Glucagon-like peptide 1 receptor (GLP-1R) imaging with radiolabeled exendin has proven to be a powerful tool to quantify b-cell mass (BCM) in vivo. As GLP-1R expression is thought to be influenced by glycemic control, we examined the effect of blood glucose (BG) levels on GLP-1R–mediated exendin uptake in both murine and human islets and its implications for BCM quantification. Periods of hyperglyce-mia significantly reduced exendin uptake in murine and human islets, which was paralleled by a reduction in GLP-1R expression. Detailed mapping of the tracer uptake and insulin and GLP-1R expression conclusively demonstrated that the observed reduction in tracer uptake directly correlates to GLP-1R expression levels. Importantly, the linear correlation between tracer uptake and b-cell area was maintained in spite of the reduced GLP-1R expression levels. Subsequent normalization of BG levels restored ab-solute tracer uptake and GLP-1R expression in b-cells and the observed loss in islet volume was halted. This manu-script emphasizes the potency of nuclear imaging techni-ques to monitor receptor regulation noninvasively. Our findings have significant implications for clinical practice, indicating that BG levels should be near-normalized for at least 3 weeks prior to GLP-1R agonist treatment or quan-titative radiolabeled exendin imaging for BCM analysis.