erence between seasons in the expression of kiss1, kiss2, and Gnih mRNA from a dissection of the brain that contained the hypothalamus (F ≤ 4.35, p ≥ 0.053, n = 4-6 per group). Although we did not detect a significant difference between seasons, expression in specific regions may differ. Therefore, using fluorescent in situ hybridization, we aim to determine the localization and expression levels of kiss1, kiss2 and Gnih expression throughout the hypothalamus. This study expands upon available data and bridges the evolutionary gaps in the roles of kisspeptin and GnIH in regulating reproduction.