The impact of GenAI-assisted instructional design on the teaching ability of pre-service physical education teachers.

The opioid epidemic will be remembered as one of the most shocking public health failures of the early 21 century, leading to the deaths of over 500,000 people in the United States between 1999-2019 []. One major difficulty in combatting the opioid crisis arises from the fact that while opioids have a high propensity for abuse, they are still the single most effective acute analgesic available. Although significant investment into both the addictive and analgesic properties of opioids has increased in recent years, other, potentially life-changing, medical domains of the endogenous opioid system have gone under-appreciated. One example of this is the role endogenous opioids play in affective neuropsychiatric disorders such as depression or post-traumatic stress disorder (PTSD) [-]. Reaching a deeper understanding of how endogenous opioids contribute to behavioral and psychiatric disorders will help uncover the complete pharmacological potential of this system that is integral to brain function. The dorsal raphe nucleus (DRN) is directly ventral to the periaqueductal gray nucleus (PAG) in the dorsal midbrain and is most well-known for its extensive serotonergic projections to the forebrain and its role in mood regulation. Though clearly important, these 5-hydroxytryptophan (5-HT) containing neurons only account for about 30% of all DRN cells, leaving the role of other neurotransmitter systems in this region unresolved []. Many of these non-serotonergic cells are enriched in opioid peptides, including enkephalin [,]. Enkephalin is encoded by the preproenkephalin (Penk) gene and is subsequently cleaved into the functional peptides met-enkephalin and leu-enkephalin [,]. Enkephalin peptides can bind and activate both delta (DOPR) and mu opioid receptors (MOPR), both of which are associated with mood, affect, motivation, and anxiety. [-]. Considering the wide-ranging interconnectedness of the DRN and the importance of endogenous opioids to behavioral tuning, it is likely that enkephalin originating from DRN neurons have a variety of behavioral effects beyond analgesia and reward. Due to technological constraints, studies investigating opioids in DRN have relied on receptor-level pharmacology to infer how endogenous peptides may contribute to behavioral effects [-]. However, whether these or other behavioral effects are based on the endogenous ligand availability has not been directly studied. Here, we investigated how enkephalin peptide from the DRN (DRN) contributes to a selection of behaviors relating to aversion and reward. Through a Cre-dependent CRISPR-Cas9 approach, we specifically knocked down DRN transcription and assessed behavioral differences in Penk-Cre+ mice and their Penk-Cre− controls. Additionally, we mapped the spatial distribution of the opioid system along with other neurotransmitters throughout the DRN through Hiplex in situ hybridization. Using this information, we also evaluated whether the behavioral effects of DRN knockdown were driven by a subpopulation of cells within the DRN.