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Cholecystokinin (CCK) is an incretin-like hormone that is also produced by pancreatic β-cells under conditions of stress and obesity. In the past, we have established a role for CCK in protection from β-cell apoptosis. However, the specific CCK receptor responsible for this protective effect was unknown. Both Cckar and Cckbr mRNA are expressed in mouse islet, although Cckbr expression is very low. Incubation of WT mouse pancreatic islets with CCK-8 (100nM) increases the transcript levels of Cckar but does not impact Cckbr expression. Conversely, 24-hour treatment with proinflammatory cytokine cocktail decreases CCKAR transcript levels but again does not modulate Cckbr expression. Therefore, we hypothesized that the CCKA receptor was the most likely mediator of CCK signaling in the β-cell. We used isolated islets from mice with germline knockout of CCK receptors to determine which receptor was required for the pro-survival effects of CCK on the β-cell. Using imaging flow cytometry of dissociated islet cells stained for annexin V and propidium iodide (PI) we find that islet cells from CCKBR knockout mice have a 19.3% (p<0.005) reduction in cytokine-mediated cell death when treated with CCK-8 (100 nM), similar to the protection seen in wild type islets. This protective effect is lost in the CCKAR and double knockout mouse islets. We also confirm this finding using insulin and TUNEL immunohistochemistry to specifically identify β-cell apoptosis. In CCKBR knockout mouse islets, CCK-8 (100 nM) reduces the % TUNEL positive β-cells from 12.98% to 4.39% (p<0.05) after cytokine exposure, similar to protection seen in WT mouse islets (10.47% to 4.48%). Again, CCK-8 does not protect β-cells from cytokine-induced apoptosis in CCKAR knockout or double CCKR null mouse islets. Taken together, CCKAR, but not CCKBR, is required for CCK-mediated protection of pancreatic β-cells from cytokine-induced stress. Disclosure H. Kim: None. S. Sacotte: None. R.A. Williams: None. A.H. de Souza: None. J. Han: None. J.T. Bartosiak: None. G.H. Yang: None. D.B. Davis: None. Funding National Institutes of Health

Cholecystokinin (CCK) is a gut peptide hormone that is also produced by pancreatic β‐cells under conditions of metabolic stress and obesity. Treatment with exogenous CCK can alleviate obesity‐related diabetes in animal models without significant side effects. We have established that CCK is expressed primarily in the pancreatic β‐cell and that CCK is highly upregulated in ob/ob mice. We have also shown that elevated cellular cAMP stimulates β‐cell CCK expression and secretion. However, the specific islet cell types that express CCK and the regulation of Cck transcription in the islet has not been thoroughly explored. A more recent study from our lab showed that high glucose concentrations can upregulate Cck expression in mouse islets and human islets. We hypothesize that Cck expression is upregulated in response to various diabetogenic conditions and that CCK upregulation by glucose can occur independent of cAMP activation. Islets from 16‐week‐old C57BL/6J mice were harvested for use in a dose‐dependent 24‐hour study using different glucose concentrations: No glucose, 2.5mM, 7mM, 11mM & 25mM. After 24‐hour treatment, islets were collected for RNA isolation and cDNA synthesis to identify changes in Cck mRNA expression after incubation with different glucose concentrations. Cck mRNA levels were elevated 4‐8 fold in islets treated with 25 mM glucose compared to islets treated with 0 to 11mM glucose. Notably, the hypoglycemic treatment of no glucose and 2.5mM showed a 4‐fold increase when compared to the 7mM and 11mM treatments. Following this experiment, we wanted to determine if high glucose would still increase Cck mRNA expression after inhibiting cAMP. Islets were pre‐incubated for 45 minutes with 10nM sulprostone, which activated prostaglandin E3 (EP3) receptor and inhibits adenylate cyclase. After inhibition of cAMP, we incubated the islets at 2.5mM and 25mM glucose concentrations like the prior experiment and observed changes in Cck mRNA expression. After treatment with sulprostone, we are still able to see a significant increase in Cck mRNA expression in islets treated with 25mM glucose compared to those treated with 2.5mM glucose, even with cAMP inhibition. This data suggests that there are alternative pathways that could lead to CCK upregulation in the pancreatic islet independent of cAMP. It also shows that hypoglycemia may also increase Cckexpression, which suggests that CCK is upregulated in response to several forms of metabolic stress. We can further explore the effectiveness of glucose to increase CCK expression by treating islets with cycloheximide and deoxyglucose to determine if glucose metabolism is required for upregulation of Cck.This information will help with our general understanding of how CCK functions in pancreatic islet and how Cck transcription is regulated through signaling pathways and transcriptional networks that regulate β‐cell function and identity under diabetogenic conditions.